Tooth Movement

Critical Reviews http://cro.sagepub.
com/
in Oral Biology & Medicine
Tooth Movement
Zeev Davidovitch
CROBM 1991 2: 411
DOI: 10.1177/10454411910020040101
The online version of this article can be found at:
http://cro.sagepub.com/content/2/4/411
Published by:
http://www.sagepublications.com
On behalf of:
International and American Associations for Dental Research
Additional services and information for Critical Reviews in Oral Biology & Medicine can be found at:
Email Alerts: http://cro.sagepub.com/cgi/alerts
Subscriptions: http://cro.sagepub.com/subscriptions
Reprints: http://www.sagepub.com/journalsReprints.nav
Permissions: http://www.sagepub.com/journalsPermissions.nav
>> Version of Record - Jan 1, 1991

What is This?
Downloaded from cro.sagepub.com by guest on July 2, 2012 For personal use only. No other uses without permission.
Critical Reviews in Oral Biology and Medicine,
2(4):411-450 (1991)
Tooth Movement
Zeev Davidovitch
Department of Orthodontics, The Ohio State University College of Dentistry, Columbus, Ohio
ABSTRACT: This article reviews the evolution of concepts regarding the biological foundation of force-induced
tooth movement. Nineteenth century hypotheses proposed two mechanisms: application of pressure and tension
to the periodontal ligament (PDL), and bending of the alveolar bone. Histologic investigations in the early and
middle years of the 20th century revealed that both phenomena actually occur concomitantly, and that cells, as
well as extracellular components of the PDL and alveolar bone, participate in the response to applied mechanical
forces, which ultimately results in remodeling activities.
Experiments with isolated cells in culture demonstrated that shape distortion might lead to cellular activation,
either by opening plasma membrane ion channels, or by crystallizing cytoskeletal filaments. Mechanical distortion
of collagenous matrices, mineralized or non-mineralized, may, on the other hand, evoke the development of
bioelectric phenomena (stress-generated potentials and streaming potentials) that are capable of stimulating cells
by altering the electric charge on their membrane or their fluid envelope. In intact animals, mechanical perturbations on the order of about 1 min/d are apparently sufficient to cause profound osteogenic responses, perhaps
due to matrix proteoglycan-related "strain memory".
Enzymatically isolated human PDL cells respond biochemically to mechanical and chemical signals. The
latter include endocrines, autocrines, and paracrines. Histochemical and immunohistochemical studies showed
that during the early places of tooth movement, PDL fluids are shifted, and cells and matrix are distorted.
Vasoactive neurotransmitters are released from periodontal nerve terminals, causing leukocytes to migrate out
of adjacent capillaries. Cytokines and growth factors are secreted by these cells, stimulating PDL cells and
alveolar bone lining cells to remodel their related matrices. This remodeling activity facilitates movement of
teeth into areas in which bone had been resorbed.
This emerging information suggests that in the living mammal, many cell types are involved in the biological
response to applied mechanical stress to teeth, and thereby to bone. Essentially, cells of the nervous, immune,
and endocrine systems become involved in the activation and response of PDL and alveolar bone cells to applied
stresses. This fact implies that research in the area of the biological response to force application to teeth should
be sufficiently broad to include explorations of possible associations between physical, cellular, and molecular
phenomena. The goals of this investigative field should continue to expound on fundamental principles, particularly on extrapolating new findings to the clinical environment, where millions of patients are subjected annually
to applications of mechanical forces to their teeth for long periods of time in an effort to improve their position
in the oral cavity. Recently developed research tools such as cell culture techniques and immunologic probes,
are the best hope for enhancing this development.
KEY WORDS: orthodontic forces, distortion of cells and matrix,
I.
INTRODUCTORY REMARKS
Throughout their natural history, teeth move

and migrate. Prior to their eruption into the oral
cavity, changes in the position of tooth buds occur primarily due to the growth of dental structures, and the concomitant remodeling of neighboring tissues, i.e., alveolar bone, gingiva, and
periodontal ligament (PDL), including the dental
follicle. Following their emergence into the oral
cavity, teeth reach a position in the dental arch,
neurotransmitters, cytokines, synergism.
dictated by the forces of the surrounding muscles

of the tongue, cheeks, and lips, and by contact
with teeth of the opposite jaw. During mastication, teeth can move slightly in the vertical and
horizontal directions, within the constraints of the
soft tissues of the PDL, and the bendability of
the alveolar bone. Despite their large magnitude,
masticatory forces do not alter the position of
teeth, due to their short duration. However, in
the presence of periodontal disease, when paradental tissues are gradually destroyed, teeth can
1045-4411/91/$.50
by CRC Press, Inc.
1991
411
migrate into new positions, where the masticatory and parafunctional forces reach equilibrium.
Often, these new positions are aesthetically
undesirable.
Tooth position may be deemed undesirable
due to functional and aesthetic considerations,
prompting patients to seek orthodontic care. In
its most simplistic translation, "orthodontics"
means straightening teeth. This "straightening",
or movement of teeth into desirable positions, is
accomplished by the application of forces to teeth,
usually of small magnitude (on the order of a few
grams per square centimeter of dental root surface) and long duration (usually about 2 years).
Millions of people are subjected annually to orthodontic treatment worldwide, making this
branch of dental care a widespread and lucrative
specialty. The number of dentists in the U.S. who
limit their practice to orthondontics is presently
around 10,000. However, many general dentists
worldwide provide orthodontic care to their patients, and all, specialists and generalists alike,
base their therapeutic means upon the time-tested
observation that teeth can be forced to move away
from their position in the dental arch to new locations by means of applied mechanical forces.
The following review focuses upon the phenomenon of tooth movement that can be brought
about by the application of continuous mechanical forces to teeth. Excluded from this review
are the phenomena of eruptive, pathological (periodontal disease related), and surgically induced
tooth movements. Specifically, this review discusses biological aspects of force-induced tooth
movement on the tissue, cellular, and molecular
levels.
II. HISTORICAL PATHFINDERS

The first recorded recommendation to use
force for orthodontic reasons was made around
the year 1 A.D. by Celsus, who suggested the
application of finger pressure to teeth for alignment purposes.1 Seventeen centuries later, Fauchard was the first to publish a description and
an illustration of an orthodontic appliance, which
generated forces by using ligatures to tie teeth to
a rigid arch.2 In the 18th century, Hunter3 provided the first biological explanation for ortho-
dontic tooth movement: "To extract an irregular

tooth would answer but little purpose, if no alterations could be made in the situation of the
rest; but we find that the very principle upon
which teeth are made to grow irregularly is capable, if properly directed, of bringing them even
again. This principle is the power which many
parts (especially bones) have of moving out of
the way of mechanical pressure."
Two significant observations were made during the 19th century concerning the biological
nature of orthodontic tooth movement. In 1815,
Delabbare4 remarked that pain and swelling of
paradental tissues occur following the application
of orthodontic forces to teeth. In contemporary
terms, Delabbare introduced the notion that inflammation is an integral part of orthodontic tooth
movement. In 1888, Farrar5 hypothesized that
tooth movement is due, at least in part, to bending
of alveolar bone by applied forces. This notion
was supported by Wolffss6 proposition in 1892
that the internal architecture of bone is dictated
by the mechanical forces that act upon it.
The first report on the histomorphology of
tissues surrounding orthodontically treated teeth
was published by Sandstedt in 1904 to 1905.7T8
That landmark experiment, which was performed
in one dog, concluded that force-induced tissue
changes are limited to the PDL and its alveolar
bone margin. At the end of 3 weeks of treatment,
Sandstedt observed new bone growth in the
stretched PDL, and bone resorption in the area
of PDL compression. Cell death occurred in the
compressed PDL when the applied force was excessive, and the alveolar bone resorbed as a result
of osteoclastic activity in adjacent marrow spaces
(undermining resorption).
Six years later, Oppenheim9 reported on a
histologic examination of the jaws of one juvenile
baboon whose teeth had been treated by orthodontic forces for 40 d. In contrast to Sandstedt,
Oppenheim saw no demarcation between the old
and new alveolar bone near the moving teeth,
but rather a trabecular structure that strongly suggested a complete transformation of the entire
alveolar bone in that region. The bony trabeculae
were all rearranged in the direction of the force.
However, Oppenheim's conclusions that orthodontic forces were capable of transforming the
entire alveolus were rejected by his contempor-
412
aries as misinterpretations. He was also criticized

for using an animal with deciduous teeth for his
experiment, suggesting that the transformation he
had seen was related to growth and development
rather than being the outcome of applied mechanical forces.
Oppenheim's "transformation" hypothesis
might have supported Farrar's earlier contention
that orthodontic forces bend the alveolar bone,
and thus are able to stimulate all the cells in and
around this bone. However, Farrar's clinical approach also was not popular, because he advocated the use of heavy forces that could indeed
bend the bone. In fact, the pendulum swung furthest to the other side, when Schwarz10 recommended the use of light orthodontic forces. He
defined those forces as being "not greater than
the pressure in the blood capillaries" (15 to 20
mmHg, or about 20 to 26 g/cm2 of root surface).
A 1957 publication by Fukada and Yasuda"l
attracted wide attention. They observed that
bending of bone by mechanical means evokes the
generation of measurable electric potential spikes
in areas of compression and tension. While this
observation caused the rebirth of the field of applied exogenous electricity to bone nonunion
fractures, it also precipitated the reintroduction
of the concept of alveolar bone bending by orthodontic forces!2,13
III. HISTOMORPHOLOGY OF TOOTH

MOVEMENT
A. Observations
by Light Microscopy
pioneering work of Sandstedt and Oppenheim opened the door for comprehensive efforts to explore in detail the morphological
changes in the stressed PDL. For over 4 decades,
Reitan'4-'9 spearheaded this thrust with authority
and confidence. The strength of his work was
derived primarily from the extensive use of human material, whereby teeth that were to be extracted for orthodontic reasons were subjected to
a variety of orthodontic force systems, i.e., light,
heavy, continuous, intermittent, tipping, and
translatory. At the end of the experimental period, the teeth were removed together with their
surrounding tissues, and processed for histologic
The
evaluation. Moreover, Reitan studied paradental

tissues of animals subjected to orthodontic forces,
particularly dogs and monkeys, exploring the effects of age, function, type of bone, force magnitude, duration, and direction, on the morphological characteristics of the tissues. He concluded
that PDL cells in sites of tension proliferate, and
that newly formed osteoid in these areas resorb
slowly when subjected to pressure. In examining
tissues from different species,20 Reitan observed
that their responses varied, and attributed this
variability to the differences in their structural
composition, i.e., alveolar bone density, frequency and distribution of marrow spaces, and
the cell and matrix constitution of the PDL.
The realization that the rate of orthodontic
tooth movement in humans varies and is unpredictable prompted Storey to suggest that it depends upon the magnitude of the applied force,21'22
or the presence of hormonal fluctuations, such
as those associated with the menstrual cycle.23
These clinical observations motivated Storey24-27
to conduct a series of experiments in rodents in
which he applied forces of different magnitudes
to the maxillary incisors, causing lateral tooth
movement and mid-premaxillary sutural widening. In the rabbit and rat the teeth moved faster,
as the force was increased, but as in man, there
seemed to be a range of force magnitudes that
could be termed optimal.24 Near teeth treated with
such a force, newly formed bone appeared more
mature, while heavy forces were associated with
the formation of a highly cellular, poorly calcified matrix. Heavy forces caused periodontal necrosis and other destructive changes in the PDL,
while light forces appeared to be favorite when
moving a tooth through a thin plate of bone, as
apposition of bone on the labial surface seemed
to lag behind the resorptive activity on the PDL
side.25 In older animals, the rate of tooth movement decreased, perhaps due to reduced cellular
activity.26 Based on these observations, Storey,27

concluded that the process of tooth translation
through bone consists of three different phenomena: bioelastic, bioplastic, and biodisruptive. The
PDL and alveolar bone, due to their fluid-fiber
composition, can be deformed elastically by external stresses, which also evoke cellular activities. When the tissue elastic limit is reached, it
starts to deform plastically, with adaptive prolif413
erative and remodeling reactions. Prolonged

forces that exceed the bioplastic limit result in
biodisruptive deformation, with ischemia, cell
death, inflammation, and repair. Clinically
speaking, Storey asserted that light forces within
the bioplastic range would cause slow tooth
movement, while optimal forces that cause teeth
to move faster are within the boundaries of the
biodisruptive range.27 He concluded that within
the optimum range, tooth movement is rapid, but
the quality of remodeling bone is poor, increasing
the potential for relapse, once external force application ceases.
In contrast to Reitan, who was able to study
human paradental tissues following the application of forces to teeth, Storey's histologic work
was conducted solely on rodents. Storey observed that in humans teeth move at different
rates when forces of various magnitudes are used,
and he used rodent incisors to explore the reasons
for this phenomenon. While generally Storey's
observations correlated well with findings of other
investigators, it is doubtful whether conclusions
derived from experiments in rodents can be directly extrapolated to man, due to marked physiological differences, as pointed out by Reitan.20
Nevertheless, Storey's reports are significant, as
he emphasized the development of an inflammatory process in the stressed PDL, even when
light forces are being used.27
Reitan's and Storey's investigations demonstrated the complexity of the tissue reaction
during tooth movement. It was no longer perceived as a simple phenomenon of applied force
causing the tooth to move within the PDL, leading to tension and compression, and subsequent
bone formation and resorption, but rather as a
dynamic set of events that involved profound alterations in cellular functions and changes in matrix composition. Thus, histology facilitated the
morphological description of changes in the dentoalveolar complex that followed the administration of orthodontic forces to teeth. While being
unable to explain why alveolar bone and PDL
cells are responsive to applied mechanical
stresses, or how these physical entities evoke biochemical responses by the cells, the histological
investigations unveiled the sites of cellular activity, and enabled other researchers to ask "why"
and "how". These questions were explored by
the use of methods such as histochemistry, electron microscopy, autoradiography, and immunohistochemistry. In addition, PDL and alveolar
bone, recognized as the prime targets for orthodontic forces, were obtained from animals and
humans and were subjected to mechanical stresses
in culture conditions, either in tissue form or as
isolated cells.
B. Histochemical Changes Associated

with Force-Induced Paradental Tissue
Remodeling
Activities of oxidative enzymes and phosphatases were localized in the PDL of rats during
force-induced tooth movement by Deguchi and
Mori28 and Takimoto et al.20'30 They caused tooth
movement by placing a piece of stretched rubber
between the first and second molars, forcing the
teeth to move in opposite directions. (This
method, introduced by Waldo and Rothblatt,31
was later adopted by a number of investigators
who used rats as experimental animals in studying tooth movement. It does not allow measurements of force magnitude, and the rubber pieces
can traumatize gingival and periodontal tissues.)
They reported on an increase in the number of
osteoclasts displaying high succinic dehydrogenase activity in PDL pressure zones after 24 h.
In contrast, they observed no changes in the distribution of acid phosphatase and lactate dehydrogenase in the stressed PDL.
Rats were also used by Lilja et al.32 to study
the distribution and activity of a number of enzymes associated with alveolar bone resorption.
One maxillary molar in each rat was moved bucally by light (5 g) or heavy (36 g) forces generated by a spring attached to the incisors for
either 10 h or 1, 3, 4, or 6 d. The activities of
acid phosphatase increased in PDL cells in
compression zones, as well as in adjacent gingival cells and alveolar crest periosteum. Staining
for acid phosphatase also increased in marrow
cells and in osteocytes near the PDL pressure
zone. Lactate dehydrogenase activity, a marker
of vital cells, disappeared from areas of PDL
pressure ("hyalinized zones"), where cells apparently died in both cases of light and heavy
forces. Interestingly, prostaglandin synthetase
414
activity was found in alveolar bone marrow cells

and in gingival cells, but not in PDL cells of the
rat. Unfortunately, quantitative analysis of the
data in this study was impossible, because each
group consisted of only one animal.
In a related experiment, Lilja et al.33 examined the PDL pressure zone of human premolars
following force application for 1 to 30 d. Dilated
blood vessels were seen near the hyalinized zone
throughout the experiment. Intense activities of
arylsulfatase and prostaglandin synthetase and
moderate activity of aminopeptidase M were
found in macrophages degrading the hyalinized
zone. Adjacent osteoclasts displayed intense activity for succinic dehydrogenase and acid phosphatase. Again, each treatment time group consisted of one to two subjects, precluding statistical
analysis of the data.
The generation of osteoclasts in the compressed PDL did not escape attention, leading
investigators to examine this zone in an effort to
elucidate specific histochemical aspects of bone
remodeling. Kurihara and Enlow34 stained rat
maxillary second molar sections with ruthenium
red, which demonstrates the presence of glycosaminoglycans (GAG). In areas of alveolar bone
resorption, reattachment of the PDL seemed to

occur as a result of GAG secretion by fibroblasts
and osteoblasts, serving to link new and old collagenous fibers. Here, too, quantitation was impossible due to inadequate sample size. In con-
trast, Martinez and Johnson35 were better able to

assess the effect of orthodontic forces on alveolar
bone GAG content in rats. They moved maxillary
molars in groups of four rats, treated for 1, 3, or
5 d with a spring (25 g). They found that GAG
staining in alveolar bone at PDL tension areas
(achieved with alcian blue) increased at day 3.
However, in an external control group that received an inactive spring, the GAG staining was
lighter than in the untreated side of the maxilla
of the rats treated with an active spring. Thus,
this well-planned experiment demonstrated the
need for a control group in which the orthodontic
appliance remains inactive.
Compression zone osteoclasts in rats were
also the targets of an investigation by Noda,36
who sought to determine the effect of calcitonin
on osteoclastic cytochrome c oxidase activity.
First, maxillary molars were moved lingually with
a spring for periods of time ranging from 15 min

to 72 h. Enzymatic activity was localized in mi-
tochondria of osteoclasts by electron microscopy.

Calcitonin injections caused an early reduction
in the number of mitochondria and enzymatic
activity in detached osteoclasts, but this effect
was abolished 72 h after calcitonin administration. These results demonstrate that locally induced bone resorption may be affected by a boneseeking hormone.
The above-mentioned histochemical studies
did not, for the most part, produce quantifiable
data. Nonetheless, they painted a picture of enzymatically active cells, engaged in the remodeling of the stressed PDL and alveolar bone. In
areas of PDL compression, oxidative enzymes
and proteinases were localized in osteoclasts and
in macrophages removing necrotic tissue from
hyalinized zones, demonstrating heightened metabolic rates in cells involved in alveolar bone
resorption and degradation of PDL matrix and
dead cells. Further details on the activities of
these cells, as well as those located in PDL tension sites, are derived from experiments in which
electron microscopy was utilized as the investigative tool, as discussed in the following section.
C. Ultrastructural Changes in Paradental

Tissues During Tooth Movement
Despite Reitan's reported observation'9 that

paradental tissues of the rat are different than
those of man in many respects, as, for example,
morphologically and physiologically, rats remained the experimental animal of choice in
transmission electron microscopic (TEM) studies
of paradental tissues during tooth movement.
Rygh and Selvig37 described finding degradation
products of erythrocytes in enlarged blood vessels and in the extravascular space of the compressed PDL. The tension site in the PDL was
studied by Ten Cate et al.,38 who later also ex-
amined stretched cranial sutures in rats.32 In both

PDL and suture they observed that fibroblasts
were apparently engaged in synthesizing as well
as degrading collagen. Fibroblasts entering areas
of matrix disruption were termed "pioneers", a
term introduced earlier by Rygh40 for identifying
cells entering hyalinized zones in the compressed
415
PDL. In the latter site, Rygh was undecided on

whether these phagocytic cells were primarily
macrophages or fibroblasts. In examining PDL
tension zones in rats that had been subjected to
force application (5 to 10 g) to a maxillary molar,
Rygh41 observed distended blood vessels and mitotic PDL cells. Spaces appeared in the stretched
PDL, which were filled with flocculent material.
Collagen fibers were primarily oriented in the
direction of tension, but many nonoriented fibrils, as well as elastic-like (oxytalan) fibers, were
visible. The latter were detected earlier by
Edwards42 in dogs.
Teeth are attached to the alveolar bone with
embedded PDL fibers (Sharpey's). Martinez and
Johnson43 examined this attachment in rats using
scanning electron microscopy (SEM). They reported that 5 d of tooth movement significantly
reduced the diameter of the attached fibers, suggesting a reduction in the mechanical strength of
the PDL. Kurihara and Enlow44 used TEM in an
attempt to elucidate the nature of the attachment
of PDL fibers to bone during resorptive activities.
They concluded that the most prevalent type of
attachment in resorptive sites is adhesive in nature. It consists of a layer of ground substance,
deposited by fibroblasts on the naked surface of
recently resorbed bone. Later, collagen fibrils are
secreted into this layer, coalescing into fibers. In
this fashion, partially released old bone matrix
fibers intermingle with the newly formed PDL
fibers.
Since Kethcham's report in 1927 that orthodontic treatment is associated with radiographic
evidence of significant dental root resorption in
many moving teeth, this phenomenon was explored by numerous investigators in man and experimental animals using light microscopy, TEM,
and SEM. Via SEM, Kvam45 examined human
premolars that had been exposed to orthodontic
forces. After 5 d of treatment, small areas of root
resorption were found on the margins of the compressed PDL of all teeth, and after 25 d all treated
teeth displayed resorption lacunae penetrating
through the cementum into the dentin. Extensive
external root resorption was observed in the teeth
of patients whose palates had been expanded rapidly by Barber and Sims46 and by Langford and
Sims.47 In this procedure, heavy forces were applied for about 14 d to teeth anchoring the ex-
pansion device, and the teeth were then retained

in their new position for a few months to allow
bone to fill the expanded mid-palatal suture. All
anchor teeth exhibited root resorption lacunae,
and the degree of resorption was directly related
to the length of the retention period. Repair of
root defects by cellular cementum was observed,
but with little evidence of PDL fiber reattachment.
Using light microscopy and TEM, Rygh48
investigated the PDL compression zones in rats
whose molars were subjected to orthodontic forces
(5, 10, or 25 g) for 2 h to 28 d. Root resorption
lacunae were seen near the hyalinized zone, in
close proximity to a rich PDL vascular network.
Rygh suggested that root resorption might be a
side effect of the cellular activity associated with
the removal of the necrotic tissue of the hyalin-
ized zone, and described the removal of the cementoid layer as the elimination of the defense
against resorption in an area that is strongly proresorptive. This proposed association between
root resorption and PDL injury was supported
recently by the results of an experiment in rats
by Nakane and Kameyama.49 In that study the
gingiva and PDL of a maxillary molar were injured repeatedly three times at 3-h intervals, by
insertion of a 2-mm-long needle. Root resorption
developed within 1 d near the traumatized, inflammatory PDL and continued through the 21
d experimental period, with concomitant repair
by cementoblasts.
Since mineralized tissues remodel under the
influence of systemic and local factors, it was
suggested that factors associated with maintenance of calcium homeostasis might regulate the
activity of root resorbing cells. To test this hypothesis, tooth movement was performed by researchers in hypocalcemic rats. Goldie and King50
created calcium deficiency in adult lactating rats
and applied 60 g force to a maxillary molar for
1 to 14 d. The teeth in these rats moved significantly faster than in the control animals, as their
bones underwent extensive resorption. However,
SEM measurements determined that the extent
of root resorption was decreased in the calciumdeficient rats, suggesting that bone resorting cells
are more responsive to bone seeking hormones
than cells that resorb roots of teeth. In a more
recent experiment, Engstrom et al.51 moved apart
maxillary incisors in growing rats (30 d old) who
416
were being fed a calcium- and vitamin-D-deficient diet. Using light microscopy, they determined that root resorption in the moving teeth
was enhanced in the hypocalcemic rats, in contrast to the finding of Goldie and King.50 This
discrepancy may be a result of the age difference
between the two groups of hypocalcemic rats, as
well as because molars were tested in one study,
while incisors were examined in the other.
D. Autoradiographic Examination of the

PDL During Tooth Movement
The PDL contains a mixed population of cells
that can synthesize or degrade bone, cementum,
and the nonmineralized PDL itself. Some of the
metabolic processes associated with these activities have been investigated by the use of autoradiography. In this method, radiolabeled amino
acids are injected into animals, and their timerelated location is determined by exposing tissue
sections to radiation-sensitive photographic
emulsions. The resulting silver-bromide salt
crystals that form over radioactive sites can be
localized microscopically. In tooth movementrelated studies, investigators utilized tritiated
proline (3H-proline) to study the kinetics of collagen synthesis and tritiated thymidine (3H-Tdr)
to study cell proliferation.
Garant and his collaborators52-55 have studied
extensively the process of collagen remodeling
by PDL fibroblasts. They administered 3H-proline into mice and rats to determine the pattern
of collagen synthesis by PDL fibroblasts by using
both light microscopy and TEM. They described
PDL fibroblasts as being elongated, polarized
cells, with the nucleus positioned at one end and
the cytoplasmic and secretory part at the other
pole. Fibroblasts were found to be distributed
evenly throughout the rodent PDL, and to migrate
between the fibers, interacting during motion with
adjacent matrix and cells. This motion appears
to be facilitated by cellular microfilaments (actin
and myosin), and by attachment to the matrix
with glycoproteins (mostly fibronectin).56 Garant
and Cho concluded that in tooth movement, PDL
fibroblasts in tension sites express the phenotype
of actively migrating and matrix-secreting cells.

Cells in the normal PDL proliferate and die
regularly,57 but these events are markedly increased during tooth movement.58 Proliferative
activities in the mechanically stressed rat PDL
were studied extensively by Roberts et al.59-66 and
by Yee et al.67'68 Uptake of 3H-Tdr by PDL cells
in tension sites was increased significantly within
2 h of the insertion of an elastic material between
the first and second maxillary molars. Most of
the mitotic activity occurred near the bone and
the middle of PDL, but not near the dental root.
Some of the newly divided cells appeared to migrate in the direction of the alveolar bone, perhaps because they were preosteoblasts.61 Based
on these observations, Roberts and his associates
concluded that stretching the PDL causes G,arrested cells to enter mitosis, while G1 cells are
stimulated to start synthesizing DNA. The latter
cells are readily labeled by 3H-Tdr.
In an effort to identify and classify PDL cells
at different stages of differentiation, Roberts and
Cox69 resorted to measurements of nuclear volume in these cells. This tedious method revealed
that the nuclei of osteoblasts are larger than those
of fibroblasts, a fact that can be used to identify
committed osteoprogenitor cells. While fibroblasts were found predominantly near PDL blood
vessels, osteoblastic progenitors were located
further away from the vessels and closer to the
bone and cementum surfaces. These reports create the impression that the PDL preosteoblastic
population resides solely within the boundaries
of the PDL. However, McCulloch et al.70 and
McCulloch and Heersche71 have attracted attention to the finding that in mice many alveolar
bone marrow spaces are directly connected
through vascular channels with the PDL. Moreover, frequent injections of 3H-Tdr into large
groups of mice, and subsequent autoradiographic
examination of their mandibles revealed that the
endosteal spaces contain many labeled progenitor
cells. Thickened areas of cementum were found
opposite openings of these channels in 64% of
the examined specimens. Although it is presently
unknown whether progenitor cells that originate
in alveolar bone marrow spaces participate in the
PDL response to mechanical stress, it is tempting
to speculate that such an association indeed exists.
417
E. Tooth Movement: Visible Tissue
Changes
Clinicians realized 2000 years ago (perhaps
earlier) that teeth can be moved from one position
in the mouth to another by being subjected to
persistent mechanical forces. Merely 250 years
ago, Hunter made the first attempt to explain the
biological basis for this dental movement. Without having seen the involved tissues magnified
by a microscope, he postulated that force-induced
tooth movement was facilitated by what we may
term today as bone remodeling. This notion was
verified in the early years of the 20th century by
Sandstedt, who described in detail the effects of
mechanical force on the PDL-alveolar bone interface. His work illustrated clearly that orthodontic tooth movement is made possible by the
resorption of alveolar bone near sites of compression in the PDL, while bone apposition occurs
where the PDL is stretched. A controversy erupted
a few years later when Oppenheim suggested that
the entire alveolar bone near a moving tooth remodels, including endosteal surfaces. Most, if
not all, of the investigators who were engaged in
exploring this issue during the first half of the
20th century overwhelmingly supported Sandstedt's hypothesis because, clearly, the most dramatic initial histologic changes could be seen in
the stressed PDL and its immediate bordering
mineralized tissue surfaces, bone and dental root.
Reitan, whose comprehensive histologic explorations spanned over 4 decades into the second
half of this century, reported on bone remodeling
in alveolar bone marrow spaces and gingival periosteum, both at a distance from the stressed PDL.
These observations appeared to support Oppenheim's transformation hypothesis, and compelled
Reitan to accept the century-old proposition of
Farrar that orthodontic forces bend the alveolar
bone. In the late 1960s Baumrind succeeded in
demonstrating that such a bending effect indeed
takes place, while others have measured spikes
of altered electric potentials in teeth, PDL, and
alveolar bone that had been subjected to orthodontic forces.
The search for an optimal orthodontic force,
a force that would be most efficient in moving
teeth, led two investigators who examined paradental tissues microscopically to make contrast-
ing recommendations. Schwarz warned against

using "heavy" forces, forces that occlude PDL
capillaries and thus can damage the tissue. However, Storey recommended utilizing forces that
do cause damage to the PDL, biodisruptive forces
that introduce inflammation into this tissue. He
also showed that within a certain range, tooth
movement could be accelerated concomitantly
with an elevation in force magnitude. Like Reitan, Storey associated slow tooth movement in
adults with a slow rate of cellular activity.
With the advent of electron microscopy, detailed information emerged in the last 2 decades
on the ultrastructure of dento-alveolar tissues.
Cells and matrices were studied in great detail
by the use of TEM and SEM. Moreover, histochemical and autoradiographic investigations shed
new light on biochemical events that occurred in
the dento-alveolar tissue complex during normal
existence and while under altered states of mechanical stress. It became evident that cells that
remodel the dento-alveolar complex are equipped
with an elaborate system of cytoplasmic organelles that enable them to synthesize and secrete
matrix components and the enzymes that participate in the degradation of this matrix. Garant
and Ten Cate and their associates demonstrated
that PDL fibroblasts can readily remodel the matrix, as well as migrate through it, while Jee and
Roberts and their collaborators identified preosteoblasts both in the PDL, following their cell
cycle kinetics, and through the stretched PDL.
In the compressed PDL, Rygh, Kvam, and others
have identified macrophages that seemed to remove necrotic tissue.
Taken together, the above microscopic studies on both light and electronic levels have described in great detail morphological changes,
and some fundamental physiological alterations
that seem to occur in dento-alveolar tissues during tooth movement. However, with the exception of the proponents of the bone bending hypothesis, none of the above investigations have
addressed the question of the mechanism of transduction of physical stimuli to biological reactions. Those who advocated the idea that bent
bone generates electric potentials proposed that
these potentials somehow stimulate cells in a mechanically stressed area by causing structural and
enzymatic changes in the cellular plasma mem-
418
brane. However, interest in this question has increased rapidly in recent years, far beyond the
boundaries of orthodontic tooth movement.
Moreover, it appears increasingly evident that the
regulation of cellular functions in most, if not
all, tissues is under the influence of local and
systemic factors that are derived from the endocrine, nervous, and immune systems. Consequently, Section IV reviews evidence in support
of the hypothesis that dento-alveolar tissue remodeling during tooth movement is an outcome
of cellular activities that are regulated by interactions between physical distortions and locally
distributed humoral factors that act as endocrines,
paracrines, or autocrines.
IV. MECHANISMS OF CELLULAR

STIMULATION IN TOOTH MOVEMENT
A. The Effect of Mechanical Stress on
Cells
Cells of all kinds are subjected at one time
or another to compression, stretch, and shearing.
There is growing evidence that most cells have
ion channels that are potentially capable of regulating active and passive variations in cellular
mechanics. According to Morris,72 these ion
channels are mechanosensitive, i.e., their openstate probability depends on stress at the membrane. Such channels were postulated decades
ago as a means of mechanoelectrical transduction
in muscle and nerve cells. However, it now seems
that most other cell types have such channel components in their membranes. These channels are
ubiquitous, occurring at uniform density, on the
order of 1/jim2 and in every cell.73-75 Calcium
ions may enter cells in significant amounts through
these channels.76-78 According to Morris and Sigurdson,79 tensions generated in patch electrodes
to activate stretch sensitive channels are of the
same order of magnitude as those measured in
migrating fibroblasts.80
Cell membrane tension may result from intracellular osmotic changes, contraction of cytoskeletal elements, or physical changes in the
extracellular matrix. In 1985, Ingber and
Folkman81 constructed three-dimensional cell
models comprised of a discontinuous array of
compression-resistant struts, pulled open by connections with tension elements. The stability of
such a structure depends on maintenance of tensional integrity. Based on these models, and observations of cellular behavior in vitro, these authors concluded that important functions are
regulated by alterations in the integrity and composition of the extracellular matrix. Interconnections between mammalian cellular nuclei and the
plasma membrane, as well as with the extracellular matrix, are through the continuous system
of cytoskeletal filaments and cell surface transmembrane receptors. Physical forces, either those
generated by the cytoskeleton or in the matrix,
appear to be important regulators of cell and tissue growth. This interaction between force and
cell function was observed to exist in skeletal
myotubes,82 lymphocytes,83 arterial smooth mus-
cle cells,84 osteosarcoma cells,85 and endothelial

cells.78
Ingbar and Folkman81 hypothesized that if
physical stimuli can be translated into metabolic
alterations through changes of intracellular structure, then mechanochemical transduction of these
signals is most likely mediated by the structural
linkages that join the cytoskeleton with the external milieu. In 1985, Ingber and Jamieson86
proposed that the cellular mechanism of mechanochemical stimuli is transduced into chemical information through local changes in thermodynamic parameters. In this fashion, activation
energy of a reaction is produced by pressure and
volume alterations, and various chemical reactions and macromolecular polymerization processes can be selectively promoted or inhibited
as a result of mechanical perturbation of the cell
surface. Indeed, Joshi et al.87 have been able to
demonstrate that intracellular cytoskeletal polymerization can be modulated by mechanical forces
applied to the cell surface in neurites. Similar
changes may be caused by cell growth factor
interactions. For instance, Herman and Pledger88
reported on alterations in the distribution of actin
and vinculin in fibroblasts as a result of exposure
to platelet-derived growth factor. Likewise, the
arrangement and function of steroid hormone receptors may be very sensitive to mechanical perturbation because they appear to be associated
physically with the nuclear protein matrix.89-91
Nicolini et al.92 reported that a specific in419
crease in nuclear size is necessary for S phase

initiation, an observation that was reported earlier
by Roberts and Cox69 to occur in PDL cells.
Nuclear shape seems also to play an important
role in regulating nuclear transport. For instance,
Jiang and Schindler93 studied the effect of various
growth factors on nuclear transport, and suggested that changes in nuclear shape may be permissive for delivery of growth factor-receptor
complexes to their site of action in the nucleus.
Experimenting with mammary epithelial
cells, Emerman and Pitelka94 observed that cell
rounding is usually associated with inhibition of
cell growth and with promotion of cytodifferentiation. Thus, cells that produce specialized
products usually appear round, a shape that might
facilitate exposure of specific parts of their genome. In orthodontic tooth movement, such transformations in cellular shape are readily visible in
mechanically stressed paradental cells (Figures 1
to 3). In unstressed PDL sites (Figure 1), alveolar
bone osteoblasts appear flat, while those in areas
of PDL tension (Figure 2) seem large and round.
In areas of PDL compression (Figure 3), PDL
fibroblasts assume a round shape. Histologic
studies by Reitan'4-16 and Rygh41 have demonstrated that activated osteoblasts in PDL tension
sites are engaged in producing a new bone matrix, while PDL cells in compression sites are
primarily involved in enzymatic degradation of
the compressed extracellular matrix.
B. The Effect of Mechanical Stress on

Mineralized and Nonmineralized
Connective Tissues
1. Regulation of Bone Remodeling In
Vivo by Applied Stresses
Typically, orthodontic forces are applied

continuously to teeth and their surrounding tissues. These forces evoke cellular activity, as has
been demonstrated by investigators who examined affected tissues mircoscopically. However,
it is unclear how long a force should be applied
to stimulate target cells in particular areas of PDL
and alveolar bone. Lanyon and his associates have
addressed this issue in a series of experiments
whereby controlled strains were applied to avian
FIGURE 1. "Flat" alveolar bone osteoblasts (arrows)

5-[pm horizontal section of cat maxilla, stained
immunohistochemically for cGMP. Tissue near control,
nonorthodontically treated canine. B, alveolar bone; P,
in a
periodontal ligament. (Magnification
1400.)
allowing the investigators to examine, radiographically, histologically, and histochemically, the effects of various strains on
bone remodeling95-04 Their experimental model
consisted of surgical removal of bone from both
proximal and distal epiphyses of the ulna in turkeys and roosters, freeing the entire diaphysis
from regular functional strains, leaving its nervous and vascular supplies intact.95 Mechanical
loads are then introduced to this bone through
stainless steel pins attached to an external loading
apparatus. The operation caused removal of loadbearing by the ulna, followed by a loss of bone
mass.96 This loss was prevented by 4 cycles per
day of an externally applied loading regimen
bone in vivo,
(10,000 to 12,000 microstrain, 0.5 Hz), for 42
d. When the number of cycles was increased to

36 per day, bone formation increased significantly. Static (continuous) loads had no effect in
420
FIGURE 2. Alveolar bone osteoblasts (arrows) at PDL

tension site after 14 d tipping force application to cat
maxillary canine. Horizontal section, 5 ixm thick, stained
immunohistochemically for cGMP. Notice the round appearance of the cells. B, alveolar bone; P, periodontal
ligament. (Magnification
1400.)
this preparation on bone remodeling, while similar loads applied intermittently for a total of only
a few minutes daily increased bone mass substantially.97 The magnitude of the strain in the
loaded bone appeared to be directly associated
with the nature and degree of the remodeling
response.4 Peak longitudinal strains below 0.001
were associated with bone loss, while peak strains
above this level were associated with substantial
enhancement of periosteal and endosteal bone
formation.98 These effects were found also in
birds suffering from calcium deficiency."
These experiments demonstrated that bone
cells in vivo are very sensitive to a small number
of strain cycles daily. A maximal osteogenic response was obtained by only 72 s of load bearing.
Moreover, it seemed like the creation of a static
load environment is essentially ignored as an os-
FIGURE 3. Periodontal cells in PDL compression zone

at the border of the necrotic "hyalinized zone" after 14
d of tipping force application to cat maxillary canine.
Horizontal section, 5 ipm thick, stained immunohistochemically for cGMP. Notice the enlarged size of the
cells in comparison to the thinner PDL cells seen in
Figure 1. (Magnification x 1400.)
teoregulatory stimulus, suggesting that functional

influence on bone architecture is derived solely
from intermittent loading. 00 Lanyon101 then proposed a hypothesis to explain the mechanism by
which bone adapts to functional load bearing. In
his opinion, the osteocytes are the most likely
candidates to sense the distribution, rate of

change, and magnitude of strain in the bone matrix. Following their recognition of a change in
the strain situation, osteocytes communicate with
the bone surface cells that remodel the bone. It
seems like the important feature of strain in this
respect is in the occurrence of an abnormally
distributed strain rather than an unusually large
strain. Evidence for osteocytic response to applied stress was found in the higher level of glucose-6-phosphate dehydrogenase in these cells,
and a sixfold increase in the number of osteocytes
421
that have incorporated 3H-uridine into their

RNA.102 Furthermore, Lanyon hypothesized that
osteocytes respond not only to the transient effects of mechanical strain, but also to the persistent effect of mechanical strain on the matrix.
As a candidate for such strain-sensitive matrix
components, he chose proteoglycans.103 Examining bone sections in a polarizing microscope,
Lanyon quantified their birefringence and observed matrix proteoglycan reorientation following short mechanical loading. These large, highly
charged matrix molecules could be forced by
strain to attach to cell surface receptors, or pass
into the cell and attach to its cytoskeleton. Since
the proteoglycan reorientation persists for 1 to 2
d, it could provide a physical basis for a "strain
memory" in bone.'03 In a more recent study,
Skerry et al.104 found that reversal reorientation
of bone matrix proteoglycans also occurs in vitro,
not only in avian bone, but in bones from rats
and dogs as well. They attributed this molecular
distortion to strain-generated fluid flow, usually
preferentially oriented with the direction of collagen fibers.
The experiments of Lanyon and his associates provide an attractive explanation of the longlasting effects of short-lived strains on bone cells.
However, the exact mechanism by which proteoglycan reorientation could influence the activity of target cells in bone remains unknown. In
orthodontics, functional appliances subject teeth
to intermittent forces, leading to their gradual
movement. In this situation, bone matrix distortion could perhaps be evoked and act in the fashion proposed by Lanyon and co-workers. In contrast, fixed-appliance orthodontics resorts to
continuous force applications. In this mode of
treatment, tissue effects often contain widespread
damage, intimately associated with inflammatory
and reparative responses. Moreover, orthodontic
tooth movement is materialized by intense resorptive activity of alveolar bone, while Lanyon
and associates' short-term strain application did
not evoke any resorptive activity, but rather extensive formative function by bone cells. Nonetheless, the implication of bone matrix in the
transduction of mechanical stimuli to physiological responses by bone cells serves to broaden
our understanding of the mechanism of cell stimulation by externally applied forces. This seem-
ingly critical involvement of bone matrix in the

response of bone cells to mechanical stress can
explain, at least in part, Oppenheim's9 earlier
observation of a transformation of the entire alveolar process during tooth movement.
2. Bioelectric Phenomena in Bone

The dependency of bone tissue integrity and
metabolism on mechanical stress has long been
recognized. At the present time, growing evidence strongly suggests that osteoporosis can be
curtailed or reversed by regular physical exercises, which subject skeletal elements to musclederived forces. 05106 Astronauts and animals that
have participated in space flights or in experiments on simulated weightlessness demonstrated
continuous loss of skeletal tissue due to the lack
of gravitational forces.107"' These observations
imply that mechanical stresses regulate the activity of skeletal cells, confirming Wolff s6 proposition that the structural architecture of bone
depends on the nature of the mechanical stresses
applied to it. Applying pressure and tension to
chick embryo long bone rudiments in vitro,
Glucksmannll2 observed in 1942 that optimal cartilaginous tissue structure was obtained only in
the presence of mechanical stress. Experiments
of this sort, which have persisted, demonstrated
that skeletal tissues and cells can respond to applied mechanical stresses in vitro, even in the
absence of other seemingly important systems,
such as the nervous and vascular systems. Thus,
bone loomed brightly as a self-contained tissue,
whose response to mechanical stress is independent of any other tissue system. An example
for this rather narrow approach, which attributed
most of the control of the response of bone to
mechanical stress on the bone cells themselves,
can be found in the proceedings of a recent conference on functional adaptation in bone tissue. 13
Isolated bones or bone cells were presented at
that conference as being fully capable of responding biochemically to applied mechanical stresses.
A proposed major link in the cascade between
the applied force and the biological response was
stress-generated electric potentials.114,115
The concept of the inherent ability of bone
to respond to applied mechanical stress was
422
boosted by Fukada and Yasuda's" report in 1957

that measurable electric potentials are evoked
temporally in bent bone. They investigated dry
specimens cut from femora of man and ox, and
were able to measure direct and converse piezoelectric effects in those bones. They concluded
that the piezoelectric effect appears only when a
shearing force is applied to the collagen fibers of
the bone lattice, causing them to slip past each
other. More recently, Marino et al."6 investigated the piezoelectric characteristics of collagen
films and concluded that these effects originate
at the level of the tropocollagen molecule, or in
molecules no larger than 50 A in diameter. Further support for the concept that collagen is the
source of the piezoelectric effect came from studies on tendons17'118 and on decalcified bone. 19
Bassett and Becker120 reported that a net negative
potential and a net positive potential appear, respectively, on the compression and tension sides
of bone. These phenomena were recorded later
by Cochran et al.'21 in a segment of a bovine
mandible, and by Gillooly et al.12 and by Zengo
et al. 122,123 in dog mandibles. Marino and Gross'24
recently compared piezoelectric surface charges
of human bone with those of the cementum and
dentin of whale teeth. They found that cementum
and dentin were capable of producing only about
12% of the surface charge produced by cortical
bone, and concluded that "piezoelectricity mediates alveolar (bone) remodeling". This is a typical statement, which narrows the effect of mechanical stress on bone to the generation of electric
potentials by the stressed collagen.
The possibility that alveolar bone is indeed
bent by the application of tipping or translatory
forces to teeth was first suggested by Farrar.5
This event was later confirmed by Baumrind13 in
rats and by Grimm125 in humans. It led DeAngelis'26 to propose that the alteration of the
electric environment within the stressed alveolar
bone may regulate differentiation of bone progenitor cells. A mechanism by which these potentials may reach the surface of bone cells was
suggested by Pollack et al. 15 According to this
concept, bone is surrounded by an electric double
layer in which electric charges flow in accordance
with a stress-related fluid flow. This stress-generated potential may affect the charge of cell
membranes, as well as that of macromolecules
in their vicinity. Borgens,127 examining intact and

damaged mouse bones, detected endogenous ionic
currents that he attributed to streaming potentials,
rather than to piezoelectricity, due to the long
(up to 30 min) current decay period. He suggested that the source of current in mechanically
stressed bone is cells rather than matrix. Otter et
al.,128 studying dry and wet specimens of bovine
tibia, concluded that while in the dry state the
current is primarily piezoelectric, in wet bone the
dominant mechanism is streaming potentials.
Bioelectric measurements in alveolar

bone'22,'23 have demonstrated that the com-
pressed (concave) side of the orthodontically

treated bone is electronegative with respect to the
tension (convex) side, suggesting that negative
potentials during bone bending can generate bone
deposition, while positive potentials are responsible for bone resorption. However, Borgens' experiments in fracture sites'27 failed to find such
a correlation. Rather, his measurements showed
that current enters the lesion, where its dispersion
(i.e., its pathway and density) remains unknown

due to the complexity of the distribution of mineralized and nonmineralized matrices.
Whichever is the source of electric potentials
in bone, it seems that these endogenous currents
are involved in bone repair, remodeling, and perhaps growth. This conclusion led numerous
investigators129-"33 to apply weak currents to nonunion bone fractures in an effort to facilitate healing. Clinical successes in orthopedics prompted
orthodontists to combine orthodontic force application with administration of weak electric
currents to jaw tissues in an effort to determine
whether a synergistic effect on the rate of tooth
movement would be achieved. In the first experiment of this kind, Beeson et al. 134 implanted
electrodes in cat mandibles and applied a 10-1pA
direct current constantly for 5 weeks. No significant differences between electrically treated and
control animals were found, regardless of whether
the cathode or the anode were placed in the vicinity of the moving tooth. This absence of an
effect seems to have stemmed from the placement
of the electrodes near the apex of the moving
teeth, rather than near the alveolar crest where
most of the force-induced bone remodeling occurs when teeth are tipped. Different results were
obtained by Davidovitch et al. ,135 136 who applied
423
a current of 20 ILA to gingiva near orthodontically

tipped maxillary canines in young adult cats. Significant acceleration of the rate of tooth movement was observed after 7 and 14 d of combined
application of force and electricity. This potentiating effect was explained by the placement of
the anode very close to the site of PDL compression, where alveolar bone resorption occurs, while
the cathode was placed in close proximity to the
site of PDL tension, where new bone is deposited.
3. Biochemical Events in Cells Affected

by Mechanical Forces In Vitro
The ability to maintain cells and organs in
culture permitted investigators to subject them to
mechanical stress and to monitor their response
to either tensile or compressive forces. These
experiments targeted specific cellular functions,
such as proliferation, synthesis, and secretion of
extracellular matrix components, or the enzymes
that degrade them. The rationale for these experiments is derived from the assumption that in
vitro conditions facilitate the exposure of sensitive target cells to a specific stimulus, in the
absence of any other factors that might exist in
vivo, which would mask or "confound" the response that is observed in vitro. As discussed
below, the situation in the intact mammalian organism differs from that of the culture system in
that bone cells in the living animal may become
subjected to mechanical stresses simultaneously
with signal molecules derived from neighboring
endothelial cells, fibroblasts, or migratory leukocytes.These molecules may amplify or diminish the effect of mechanical stress on the shape
and cytoskeletal structure of the bone cells. Thus,
for an in-depth understanding of the nature of the
response of cells to mechanical stress it is essential to study combinations of these interactions,
a design that is rarely performed for in vitro
investigations.
Among the main investigative yardsticks that
were utilized to explore the mechanism of activation of cells by mechanical forces were cyclic
nucleotides, prostaglandins, DNA, phosphatases, and metalloproteinases. Adenosine 3',5'monophosphate (cyclic AMP, or cAMP) and
guanosine 3',5'-monophosphate (cyclic GMP,
or cGMP) have been identified as mediators of

the effects of external stimuli on bone cells in
vivo'37-'39 and in vitro.'40'141 Fluctuations in the
levels of these substances have been found to
occur in bone treated by parathyroid hormone
(PTH),120 calcitonin,138 and vitamin D3,139 as well
as electric currents'40 and mechanical force.'41
Prostaglandins, particularly those of the E series, have been associated with bone remodeling
activities resulting from malignancies,142 gingival inflammation,143 rheumatic joint disease,'44
and fracture.'45 Thus, fluctuations in the levels
of cAMP and cGMP in mechanically stressed
cells and PGE2 in their culture media were used
frequently as indicators of cellular responsiveness.
In 1975, Rodan et al.141 applied compressive
forces to chick cartilaginous bone rudiments and
observed a reduction in the levels of cAMP and
cGMP in these cells within 15 min. Uchida et
al.146 stretched rat costochondral chondrocytes
from 1 min to 24 h, and reported an initial increase in cAMP content at 3 to 5 min, with a
decline to control levels shortly thereafter. The
incorporation of 35S into GAG by the stretched
cells increased significantly, but their rate of DNA
synthesis was not altered. Interestingly, when
stretched chondrocytes were incubated in the
presence of calcitonin, their cAMP levels at 24
h were much higher than those of control cells
or cells treated with PTH.
In 1980, Somjen et al.'47 stretched rat embryonic calvarial cells for 1 to 60 min. They
observed sharp increases in cAMP and PGE2 levels in cells and media, respectively, with peaks
at 15 to 20 min and subsequent declines. Both
cAMP and PGE2 levels failed to increase when
bone cells were stressed in the presence of indomethacin, a potent inhibitor of prostaglandin
synthesis. More recently, Shen et al.'48 exposed
osteoblasts cultured from rat fetal calvaria to different concentrations of PGE2, and within 10 min
observed a transient change in shape from an
epithelioid to stellate, with markedly increased
numbers of gap junctions.
Rat calvarial cells were also used by Hasegawa et al.149 in an effort to determine the effect
of stretching on DNA and matrix component synthesis. Continuous or intermittent stretching for
2 h increased both the number of DNA synthes-
424
izing cells (by 64%) and the production of osteonectin-like molecules. These results suggest
that bone cells respond to mechanical stress by
increasing their numbers and by rearranging their
contacts to neighboring structures. Similar effects on DNA synthesis were obtained with avian
calvarial osteoblasts that were stretched for up to
5 d by Buckley et al. 50 In addition, the stretched
cells were uniformly aligned perpendicular to the
direction of the strain field.
Compressive forces were applied to bone cells
in a number of studies, usually by compressing
the gas phase above the culture medium. In this
fashion, Klein-Nulend et al.'51 applied intermittent pressure to mouse calvaria for 5 d. This
treatment increased alkaline phosphatase activity
and 45Ca uptake by the calvaria, while resorptive
activities decreased. The net result was a 16%
increase in the calvarial mineral content. Similar
effects were observed by these investigators following the application of intermittent compressive forces (132 g/cm2, 0.3 Hz) to fetal mouse
metatarsal bone rudiments.'52 They concluded that
compression inhibits the migration and activity
of osteoclasts and their precursors. Heavier, continuous compressive forces (3 atm) were applied
by Ozawa et al.'53 to osteoblast-like cells, resulting in suppression of osteoblastic activities
and marked enhancement of PGE2 production.
Unquestionably, the main arena of tissue remodeling during tooth movement is the PDL. It
is the prime target of tooth-moving mechanical
forces, and has been the subject of numerous
investigations aimed at elucidating details of the
biological response of its cells to applied mechanical stress. Duncan et al.154 applied mechanical forces to mouse molars in vivo as well as in
vitro. After 3 to 5 d in culture, large amounts of
PGE2 and type II collagen were synthesized by
the ligaments. A substitute model for the PDL
was introduced by Meikle et al.,155 who used a
spring to apply tensile stress to rabbit calvarial
sutures in vitro. They reported an increase in the
tissue levels of collagenase and a reduction in
the level of tissue inhibitors of metalloproteinases

(TIMP) in these sutures.
Fibroblasts isolated from chick embryos were
grown and stretched on nylon meshes by Curtis
and Seehar.'56 Intermittent stretching at 0.25 to
1.0 Hz caused significant increases in mitotic
frequency and in the proportion of cells in S

phase. It was concluded that stress speeds up the
mitotic cycle of fibroblasts, rather than switching
cells from G, to S. However, Norton et al.157
reported recently that tensile forces failed to
change the cytoskeletal configuration in PDL fibroblasts (as determined by immunofluorescence
of tubulin, vimentin, and actin), suggesting that
these cells are not responsive to tensile forces per
se, but rather to injurious effects resulting from
these forces. Human gingival fibroblasts were
stretched by Ngan et al.,158 who reported that a
5% increase in cellular surface area for 5 min to
2 h caused significant elevations in the levels of
cAMP and PGE2 in the cells and their media,
respectively. In a related study, Ngan et al.'59
reported recently on similar effects in stretched
human PDL fibroblasts.
Despite the major role played by PDL fibroblasts in tooth movement, a surprisingly small
number of investigations on the response of these
cells to mechanical forces in vitro have been performed to date. However, the relative ease of
obtaining these cells from extracted healthy human teeth,l60 and the availability of a number of
mechanical systems that can apply various modes
of compressive and tensile stresses to cells in
culture promise to facilitate new experiments in
the near future. However, since the PDL fibroblast-like cell population is heterogeneous, consisting of cells with differing phenotypes, and,
as the PDL also includes numerous precursors of
osteoblasts as well as epithelial and endothelial
cells, strict identification and phenotyping of these
cells will be required so that the data may be
interpreted more meaningfully.
Of great curiosity and perhaps importance is
the possible role of the epithelial rests of Malassez in tooth movement. These clusters of epithelial cells, left behind during the growth of the
dental root, are distributed throughout the PDL
in close proximity to the root surface cementum
layer. Their functional role continues to be an
enigma. Reitan'7 noted that these epithelial clusters are eliminated from necrotic areas of compressed PDL and do not regenerate. He speculated that these cells may play a protective role
in preventing force-induced root resorption. Brunette et al.'61 cultured monkey epithelial cells
derived from rests of Malassez, together with
425
PDL fibroblasts. They observed no junctional

structures between epithelial cells and fibroblasts, but in some cases islands of epithelial cells
were sandwiched between two layers of fibroblasts. Later, Brunette et al.'62 detected large
quantities of PGE and PGF in media in which
epithelial cells derived from porcine rests of Malassez had been incubated. They proposed that
these prostaglandins might participate in the regulation of alveolar bone remodeling, either by
affecting bone cells directly or indirectly by interacting with PDL fibroblasts and endothelial
cells. Support for the concept of interaction between PDL fibroblasts, epithelial, and endothelial cells was provided by Merrilees et al. 163 They
found that the GAG synthesized in vitro by PDL
fibroblasts is predominantly chondroitin sulfate,
whereas epithelial cells produced primarily hyaluronic acid. However, endothelial cells or their
conditioned media, when cocultured with fibroblasts, stimulated increased GAG synthesis, particularly hyaluronic acid.
The question of whether epithelial cells from
the rests of Malassez can produce factors that
enhance bone resorption was addressed by Birek
et al.'64 They cultured epithelial cells or their
conditioned media with mouse calvaria for 4 d,
causing significant increases in calcium release
from the bones. Indomethacin inhibited this resorptive effect only partially, suggesting that factors other than prostaglandins are synthesized by
the epithelial cells, which may account for the
osteolytic effects of the epithelial cells. The fact
that epithelial cells from the rests of Malassez
respond in vitro to 65tensile forces was demonstrated by Brunette. In his experiment the number of 3H-Trd labeled cells doubled after 2 h of
stretching. Moreover, stretched cells had a higher
volume of filamentous structures and more desmosomes per unit length of cell membrane than
unstretched cells.
The above review indicates that bone cells,
PDL fibroblasts, and epithelial cells from the rests
of Malassez respond readily to applied mechanical stresses. These cellular responses include
identifiable biochemical events that span the entire cellular domain. The plasma membrane, the
cytoplasmic organelles, the filamentous skeleton,
and the nucleus all seem to participate in the
physical-to-chemical transduction process. How-
ever, one of the main foci of attention in this
investigative field has been the role of prostaglandins in this process. The following section
briefly reviews this issue.
4. Prostaglandins and Force-Induced
Bone Remodeling
Since the report by Klein and Raisz'66 in 1970
that prostaglandins stimulate bone resorption in
tissue culture, numerous publications have implicated prostaglandins, particularly of the E series, in the response of bone cells to chemical
and mechanical stimuli. Out of the plethora of
investigations emerged a hypothesis, introduced
by Rodan and Martin,'67 which proposed that
osteoblasts regulate the resorptive activities of
osteoclasts. This hypothesis was based upon the
findings that osteoblasts carry receptors to all the
hormones involved in the maintenance of calcium
homeostasis, such as PTH, calcitonin, and vitamin D3. Osteoblasts respond to these endocrine
molecules, as well as to locally produced agents
such as growth factors, with elevations in cAMP
contents and prostaglandin synthesis. Thus, prostaglandins could serve as a stimulatory link or a
coupling factor between osteoblasts and osteoclasts. Applying tensile forces to cells derived
from mouse embryo calvaria, Binderman et al.168
stimulated the production of PGE2 and cAMP by
these cells. This effect was abolished by agents
that bind to membrane phospholipids (gentamicin
and antiphospholipid antibodies), and thus reduce their availability for enzymatic changes.
They concluded that mechanical forces exert their
effect on bone cells by the following chain of
events: activation of phospholipase A2, release
of arachidonic acid, increased PGE synthesis,
and elevated cAMP production. Taken together,
these observations assign to PGE2 a central role
in the regulation of force-induced bone cell activation. This is clearly an oversimplified concept
that may apply to cell culture conditions, in which
many factors found in the intact, living mammalian organism are absent. PGE was localized
immunohistochemically in the cat PDL by Davidovitch et al.169'170 The application of tipping
forces to cat canines for periods of time ranging
from 1 h to 14 d caused a significant increase in
426
the staining for PGE in PDL and alveolar bone

cells at sites of both tension and compression,
suggesting that PGE may indeed be involved in
the response of PDL and bone cells to mechanical
stress. However, as discussed below, other local
factors with bone cell stimulation capabilities have
been localized recently in the mechanically
stressed PDL, suggesting that prostaglandins may
be only one agent in a battery of factors that
regulate the response of cells to force.
The great emphasis in the literature on the
role of prostaglandins in the response of bone
cells to applied force prompted Yamasaki and his
associates to perform a series of experiments on
rats, monkeys, and humans, where PGE was injected into the gingiva near orthodontically treated
teeth.171-174 In rats,171 PGE, or PGE2 administration increased the number of osteoclasts in mechanically stressed alveolar bone within 12 h. In
monkeys,'72 the rate of tooth movement doubled
during 18 d of treatment. The drawback of this
experiment was the sample size, which consisted
of two subjects. Similar results were obtained in
a study on human patients. 173174 Here, PGE, was
injected monthly for 5 months into the gingiva
near orthodontically treated canines, resulting in
doubling of the rate of tooth movement. In explaining why they chose PGE administration as
an adjunct to orthodontic tooth movement, Yamasaki et al. stated that the rationale for the decision had been the reported evidence, primarily
from tissue-cell culture experiments, that implicated PGE2 in bone resorption. However, PGE2
has been shown to enhance metaphyseal bone
growth in young rats,175 and PGE1 infusion for
3 weeks enhanced alveolar bone growth in beagle
dogs.'76 Thus, in vivo, PGE may regulate bone
resorption and formation. To investigate this dual
role of PGE in bone as a stimulator of both resorption and formation, Nefussi and Baron'77 cultured 45Ca-labeled rat fetal long bones with PGE2
for 4 d. This treatment caused enhanced periosteal osteoblastic activity (in terms of percentage of osteoblastic surfaces), but increased osteoclastic resorption in medullary cavities. Thus,
the effect of PGE on bone cells may differ, depending on their location.
Pursuing this avenue further, Lee178 moved
teeth in rats by inserting an elastic band between
the first and second maxillary molars, according
to the method of Waldo and Rothblatt,31 for periods of 6 h to 5 d. In addition, the animals
received either local gingival injections of PGE1
twice daily, or a constant systemic administration
by a mini-osmotic pump. In both cases the number of alveolar bone osteoclastic lacunae in PDL
pressure sites was increased significantly, compared to non-PGEl-treated animals, but the effect
was more pronounced in the animals receiving
PGE, systemically. This finding raises the possibility of administering PGE, or PGE2 systemically during orthodontic treatment in an effort
to enhance the rate of tooth movement. However,
side effects of such a treatment must not be ignored. These effects may include diarrhea, vomiting, corneal congestion, and phlebitis.
Another feature of prostaglandins related to
bone remodeling has come to the foreground in
recent years. Prostaglandins have long been
known as being potent mediators of the inflammatory process in many tissues, including cartilage and bone. This fact led to the widespread
use of nonsteroidal anti-inflammatory drugs in
combatting rheumatoid arthritis'79'180 and jawbone destruction due to endodontic lesions'8' and
periodontal disease.182-'84 If tooth-moving forces
indeed cause an inflammatory reaction in the PDL,
then it should be expected that not only would
prostaglandins be found there, but other inflammatory mediators as well. The following sections
examine this issue.
V. REGULATION OF TOOTH MOVEMENT

BY INFLAMMATORY MEDIATORS
A. The Cells and Fluids of the
Periodontal Ligament
The PDL is a soft tissue envelope separating
the tooth from the alveolar bone. Like all other
connective tissues, it is comprised of cells and
extracellular matrix, which consists of collagen
and ground substance.'85 It contains an intricate
network of blood vessels and nerve endings, and
is very cellular. The majority of the PDL cells
are fibroblasts, but some of these fibroblast-like
cells are actually osteoprogenitor cells.65 Osteoblasts, either active or in the form of lining cells,
occupy the alveolar bone surface bordering the
427
PDL, while cementoblasts cover the dental root

surface that interfaces with the PDL. Clusters of
epithelial cells, the rests of Malassez, are spread
in the PDL in the vicinity of the root surface,
while capillaries are usually more numerous in
the center of the ligament and in the zone closer
to the alveolar bone. Cells migrating out of these
capillaries, such as lymphocytes, macrophages,
and mast cells, can be observed throughout the
PDL. Cells may also migrate into the PDL from
neighboring marrow spaces in the alveolar
bone.70,71
The PDL contains an elaborate network of
neural filaments'86 that arise from the trigeminal
nerve and send neural bundles through the apical
alveolar bone in a coronal direction to the gingiva
and PDL. Myelinated and unmyelinated fibers
are found in the PDL, some terminating as "free"
nerve endings, mostly in the inner part of the
PDL, while others terminate as knob-like enlargements or as coiled nerve endings. Unmyelinated fibers usually follow PDL blood vessels
and may have a vasomotor function. In this capacity, PDL nerve fibers may release, when
stressed mechanically, vasoactive peptides that
regulate movement of leukocytes out of
capillaries.
Nerve impulses resulting from tooth movement can be detected in afferent fibers. These
impulses originate primarily in the PDL and not
in the dental pulp, as shown in experiments where
the pulp had been removed.'87 Mechanical stimulation activates PDL fibers that are associated
with large-sized myelinated fibers. 188 The smaller
C fibers also react to mechanical forces, but with
a larger magnitude or longer duration.189 In forceinduced tooth movement, the PDL nerve fibers
perform two main functions: transmission of nociceptive impulses centrally and release of neuropeptides peripherally. The latter (discussed below) may have an important role in regulating
the local inflammatory response, primarily by
interacting with cells of the vascular system.
Examining the PDL in mouse molars, Freezer
and Sims'90 observed that 88% of the blood vessels consisted of venules and 12% were capillaries. Gould et al. 91 and McCulloch and
Melcher'92 found that 75 to 80% of the blood
vessels were positioned in the bony portion of
the PDL. To test the effect of orthodontic forces
(1 to 7 d) on the PDL vascular bed, Khouw and

Goldhaber'93 perfused dogs and monkeys with a
colloidal suspension of carbon particles. They
reported on dilation of vessels in both areas of
PDL tension and compression, in close proximity
to sites of alveolar bone apposition and resorp-
tion, respectively. Examination by TEM of se-
vere PDL compression sites in rats94 revealed

stasis and erythrocytic breakdown, with disintegration of vessel walls. However, this initial
response was followed by a repair process, typified by an invasion of the hyalinized zone by a
front of cellular and vascular elements. A markedly increased PDL vascularity was also detected
in the inflamed PDL of beagles by Jeffcoat et
al.195 using an angiographic method.
In addition to providing the PDL with a variety of leukocytes, the vascular system also contributes to its fluid composition. Bien'96 thoroughly analyzed the dynamics of PDL fluid in
relation to tooth movement, and identified three
sources of fluid in the PDL: cellular, vascular,
and interstitial. The latter is localized in the ground
substance and acts as a thixotropic gel, which is
jelly-like when not in motion, and flows quite
easily under pressure. When subjected to a steady
force, this fluid flows within the PDL out of areas
of compression and into areas of tension. This
fluid flow, which starts as soon as the force is
applied to a tooth and is maintained over extended periods of time, is apparently a crucial
step in the physicochemical behavior of the PDL.
This fluid motion and rearrangement signifies the
onset and progress of distortion of PDL cells and
fibers. This distortion of PDL, which is seen
microscopically as widening in areas of tension
and narrowing in sites of compression, may result
in the release of vasoactive neuropeptides, appearance of stress-generated potentials, and alterations of cellular shape. Storey27 observed vasodilation and migration of carbon particles out
of capillaries in stressed PDL within 20 min of
the application of an orthodontic force to guinea
pig incisors. Thus, orthodontic forces seem to
evoke an early response in the stressed PDL,
which encompasses fluids, matrix fibers and
ground substance, and cells.
The chain of events above-reported led us to
propose the following scheme to describe the initial effects of force on paradental tissues (Figure
428
4): (1) movement of fluids within the PDL; (2)

gradual distortion of the PDL; (3) generation of
streaming potentials; (4) alteration of cellular
shape and ion channel permeability; (5) neuropeptide release from nerve endings; (6) capillary
vasodilation and migration of leukocytes into extravascular areas; and (7) bending of the alveolar
bone and generation of piezoelectric spikes.
The outcome of such events is the introduction into the stressed PDL of agents derived from
cells of the nervous and immune systems. In addition, products of the endocrine system are routinely delivered to the PDL through the circulation. Thus, on the biochemical level, mechanical
forces can result in the simultaneous exposure of
PDL cells to signals from the nervous, immune,
and endocrine systems, leading to intricate and
fascinating interactions and cellular responses.
B. Interactions between Cells and

Products of the Nervous, Immune, and
Endocrine Systems
Recent attempts to understand the mode of
regulation of cellular activities in various tissues
have resulted in a rapidly growing volume of
evidence in support of the contention that interactions between cells of the nervous, immune,
and endocrine systems are pivotal parts of this
mechanism. In our own research we have focused
on the possibility of such interactions in the
stressed PDL between neurotransmitters, cytokines, and hormones. The neurotransmitters we
targeted are substance P (SP), vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP), and methionine enkephalin (ME).
The cytokines we chose are interleukin-lao and -
ORTHODONTIC FORCE
Movement of PDL fluids
Generation of streaming
potentials that affect PDL
Gradual Distortion of
PDL matrix and cells
1~~~~~~~\\^~
~and alveolar bone cells
Alteration of cellular
Neuropeptide release from
shape, cytoskeletal
configuration, and ion
channel permeability
PDL afferent nerve endings
Bending of the alveolar
Capillary vasodilation,
migration of leukocytes into
bone
Piezoelectric effects
extravascular areas
Synthesis and release of

cytokines, growth factors, PG's
FIGURE 4. Initial effects of orthodontic forces on
paradental tissues.
429
113 (IL-la and IL- 13), interleukin-2 (IL-2), tumor necrosis factor-a (TNF-a), and gamma interferon (IFN--y). The reason for choosing these
particular molecules was the existing evidence
implicating them in bone remodeling.
Incubation of human blood monocytes with
SP by Lotz et al.197 induced the release of IL-1,
TNF-a, and IL-6 by these cells, demonstrating
recognition of neurotransmitters by immune cells.
Similar effects were seen in B lymphocytes that
were stimulated to differentiate by SP,198 and in
neutrophils that were stimulated by SP to enhance
their oxidative metabolism. 199 ME had a biphasic
effect on the proliferation of peripheral blood
monocytes200 and stimulated 02 release by polymorphonuclear cells.201
Cells of the immune system were found to
synthesize neurotransmitter-like molecules. Substance P was extracted from mouse liver granulomas by Weinstock et al.,202 203 and its messenger RNA (mRNA) was localized to the granuloma
eosinophils by in situ hybridization. Roth et al.24
reported that the opioid precursor proenkephalin
was secreted by activated T helper cells. Cells
of the nervous system were reported to be reactive to products of immune cells.205 For instance, incubation of mouse anterior pituitary cells
with IL-1 induced protein phosphorylation, but
without cAMP elevations, which appears to be
an early signal for the secretion of 13-endorphin.
In another experiment, Fagarasan and Axelrod206
treated pituitary cells with IL-I in the presence
of norepinephrine or isoproterenol, causing an
additive effect on 3-endorphin secretion. Su et
al.207 found identical steroid receptors in guinea
pig brain and spleen, postulating that steroids
can, in this fashion, alter the immune function
and cause psychological changes.
C. Neuropeptides and Mineralized

Tissues
SP, a neuropeptide present in parts of the

CNS and PNS, such as C- type sensory fibers
and autonomic afferents and efferents, is released
from nerve endings in response to various stimuli. Its effects in peripheral tissues include vasodilation and an increase in capillary permeability. These effects may contribute to the plasma
extravasation and increased local blood flow that

accompany inflammation. Moreover, SP can
stimulate histamine release from mast cells at
sites of injury and inflammation. These biological capabilities turned SP into a suspected prime
contributor to the pathogenesis of rheumatoid arthritis. In 1984, Levine et al.208 caused adjuvant
arthritis in rats, and observed that the severity of
the disease was pronounced in joints that were
densely innervated by SP-containing afferent
neurons. Injection of SP into joints increased the
severity of arthritis. Lotz et al.209 incubated synoviocytes from human arthritic joints with SP,
causing increases in PGE2 and collagenase release. Yokoyama and Fujimoto210 demonstrated
that lymphocytes from rheumatoid arthritis patients could be activated by SP to present high
levels of HLA markers. Monocytes of these patients, when treated with SP, released high
amounts of oxygen intermediates.
A significant observation was made by
O'Bryne et al.,211 who injected IL-la into rabbit
knees and measured SP and PGE2 in the joint
fluid at 4, 24, and 48 h. By 4 h, SP was increased;
it further increased by 24 h and remained elevated
at 48 h. PGE2 levels were highest at 4 h and
remained elevated at 48 h. These results highlight
the intimate association, at sites of inflammation,
between cytokines, neurotransmitters, and prostaglandins. The neurogenic component of joint
inflammation was demonstrated by Lam and Ferrell, who in one experiment212 have inhibited carrageenan-induced knee joint inflammation in rats
by denervation, while in the other,213 abolished
it by an intraarticular injection of capsaicin, a SP
releaser and inhibitor.
Immunolocalization of SP in dental tissues
was first reported by Olgart et al.,214'215 who
observed its presence in the feline dental pulp.
In a more recent series of articles, Wakisaka et
al.216-2'8 reported on the distribution and origin
of SP in the rat molar pulp and PDL. They
observed SP-containing nerves along blood vessels, primarily in the middle and apical regions
of the PDL. During orthodontic tooth movement
in cats, intense immunohistochemical staining
for SP in PDL tension sites was seen by Davidovitch et al.219 within 1 h of treatment. Furthermore, administration of SP to human PDL
fibroblasts in vitro significantly increased the
430
concentration of cAMP in the cells and PGE2

in the medium within 1 min.219
Another neurotransmitter that has been implicated recently as a stimulator of bone resorption is VIP, a 28-amino acid residual peptide,
which was originally extracted from porcine duodenum.220 In vitro studies have demonstrated that
VIP stimulates bone resorption dramatically, and
that this activity is not mediated by PGE2.221
Moreover, this effect of VIP involves an 8- to
13-fold increase in bone cAMP levels.221 Binding
studies222 revealed high-affinity receptors for VIP
on human osteosarcoma cells. Hohmann et al.223
localized VIP in nerve fibers in the periosteum
of porcine rib, tibia, and vertebra. Here, VIP was
traced to sympathetic postganglionic neurons.
Herness224 localized it in mouse PDL, mainly in
the apical part around the blood vessels. During
orthodontic tooth movement in cats,22 intense
staining for VIP was localized in the compressed
PDL near sites of bone resorption and in the pulp
of moving teeth.
The recent discovery of CGRP by Rosenfeld
et al.226 raised the interest of investigators of neural
control of bone remodeling. This neurotransmitter was localized in small to medium diameter
sensory ganglion neurons and appeared to coexist
with SP.226 In the cat, local intraarterial infusion
of SP or CGRP caused a concentration-dependent
increase in nasal blood flow.227 The widespread
distribution of this 37-amino acid peptide has
been demonstrated in cardiovascular tissues of
several species, as well as in perivascular nerves
in the rat mesenteric artery.22 In arthritic patients, Larsson et al.229 detected CGRP-like immunoreactivity in the knee synovial fluid; however, similar amounts of CGRP were found in
synovial fluids of control patients. Recently, Silverman and Kruger230 localized CGRP in many
rat orofacial sensory structures, while Wakisaka
et al.231 studied its distribution in the feline dental
pulp. In the latter tissue CGRP-like immunoreactivity was observed in nerves along blood
vessels as well as in the subodontoblastic zone,
while dentinal injury in rat molars evoked sprouting of CGRP-containing nerve fibers.232 CGRP
was also localized in the rat PDL233 and mandibular periosteum.234 In the cat dental pulp, intraarterial infusion of SP and CGRP produced vasodilation, as measured by both laser Doppler
and 125I clearance.235 The effect of

CGRP was ten times larger when given after SP
than before it. Regarding an association between
CGRP and acute inflammation, Takahashi et al.236
found CGRP-containing nerves in contact with
dynorphin-containing dendrites in acutely inflamed rat hindpaw.
In relation to tooth movement, Kvinnsland
and Kvinnsland237 localized CGRP in the pulp
and PDL of rats receiving orthodontic forces to
maxillary molars for 5 d. In unstressed teeth,
CGRP immunoreactivity was localized primarily
in pulp and PDL nerves surrounding blood vessels. In moving teeth, the number of CGRP-containing nerves in both pulp and PDL increased,
and their staining intensified, particularly in PDL
tension sites. In these areas, dark "spots" were
seen, which were probably fibroblasts that have
bound CGRP released from stressed sensory nerve
endings. Such a pattern of cellular staining for
CGRP was observed in the pulp and PDL of
moving teeth in cats by Okamoto et al.238 in the
author's laboratory. In this group of 28 animals
that had been treated by a translatory force ap-
flowmetry
plication to one maxillary canine for 1 h and/or

2, 7, or 28 d, CGRP immunoreactivity in the
PDL intensified within 1 h in tension sites, but

was particularly intense in compression areas at
day 28, in PDL cells located at the edge of the
hyalinized zone and near osteoclasts (Figures 5
to 8). In the pulp of nonmechanically stressed
teeth, staining for CGRP was widespread and
distinct, localized in perivascular fibers and in
numerous fibroblasts (Figure 9). Force application to a tooth did not seem to alter the CGRP
immunoreactivity in dental pulp cells (Figure 10).
While SP appears to be intimately involved
in the transmission of painful sensations in the
primary afferent system, enkephalins are related
to morphine- and stimulation-produced analgesia. Both of these neuropeptides have been shown
immunohistochemically to have a similar distribution in many regions of the rat CNS.239 This
relationship between SP and ME could be of interest in investigating the regulation mechanism
of bone remodeling, since Jessell and Iversen240
have reported that opiate analgesics inhibit SP
release in the rat trigeminal nucleus. One group
recently focused on this issue.241-243 In one
experiment241 they measured ME levels in dental
431
FIGURE 5. Immunohistochemical localization of CGRP in unstressed cat PDL. Notice

dark staining of nerve fiber, but light staining of PDL cells (arrows). (Magnification x 1400.)
FIGURE 6. Localization of CGRP in PDL tension site after 1 h of application of a translatory

force to a cat maxillary canine. Notice dark staining over cells' periphery and cytoplasm
(arrows). (Magnification
1400.)
432
FIGURE 7. Dark staining for CGRP of cells in PDL tension zone after 28 d of translatory
force application to cat maxillary canine (arrows). (Magnification x 1400.)
FIGURE 8. Cells in compressed PDL at edge of hyalinized zone, stained very darkly for
CGRP, after 28 d of translatory force application to cat maxillary canine. (Magnification x
1400.)
433
FIGURE 9. Nerve fiber in unstressed cat canine pulp, intensely stained for CGRP, approaching blood vessel wall (B). (Magnification x 1400.)
FIGURE 10. Immunohistochemical localization of CGRP in pulp of cat canine subjected

to translator force for 28 d. Staining deposits are seen in nerve fiber approaching blood
vessel (B), as well as in adjacent cells. (Magnification x 1400.)
434
pulps of human teeth that had been moved by an

orthodontic spring for a few hours prior to extraction. They found drastic declines in the concentrations of ME in the moved teeth, perhaps
correlated with the early development of painful
sensations in orthodontic treatment. In another

study242 ME was extracted from pulpless, decalcified human third molars, suggesting that ME
is present in dentinal fibers that connect with the
dental pulp. In a more recent investigation Robinson et al.243 measured the concentrations of
P-endorphin, another opioid-like neuropeptide,
in premolars of 30 young patients whose premolars were to be extracted for orthodontic reasons. As in the case of ME, an acute mechanical
stress caused a gradual decrease in the level of
3-endorphin in the pulp. In an effort to determine
whether ME fluctuates in the PDL of moving
teeth, we attempted to localize it immunohistochemically in orthodontically treated teeth; however, only scarce and faint immunoreactivity for
ME could be detected in the unstressed PDL,
with a similar appearance in the PDL of the moving tooth (unpublished results).
D.
Cytokines and Mineralized Tissues

The evidence implicating cytokines in the
regulation of the activities of mineralized and

nonmineralized connective tissue cells is overwhelming. Following their initial discovery, cytokines were believed to be signal molecules produced by leukocytes, serving primarily as
communication links between cells of the immune system. However, many other cell types
were later found to synthesize cytokine-like molecules that could function in an autocrine or paracrine capacity. With respect to bone metabolism, cytokines with demonstrated or suspected
effects are IL-1, IL-2, IL-3, IL-6, TNF-a, and
IFN--y. Of these cytokines, the most potent stimulator of bone resorption in vitro is IL-1. It is
produced by many cell types, including
osteoblasts244.245 and chondrocytes.246 Secretion
of IL-1 is triggered by a variety of stimuli, including other cytokines and whole microorganisms. It has two distinct forms, IL-la and IL1p, which are coded by separate genes, but both
forms have similar biologic actions. In a recent
report Kaplan et al.247 listed the systemic effects

of IL-1. This long list encompasses the CNS, as
well as the vascular and immunologic constel-
lations. On the local level, IL-1 attracts leukocytes, stimulates fibroblast proliferation, and enhances bone resorption. It thus seems to be one
of the major components of the inflammatory
response.
Osteoblast-like cells, derived from human
trabecular bone, were incubated for 1 to 3 d with
various doses of IL-1 by Gowen et al.48 They
reported a significant increase in the uptake of
[3H] Tdr by these cells in comparison to controls,
and a marked increase in cell counts at day 3.
Infusion of IL-1 into normal mice by Boyce et
al.249 first resulted in a PGE2-related hypocalcemia after 3 h, followed by a hypercalcemia at
24 h. In another related experiment Sabatini et
al.250 infused IL-1 into mice, causing hypercalcemia at 72 h, with increased numbers of osteoclasts and bone resorption surfaces. Continuous
infusion of IL-1 for 14 d into rabbit knee joints
by Feige et al.251 induced severe arthritic changes.
In vitro, synovial fibroblasts were stimulated by
IL-1 to produce PGE2 and collagenase,252 and
Rafter253 proposed that polymorphonuclear leukocytes are the main source for IL-1 in arthritic
joints. However, in a recent interesting experiment Johnson et al.254 discovered that rat synovial
fibroblasts could be stimulated by lipopolysaccharides to produce and secrete IL-1, only following an initial exposure of the cells to IFN-y.
In samples of gingival cervicular fluid and in
gingival tissue, IL-la and IL-113 were detected
in patients with periodontal disease.255 Marked
reductions in IL-1 levels followed effective periodontal treatment.
In bone, target cells for IL-1 appear to be
osteoblasts, according to Thomson et al.,256 who
incubated neonatal mouse tibial osteoclasts with
human cortical bone and with IL-1 in the presence or absence of calvarial osteoblasts. Resorption of bone occurred only when osteoclasts and
osteoblasts were present. When fetal rat long
bones were incubated with IL-l a or IL-13p and
with PTH by Dewhirst et al.,257 a synergistic
effect on 45Ca release was recorded when both
cytokine and hormone were introduced simultaneously. Moreover, the presence of small concentrations of IL-1 necessitated only a very small
435
amount of PTH to cause a marked resorptive

effect. This finding is potentially very significant, because it means that in cases of induced
bone resorption, such as that occurring during
tooth movement, suboptimal amounts of key local mediators in the PDL may be sufficient to
evoke alveolar bone resorption, provided that
minute amounts of systemic bone-seeking hormones also happen to be present in the same site
at that time.
The effects of interleukins on bone resorption
were investigated by Gowen and Mundy258 using
the mouse calvarial assay system. They observed
marked stimulation of resorption by IL-1, but not
by IL-2. Tatakis et al.259 administered IL-1, IL2, or IL-3 to osteoblasts isolated from rat fetal
calvaria. Only IL-1 stimulated the release of large
amounts of PGE2 by the cells, and this effect was
augmented by PTH. However, when osteoclasts
were incubated with mouse calvaria in the presence of IL-2, their acid production increased.260
During tooth movement in cats261 we did not observe immunohistochemical staining for IL-2 in
the unstressed PDL, neither did we notice it in
PDL tension sites. However, after 7 and 14 d of
force application to teeth, intense staining for IL2 was observed in clusters of mononucleated cells
in PDL compression sites in close proximity to
osteoclasts and near the necrotic hyalinized zone.
The timing of the appearance of IL-2 immunoreactivity in the compressed PDL may coincide
with the localization of osteoclasts in Howship's
lacunae in the alveolar bone. Similar timing was
reported by Nieto-Sampedro and Chandy262 to
occur in injured rat brain, where IL-2 activity in
the tissue immediately adjacent to the injury
reached a peak 10 d postlesion. This peak coincided with axonal sprouting and astrocyte division. In tooth movement, IL-2 may thus be
associated with attraction and/or proliferation of
osteoclast progenitors, as well as with a stimulation of acid production by active osteoclasts.
TNF-a is another cytokine with well-documented potential as a stimulator of bone resorption. This 17-kDa molecule is synthesized mainly
by monocytes and macrophages, and is an im-
portant component of the inflammatory process.

It stimulates endothelial cells to secrete IL-1263
and increases their adherence to leukocytes.264 In
fetal mouse calvaria TNF-a increased the pro-
duction of procollagenase and the activity of collagenase.266 In human osteoblast-like cells TNFa inhibited proliferation and alkaline phosphatase
activity.267 This effect was not mediated by
cAMP, and, moreover, TNF-a reduced the cAMP
elevation caused by PTH.268 In terms of bone
resorbing potency, IL-1 is much stronger than
TNF-a; however, Stashenko et al.269 reported that
suboptimal concentrations of IL-1 3 or IL-la, in
combination with suboptimal concentrations of
TNF-a, stimulate the formation of osteoclast-like
cells in vitro from human marrow cells,270 and
this effect is synergistic when both agents are
added simultaneously to the cell cultures.
In living animals, TNF-a administration has
an inhibitory effect on bone fracture healing271
and it induces hypercalcemia.272 Using monoclonal antibodies, Hopkins and Meager273 detected low levels of TNF-a and INF-y in synovial
fluids of patients with rheumatoid arthritis, while
Maury and Teppo274 measured elevated levels of
TNF-a in the circulation of 46% of patients with
rheumatoid arthritis and 29% of patients with
lupus erythematosus. Using immunohistochemical staining, Yocum et al.275 localized TNF-a in
mononuclear cells of the joint lining layer, sublining, and perivascular areas. During tooth
movement in cats, intense cellular staining for
TNF-a was observed in the compressed PDL after
7 and 14 d of force application, particularly in
fibroblasts near alveolar bone osteoclasts. However, PDL cells in tension sites also contained
positive TNF-a immunoreactivity.
While IL-1, IL-2, and TNF-a have been found
to stimulate and enhance bone resorption in vivo
and in vitro, IFN-y was discovered to interfere
with resorptive processes. This cytokine is a lymphocytic product, which antagonizes the effects
of a number of growth factors. Gowen et al.276
reported that IFN-y completely abolished the resorptive effects of IL-1 and TNF-a in mouse
calvaria, but that it did not inhibit the resorptive
effects of PTH and vitamin D3. Direct effects of
IFN-y and TNF-a on human osteoblast-like cells
were demonstrated by Gowen et al.277 Cellular
proliferation and PGE2 production were stimulated by TNF-a, while alkaline phosphatase activity and osteocalcin release were inhibited;
however, the effects IFN-y had on bone cell activities were the polar opposite. In osteosarcoma
436
cells with an osteoblastic phenotype, both TNFa and IFN-y inhibited DNA synthesis, and their
inhibitory effect was even greater when their
administration was simultaneous.278 A similar inhibitory pattern on collagen synthesis was also
noted. In mice who received bone particle implants near their tibiae and seven daily injections
of IFN-y, macrophages that fused to form osteoclast-like cells increased in number, although IFNy inhibited the fusion of alveolar macrophages
in vitro, suggesting that IFN-y limits inflammation and favors tissue repair.279
In dental tissues, IFN-y was found by Melin
et al.280 to stimulate proliferation of human dental
pulp fibroblasts and to inhibit the synthesis of
type I and III collagen and of fibronectin. We
stained cat jaw sections for IFN-y,281 and found
practically no immunoreactivity in the unstressed
PDL or in PDL tension sites in moving teeth.
However, after 7 and 14 d of tooth movement,
numerous cells in PDL compression sites displayed positive staining for IFN--y. These cells
were located primarily near alveolar bone osteoclasts, suggesting that IFN-y is involved in the
regulation of bone resorption in vivo.
E. Neurotransmitters,
Tooth Movement
Cytokines, and
The above review suggests strongly that a

number of neurotransmitters and cytokines play
regulatory roles in the remodeling of mineralized
and nonmineralized connective tissues. Results
from experiments in the author's laboratory have
demonstrated that most of these signal molecules
can be localized immunohistochemically in cat
PDL and alveolar bone cells, and that their distribution and relative concentration are modified
when the tissues are stressed mechanically. Taken
together, these results imply that interactions between neurotransmitters, cytokines, and target
cells occur in paradental tissues during tooth
movement. However, localization of these molecules does not prove that some or all play active
roles in stimulating or inhibiting cells in mechanically stressed paradental tissues. To determine whether these signal molecules can evoke
a biochemical response by PDL fibroblasts, and
whether such stimulation can affect the activity
of bone-resorbing and bone-forming cells, we

resorted to cell and tissue culture experiments,
which are summarized briefly below.
VI. THE EFFECTS OF

NEUROTRANSMITTERS AND
CYTOKINES ON BONE AND PDL
FIBROBLASTS IN VITRO
In these experiments, (1) neonatal mouse calvaria were subjected to increasing concentrations
of neurotransmitters and cytokines to determine
the effect of 45C release or [3H] proline incorporation; (2) human PDL fibroblasts were incubated with neurotransmitters or cytokines to determine whether this treatment altered the
concentrations of cellular cAMP and PGE2; and
(3) conditioned media from cytokine-treated or
mechanically stressed PDL fibroblasts were tested
to determine whether they would enhance bone
resorption in vitro.
A. The Effects of Neurotransmitters and

Cytokines on Bone Resorption and
Formation In Vitro
In an effort to determine whether bone cells
can respond to a direct application of neurotransmitters or cytokines, we282 administered these
agents to neonatal mouse calvaria in vitro for a
24 h incubation period. IL-la and IL-1iI stimulated bone resorption more potently than other
cytokines. Bone formation was inhibited by PTH,
IL-la, IL-1p, and TNF-a, but not by IFN-y.
None of the neurotransmitters had any effect on
the rate of bone formation. These data demonstrate that cytokines and neurotransmitters that
have been found in the PDL during tooth movement can directly affect bone remodeling in vitro.
B. Effects of SP and Cytokines on PGE

and cAMP in PDL Fibroblasts
PDL fibroblasts, obtained from extracted
premolars of young orthodontic patients, were

removed enzymatically and incubated with SP (1
x 10-6 M) for 1 to 120 min.219 Significant in437
creases in the levels of cellular cAMP and PGE

in the medium occurred within 1 min, and persisted throughout the incubation period. Likewise, human PDL fibroblasts were incubated with
graded doses of IL-la, IL- 13, TNF-a, and IFNy283 for 15, 30, and 60 min, or 2, 4, 24, 48, and
72 h. The cells responded to all the cytokines
with dose- and time-related increases in the levels
of cAMP and PGE. These increases were inhibited by indomethacin. In another experiment284
cytokines were added to cell-containing media,
alone or in combination. The interactions between these cytokines varied in degree, depending on the particular combinations of cytokines.
Moreover, the administration of cytokine combinations was found to be additive, synergistic,
subtractive, or inhibitory on the production of
PGE, depending on the length of incubation time.
For instance, 24 h after the simultaneous administration of IL-1 p and TNF-a, the level of PGE
in the medium increased synergistically, but at
72 h it was lower than that produced by control
cells. In most of the time periods the addition of
IFN-y in the presence of IL-1 or TNF-a caused
a reduction in the level of PGE in the medium.
C. Bone Resorbing Activity Produced

Human PDL Fibroblasts
by
To determine whether cytokine-stimulated

PDL cells can enhance bone resorption, we285
administered IL- la, IL- 1 3, TNF-a, or IFN-y to
human PDL fibroblasts for a 1-h incubation. The
medium was then replaced with a fresh medium
for an additional 24 h incubation period, and the
latter medium (conditioned medium, CM) was
added to mouse calvaria containing media. CM
derived from unstimulated PDL fibroblasts increased the rate of bone resorption (45Ca release)
by 2.5-fold, as compared to control (no CM)
resorptive rate. CM derived from IL- lp-stimulated PDL fibroblasts caused a rate of resorption
3.5 times greater than control. Resorption in the
presence of CM obtained from cells stimulated
by IFN-y or IL- 13 + IFN--y was at the same
level as that caused by CM of unstimulated cells.
Indomethacin administration to the calvaria-containing medium abolished the synthesis of PGE,
but reduced only partially the enhanced resorp-
tion rate caused by the IL-1 CM. These results

suggest that PDL cells produce nonprostaglandin
bone-resorbing factor(s), in addition to PGE2.
The addition of anti-IL-lp monoclonal antibodies to the CM failed to inhibit the resorptive effect
on the calvaria, suggesting that the resorptive
factor produced by PDL fibroblasts was not IL1 3 (unpublished results). In another recent
study286 we found that the resorptive effects of
CM derived from PDL fibroblasts stimulated by
either IL-la, IL-IL, or TNF-a could be augmented by the addition of PTH (1 U/ml) to the
CM. This observation suggests that extensive resorption of bone may result from products of
cytokine-stimulated PDL fibroblasts, when a systemic hormone such as PTH is also present in
the vicinity of the bone target cells.
D. Interactive Effects of IL-1, and

Mechanical Stress in PDL Fibroblasts
Human PDL fibroblasts were stretched in viby the use of convex templates for 2 to 60
min.'59 The stretching caused significant elevations in PGE and cAMP within 15 min; however,
the addition of IL-Il caused synergistic, additive, or even subtractive effects, depending on
the duration of the incubation time. In another
recent study287 CM derived from PDL cells stimulated by mechanical stress in the presence of
IL-1 caused significantly more bone resorption
than CM obtained from cells that had been treated
by either factor alone. Again, indomethacin inhibited PGE synthesis, but reduced bone resorption only partially.
Taken together, the results of the experiments
summarized in this section demonstrate that human PDL fibroblasts are sensitive to mechanical
stress, as well as to a variety of chemical signals
derived from the nervous, immune, and endocrine systems. If levels of cellular cAMP or PGE
in the medium are used as indicators, or if the
resorptive activity of CM derived from stimulated
or unstimulated PDL cells is used as such, it
becomes clear that when two or more of these
signals, mechanical or chemical, are presented
to the PDL cell simultaneously, their response
would usually differ from that seen following the
application of one signal alone. Thus, it may be
tro
438
possible to observe synergistic, additive, or subtractive effects. In the in vivo situation there are
usually several factors present simultaneously in
the cell's environment. Therefore, it may be reasonable to assume that in tooth movement PDL
fibroblasts do not respond merely to tension or
compression, but also to other signals; products
of neighboring cells; members of other systems,
such as endothelial cells, monocytes, and nerve
cells; and cells of the epithelial rests of Malassez.
Moreover, it seems likely that prostaglandins are
not the sole link between PDL fibroblasts and
bone cells; however, the identity of the other
connecting molecules is unknown at the present
time, and deserves further probing.
VII. TOOTH MOVEMENT: CONCLUSIONS

AND DIRECTIONS FOR FUTURE
RESEARCH
For at least 2000 years it has been known
that a tooth can be moved gradually from one
spot in the oral cavity to a more desirable one
by the application of mechanical forces to the
tooth's crown. In the middle of the 18th century,
John Hunter explained that this movement is an
outcome of the habit of the bone "to move out
of the way of pressure". About 100 years later,
John Farrar attributed force-induced tooth movement to "decalcification of the root socket and
bending of the alveolar bone". In the years since,
extensive investigations have confirmed that indeed physical and chemical alterations occur concomitantly in the paradental tissues, resulting in
cellular activities that culminate in tissue remodeling and tooth movement.
Histologic studies made it clear that tissues
can be remodeled only by the action of cells. In
the case of the PDL, the cells that form and
degrade the periodontal extracellular matrix are
primarily the fibroblasts. In the case of the alveolar bone, the cells that remodel it are the osteoblasts, osteoclasts, and osteocytes. When the
PDL and alveolar bone are stressed by continuously applied forces, both cells and matrix are
distorted and extracellular fluids are mobilized.
Bone matrix distortion in vivo is associated with
reorientation of its proteoglycans, a phenomenon
that may serve as "strain memory", as the dis-
torted molecules return slowly to their original

configuration. This strain-related distortion is also
associated with the appearance of piezoelectric
spikes, while fluid flow leads to slow-dissipating
streaming potentials. These bioelectric phenomena can cause alterations in the polarity of the
plasma membranes of cells, leading to activation
of membrane enzymes and cell-matrix interactions. Stress-caused changes in the shape of cells
can result in the crystallization of cytoskeletal
filaments and opening or closing of stress-related
membrane ion channels. Thus, fibroblasts and
bone cells can respond, in vivo and in vitro, to
the distortive effects of mechanical stress, which
are expressed both in the cells and their surrounding matrix. Significantly, bone cells appear to be
sensitive to short-duration exposures to mechanical loads whose distribution in the bone matrix
deviates from their regular dispersion pattern. This
finding may suggest that properly designed force
systems may obviate the need for prolonged periods of force application, as is customary in most
of the prevalent orthodontic techniques. However, while such brief loads are capable of evoking osteoblastic activity, it is not evident that
resorptive functions can also be stimulated by
these strains. The latter may perhaps necessitate
the application of prolonged strains, particularly
those that cause periodontal injury. Thus, from
the orthodontic standpoint, a clinically effective
force should be somewhat biodisruptive, i.e., be
capable of causing an inflammatory/reparative
reaction in the PDL and alveolar bone. Osteoclastic activity is quite intense in such a case,
and it is the activity of these multinucleated cells
that removes the alveolar bone that stands in the
way of the moving teeth.
The applied mechanical stress causes gradual
fluid shifts within the PDL, resulting in nerve
fiber distortion, which leads to the release of
neuropeptides from nerve endings. Some of these
neuropeptides have vasoactive properties, and
their interaction with PDL capillaries causes vasodilation, plasma extravasation, and migration
of leukocytes from the capillaries into the extravascular spaces of the PDL. These migratory leukocytes synthesize and secrete a variety of cytokines capable of stimulating fibroblasts,
endothelial cells, and alveolar bone cells. The
PDL fibroblasts are responsive to the neuropep439
tides, cytokines, and several growth factors produced by endothelial cells and bone cells. These
fibroblasts, in areas of tension, proliferate and
synthesize new matrix components, and in areas
of compression they degrade the necrotic PDL.
However, in both sites of tension and compression, the fibroblasts seem to produce factors that
activate neighboring bone cells. Recent evidence
suggests that osteoclasts are regulated by factors
derived from adjacent osteoblasts, and PGE2 was
proposed as being a major part of this bridge.
However, in vitro experiments with PDL fibroblasts and mouse calvaria have demonstrated that
factors produced by unstimulated or stimulated
(mechanically or chemically) PDL fibroblasts can
markedly enhance the rate of bone resorption.
Thus, in tooth movement, PDL fibroblasts may
not only be responsible for the remodeling of the
periodontal matrix, but may also be actively involved in the regulation of the activity of the cells
that remodel the alveolar bone. Osteocytes also
seem to be sensitive to applied loads, and it was
suggested that these cells, which are capable of
recognizing and responding to molecular reorientation in their surrounding matrix, communicate these alterations to bone surface cells (primarily osteoblasts), providing them with an
osteogenic stimulus.
Orthodontic tooth movement is based predominantly on the application of mechanical
forces to teeth in a judicious fashion supported
by biomechanical principles. The rationale for
investigating associated biological phenomena is
derived from the desire to gain a thorough insight
into these events in order to determine whether
our clinical means to move teeth are effective
and unharmful. Furthermore, this rationale is derived from our everlasting quest to improve our
clinical approaches and from the realization that
such progress can be derived from increased
knowledge of biological principles on the tissue,
cellular, and molecular levels.
Many questions remain, however, with the
biochemical and physical systems, only partly
understood, but certainly deserving much further
investigation. Among these issues are the effects
of mechanical stresses on cells of the epithelial
rests of Malassez, and on alveolar bone marrow
spaces and the interaction of these cells with cells
of the PDL. Another poorly understood phenom-
enon is the tooth movement-related dental root

resorption. Above all remains our inability to
predict the nature and pattern of the individual
biological response to tooth moving forces. It
seems likely that new information on these issues
will be derived from experiments that correlate
and integrate in vivo and in vitro systems and
findings. The ability to maintain and challenge
human PDL fibroblasts in vitro offers an opportunity to thoroughly examine one of the major
cell types that is directly affected by orthodontic
forces. Similar approaches can be applied to other
PDL cell types, i.e., endothelial and epithelial
cells. Interactions between these cell types and
bone cells remain an intriguing issue within the
framework of force-induced cellular activation
and tissue remodeling. Ultimately, further explorations in this area should elucidate the cellular/molecular basis of tooth movement, and enable clinicians to include biological considerations
in planning treatment for individual patients.
ACKNOWLEDGMENT
Supported by grant number DE-08428 from

Institute of Dental Research.
National
the
REFERENCES
Weinberger, B. W., Orthodontics, An Historical
Review of its Origin and Evolution, C. V. Mosby,
St. Louis, 1926, 54.
2. Weinberger, B. W., Orthodontics, An Historical
Review of its Origin and Evolution, C. V. Mosby,
St. Louis, 1926, 138.
3. Hunter, J., The Natural History of the Human Teeth,
J. Johnson, London, 1778, 99.
4. Delabarre, C. F., Odontologie ou Observations sans
les Dents Humaines, Paris, 1815.
5. Farrar,J. N., Treatise on Irregularities of the Teeth
and Their Correction, Vol. 2, DeVinne Press, New
York, 1888, 758.
6. Wolff, J. D., Das Gesetz der Transformation der
Knochen, Hirshwald, Berlin, 1982.
7. Sandstedt, C., Einige beitrage zur theori der zahnregulierung, Nord. Tandlaeg, Tidskr., 5, 236, 1904.
8. Sandstedt, C., Einige beitrage zur theori der zahnregulierung, Nord. Tandlaeg, Tidskr., 6, 1, 1905.
9. Oppenheim, A., Tissue changes, particularly of the
bone, incident to tooth movement, Trans. Eur. Orthodont. Soc., 8, 11, 1911.
1.
440
Schwarz, A. M., Tissue changes incidental to orthodontic tooth movement, Int.J. Orthodont., 18, 331,
1932.
11. Fukada, E. and Yasuda, I., On piezoelectric effect
of bone, J. Phys. Soc. Jpn., 12, 1158, 1957.
12. Gillooly, C. J., Hosley, R. T., Mathews, J. R.,
and Jewett, D. L., Electric potentials recorded from
mandibular alveolar bone as a result of forces applied
to the tooth, Am. J. Orthodont., 54, 649, 1968.
13. Baumrind, S., A reconsideration of the propriety of
the "pressure-tension" hypothesis, Am. J. Orthodont., 55, 12, 1969.
14. Reitan, K., The initial tissue reaction incident to
orthodontic tooth movement as related to the influence of function, Acta Odont. Scand., Suppl. 6, 1951.
15. Reitan, K., Tissue reaction as related to the age
factor, Dent. Rec., 74, 271, 1954.
16. Reitan, K., Experiments of rotation of teeth and their
subsequent retention, Trans. Eur. Orthodont. Soc.,
34, 124, 1958.
17. Reitan, K., Behavior of Malassez' epithelial rests
during orthodontic tooth movement, Acta Odont.
Scand., 19, 425, 1961.
18. Reitan, K., Effects of force magnitude and direction
of tooth movement on different alveolar bone types,
Angle Orthodont., 34, 244, 1964.
19. Reitan, K., Initial tissue response behavior during
apical tooth resorption, Angle Orthodont., 44, 68,
1974.
20. Reitan, K. and Kvam, E., Comparative behavior
of human and animal tissue during experimental tooth
movement, Angle Orthodont., 11, 1, 1971.
21. Storey, E. and Smith, R., Force in orthodontics and
its relation to tooth movement, Aust. J. Dent., 56,
11, 1952.
22. Smith, R. and Storey, E., The importance of force
in orthodontics, Aust. J. Dent., 56, 291, 1952.
23. Storey, E., Bone changes associated with tooth
movement: the influence of the menstrual cycle on
the rate of tooth movemement, Aust. J. Dent., 58,
80, 1954.
movement: a histological study of the effect of force
in the rabbit, guinea pig and rat, Aust. J. Dent., 59,
147, 1955.
movement: a histological study of the effect of force
for varying durations in the rabbit, guinea pig and
rat, Aust. J. Dent., 59, 209, 1955.
movement: a histological study of the effect of age
and sex in the rabbit and guinea pig, Aust. J. Dent.,
59, 220, 1955.
27. Storey, E., The nature of tooth movement, Am. J.
Orthodont., 63, 292, 1973.
28. Deguchi, T. and Mori, M., Histochemical observations on oxidative enzymes in periodontal tissue
during experimental tooth movement in rat, Arch.
Oral Biol., 13, 49, 1968.
10.
29.
Takimoto, K., Deguchi, T., and Mori, M., Histochemical detection of succinic dehydrogenase in
osteoclasts following experimental tooth movement,

J. Dent. Res., 45, 1473, 1966.
Takimoto, K., Deguchi, T., and Mori, M., Histochemical detection of acid and alkaline phosphatase
in periodontal tissue following experimental tooth
movement, J. Dent. Res., 47, 340, 1968.
31. Waldo, C. M. and Rothblatt, J. M., Histologic
response to tooth movement in the laboratory rat:
procedure and preliminary observation, J. Dent. Res.,
33, 481, 1954.
32. Lilja, E., Lindskog, S., and Hammarstrom, L.,
Histochemistry of enzymes associated with tissue
degradation incident to orthodontic tooth movement,
Am. J. Orthodont., 83, 62, 1983.
33. Lilja, E., Bjornestedt, T., and Lindskog, S., Cellular enzyme activity associated with tissue degradation following orthodontic tooth movement in man,
Scand. J. Dent. Res., 91, 381, 1983.
34. Kurihara, S. and Enlow, D. H., A histochemical
and electron microscopic study of an adhesive type
of collagen attachment on resorptive surfaces of alveolar bone, Am. J. Orthodont., 77, 532, 1980.
35. Martinez, R. H. and Johnson, R. B., Effects of
orthodontic tooth movement on the alcian blue staining patterns of rat alveolar bone: a histochemical study,
Histol. Histopathol., 3, 39, 1988.
36. Noda, K., Localization of cytochrome c oxidase activity in osteoclasts treated with calcitonin during experimental tooth movement, Acta Histochem. Cytochem., 21, 301, 1988.
37. Rygh, P. and Selvig, K. A., Erythrocytic crystallization in rat molar periodontium incident to tooth
movement, Scand. J. Dent. Res., 81, 62, 1973.
38. Ten Cate, A. R., Deporter, D. A., and Freeman,
E., The role of the fibroblasts in the remodeling of
periodontal ligament during physiological tooth
movement, Am. J. Orthodont., 69, 155, 1976.
39. Ten Cate, A. R., Freeman, E., and Dickinson,
J. B., Sutural development: structure and its response
to rapid expansion, Am. J. Orthodont., 71,622, 1977.
40. Rygh, P., Elimination of hyalinized periodontal tissues associated with orthodontic tooth movement,
Scand. J. Dent. Res., 82, 57, 1974.
41. Rygh, P., Ultrastructural changes in tension zone of
rat molar periodontium incident to orthodontic tooth
movement, Am. J. Orthodont., 70, 269, 1976.
42. Edwards, J. E., A study of the periodontium during
orthodontic rotation of teeth, Am. J. Orthodont., 54,
441, 1968.
43. Martinez, R. H. and Johnson, R. B., Effects of
orthodontic forces on the morphology and diameter
of Sharpey fibers on the alveolar bone of the rat,
Anat. Rec., 219, 10, 1987.
44. Kurihara, S. and Enlow, D. H., An electron microscopic study of attachments between periodontal
fibers and bone during alveolar remodeling, Am. J.
Orthodont., 77, 516, 1980.
30.
441
45.
Kvam, E., Scanning electron microscopy of tissue
changes on the pressure surface of human premolars

following tooth movement, Scand. J. Dent. Res., 80,
ulated
61.
357, 1972.
46. Barber, A. F. and Sims, M. R., Rapid maxillary
expansion and external root resorption in man: a scanning electron microscope study, Am. J. Orthodont.,
79, 630, 1981.
47.
Langford, S. R. and Sims, M. R., Root surface

resorption, repair, and periodontal attachment following rapid maxillary expansion in man, Am. J.
Orthodont., 81, 108, 1982.
Rygh, P., Orthodontic root resorption studied by

electron microscopy, Angle Orthodont., 47, 1, 1977.
49. Nakane, S. and Kameyama, Y., Root resorption
caused by mechanical injury of the periodontal soft
48.
tissue in rats, J. Periodont Res., 22, 390, 1987.
Goldie, R. S. and King, G. J., Root resorption and

tooth movement in orthodontically treated, calciumdeficient, and lactating rats, Am. J. Orthodont., 85,
424, 1984.
51. Engstrom, C., Granstrom, G., and Thilander, B.,
Effect of orthodontic force on peridontal tissue metabolism, Am. J. Orthodont. Dentofacial Orthoped.,
93, 486, 1988.
52. Garant, P. R. and Cho, M. I., Cytoplasmic polarization of periodontal ligament fibroblasts. Implications for cell migration and collagen secretion, J.
50.
53.
54.
55.
56.
57.
58.
59.
60.
Periodont. Res., 14, 95, 1979.

Garant, P. R., Cho, M. I., and Cullen, M. R.,
Attachment of periodontal ligament fibroblasts to the
extracellular matrix in the squirrel monkey, J. Periodont. Res., 17, 70, 1982.
Cho, M. I. and Garant, P. R., Mirror symmetry
of newly divided rat periodontal ligament fibroblasts
in situ and its relationship to cell migration, J. Periodont. Res., 20, 185, 1985.
Garant, P. R. and Cho, M. I., Fibroblast migration,
cytoplasmic polarity, and matrix secretion in the periodontal ligament, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC
Press, Boca Raton, FL, 1989, 29.
Willingham, M. C., Yamada, K. M., Yamada,
S. S., Pouyssegur, J., and Pastan, I., Microfilament bundles and cell shape are related to adhesiveness to substratum and are dissociable from growth
control in cultured fibroblasts, Cell, 10, 375, 1977.
McCulloch, C. A. G., Barghava, U., and Melcher,
A. H., Cell death and the regulation of populations
of cells in the periodontal ligament, Cell Tissue Res.,
255, 129, 1989.
Kvam, E., Cellular dynamics on the pressure side
of the rat periodontium following experimental tooth
movement, Scand. J. Dent. Res., 80, 369, 1972.
Roberts, W. E. and Jee, W. S. S., Cell kinetics of
orthodontically-stimulated periodontal ligament in the
rat, Arch. Oral Biol., 19, 17, 1974.
Roberts, W. E., Chase, D. C., and Jee, W. S. S.,
Counts of labeled mitoses in the orthodontically stim-
periodontal ligament of the rat, Arch. Oral
Biol., 19, 665, 1974.
Roberts, W. E., Cell kinetic nature and diurnal periodicity of the rat periodontal ligament, Arch. Oral
Biol., 20, 456, 1975.
Roberts, W. E., Aubert, M. M., Sparaga, J. A.,

and Smith, R. K., Circadian periodicity of cell kinetics of rat molar periodontal ligament, Am. J. Orthodont., 76, 316, 1979.
63. Smith, R. K. and Roberts, W. E., Cell kinetics of
the initial response of orthodontically induced osteogenesis in rat molar periodontal ligament, Calcified
Tissue Int., 30, 51, 1980.
64. Roberts, W. E. and Chase, D. C., Kinetics of cell
proliferation and migration associated with orthodontically-induced osteogenesis, J. Dent. Res., 60,
174, 1981.
65. Roberts, W. E., Klinger, E., and Mozsary, P. G.,
Circadian rhythm of mechanically mediated differentiation of osteoblasts, Calcified Tissue Int., 36,
S63, 1984.
66. Roberts, W. E. and Ferguson, D. J., Cell kinetics
of the periodontal ligament, in The Biology of Tooth
Movement, Norton, L. A. and Burstone, C. J., Eds.,
CRC Press, Boca Raton, FL, 1989, 55.
67. Yee, J. A., Kimmel, D. B., and Jee, W. S. S.,
Periodontal ligament cell kinetics following orthodontic tooth movement, Cell Tissue Kinet., 9, 293,
1976.
68. Yee, J. A., Response of periodontal ligament cells
to orthodontic force: ultrastructure identification of
proliferating fibroblasts, Anat. Rec., 194, 603, 1979.
69. Roberts, W. E. and Cox, W. B., Morphometric
quantification of nuclear volume changes during osteoblast histogenesis, in Bone Histomorphometry,
Meunier, P. J., Ed., Armour Montagu, Paris, 1977,
267.
70. McCulloch, C. A. G., Nemeth, E., Lowenberg,
B., and Melcher, A. H., Paravascular cells in endosteal spaces of alveolar bone contribute to periodontal ligament cell populations, Anat. Rec., 219,
233, 1987.
71. McCulloch, C. A. G. and Heersche, J. N. M.,
Lifetime of the osteoblast in mouse periodontium,
Anat. Rec., 222, 129, 1988.
72. Morris, C. E., Mechanosensitive ion channels, J.
Memb. Biol., 113, 93, 1990.
73. Brezden, B. L., Gardner, D. R., and Morris,
C. E., A potassium-selective channel in isolated
Lymnaea stagnalis heart muscle cells, J. Exp. Biol.,
62.
123, 175, 1986.
74.
Moody, W. J. and Bosma, M. M., A nonselective

cation channel activated by membrane deformation
in oocytes of the ascidian Boltenia Villosa, J. Membr.
75.
Biol., 107, 179, 1989.

Sachs, F., Mechanical transduction in biological systems, Crit. Rev. Biomed. Eng., 16, 141, 1988.
76.
Christensen, O., Mediation of cell volume regula-
442
tion
by Ca2+
influx
through stretch-activated chan-
nels, Nature, 330, 66, 1987.

77. Cooper, K. E., Tang, J. M., Rae, J. L., and
Eisenberg, R. S., A cation channel in frog lens epithelia responsive to pressure and calcium, J. Membr.
Biol., 93, 259, 1986.
78. Lansman, J. B., Hallam, T. J., and Rink, T. J.,
79.
80.
81.
82.
83.
84.
85.
86.
87.
88.
89.
90.
Single stretch-activated ion channels in vascular endothelial cells as mechanotransducers, Nature, 325,
811, 1987.
Morris, C. E. and Sigurdson, W. J., Stretch-inactivated ion channels coexist with stretch-activated
ion channels, Science, 243, 807, 1989.
Harris, A. K., Wild, P., and Stopak, D., Silicone
rubber substrata: a new wrinkle in the study of cell
locomotion, Science, 208, 177, 1980.
Ingber, D. E. and Folkman, J., Tension and
compression as basic determinants of cell form and
function: utilization of a cellular tensegrity mechanism, in Cell Shape: Determinants, Regulation, and
Regulatory Role, Stein, W. D. and Bronner, F., Eds.,
Academic Press, San Diego, CA, 1989, 3.
Vanderburgh, H. H. and Kaufman, S., Stretchinduced growth of skeletal myotubes correlates with
activation of the sodium pump, J. Cell Physiol., 109,
205, 1981.
Cogoli, A., Valluchi-Morf, M., Mueller, M., and
Briegleb, W., Effect of hypogravity on human lymphocyte activation, Aviat. Space Environ. Med., 51,
29, 1980.
Leung, D. Y. M., Glagov, S., and Mathews, M. B.,
Cyclic stretching simulates synthesis of matrix components by arterial smooth muscle cells in vitro, Science, 191, 475, 1976.
Quinn, R. S. and Rodan, G. A., Enhancement of
ornithine decarboxylase and Na+, K+ ATPase in osteosarcoma cells by intermittent compression,
Biochem. Biophys. Res. Commun., 100, 1696, 1981.
Ingber, D. E. and Jamieson, J. D., Cells as tensegrity structures: architectural regulation of histodifferentiation by physical forces transduced over
basement membrane, in Gene Expression During
Normal and Malignant Differentiation, Andersson,
L. C., Gahmberg, C. G., and Ekblom, P., Eds.,
Academic Press, Orlando, FL, 1985, 13.
Joshi, H. C., Chu, D., Buxbaum, R. E., and
Heidermann, S. R., Tension and compression in the
cytoskeleton of PC 12 neurites, J. Cell Biol., 101,
697, 1985.
Herman, B. and Pledger, W. J., Platelet-derived
growth factor-induced alterations in vinculin and actin distribution in BALB/c-3T3 cells, J. Cell Biol.,
100, 1031, 1985.
Berezney, R. and Coffey, D. S., Nuclear protein
matrix: association with newly synthesized DNA,
Science, 189, 291, 1975.
Pardoll, D. M., Vogelstein, B., and Coffey, D. S.,
A fixed site of DNA replication in eucaryotic cells,
Cell, 19, 527, 1980.
Barrak, E. R., and Coffey, D. S., Biological properties of the nuclear matrix: steroid hormone binding,
Recent Prog. Hormone Res., 38, 133, 1982.
92. Nicolini, C., Belmont, A. S., and Martelli, A.,
Critical nuclear DNA size and distribution associated
with S phase initiation, Cell Biophys., 8, 103, 1986.
93. Jiang, L.-W. and Schindler, M., Nuclear transport
in 3T3 fibroblasts: effects of growth factors, transformation and cell shape, J. Cell Biol., 106, 13,
1988.
94. Emerman, J. T. and Pitelka, D. R., Maintenance
and induction of morphological differentiation in associated mammary epithelium on floating collagen
membranes, In Vitro, 13, 316, 1977.
95. Lanyon, L. E., Goodship, A. E., Pye, C. J., and
MacFie, J. H., Mechanically adaptive bone remodeling, J. Biomech., 15, 767, 1982.
96. Rubin, C. T. and Lanyon, L. E., Regulation of
bone formation by applied dynamic loads, J. Bone
Jt. Surg., 66-A, 397, 1984.
97. Lanyon, L. E. and Rubin, C. T., Static vs. dynamic
loads as an influence on bone remodeling, J. Biomech., 17, 897, 1984.
98. Rubin, C. T. and Lanyon, L. E., Regulation of
bone mass by mechanical strain magnitude, Calcified
Tissue Int., 37, 411, 1985.
99. Lanyon, L. E., Rubin, C. T., and Baust, G., Modulation of bone loss during calcium insufficiency by
controlled dynamic load, Calcified Tissue Int., 38,
209, 1986.
100. Rubin, C. T. and Lanyon, L. E., Osteoregulatory
nature of mechanical stimuli: function as a determinant for adaptive remodeling in bone, J. Orthoped.
Res., 5, 300, 1987.
101. Lanyon, L. E., Functional strain in bone tissue as
an objective, and controlling stimulus for adaptive
bone remodeling, J. Biomech., 20, 1083, 1987.
102. Pead, M. J., Suswillo, R., Skerry, T. M., Vedi,
S., and Lanyon, L. E., Increased 3H-uridine levels
in osteocytes following a single short period of dynamic bone loading in vivo, Calcified Tissue Int., 43,
92, 1988.
103. Skerry, T. M., Bitensky, L., Chayen, J., and
Lanyon, L. E., Loading-related reorientation of bone
proteoglycan in vivo. Strain memory in bone tissue?,
J. Orthoped. Res., 6, 547, 1988.
104. Skerry, T. M., Suswillo, R., El Haj, A. J., Ali,
N. N., Dodds, R. A., and Lanyon, L. E., Loadinduced proteoglycan orientation in bone tissue in
vivo and in vitro, Calcified Tissue Int., 46, 318, 1990.
105. Smith, E., Reddan, W., and Smith, P., Physical
activity and calcium modalities for bone mineral increased in aged women, Med. Sci. Sports Exer., 13,
60, 1980.
106. Smith, E. L., Smith, P. E., Ensign, C. J., and
Shea, M. M., Bone involution decrease in exercising
middle-aged women, Calcified Tissue Int., 36, S 129,
91.
1984.
107. Mack, P. B., LaChance, P. A., Vose, G. P., and
443
Vogt, F., Bone demineralization of foot and hand of

Gemini-Titan IV, V, and VII astronauts during orbital
flight, Am. J. Roentgenol., 100, 503, 1967.
108. Cann, C. E. and Adachi, R. R., Bone resorption
and mineral excretion in rats during space flight, Am.
J. Physiol., 244, R327, 1983.
109. Rambaut, P. C. and Goode, A. W., Skeletal changes
during space flight, Lancet, p. 1050, 9, November
1985.
110.
Bikle, D. D., Halloran, B. P., Cone, C. M., Globus,

R. K., and Morey-Holton, E., The effect of sim-
ulated weightlessness on bone maturation, Endocrinology, 120, 678, 1987.

111. Yamaguchi, M., Ozaki, K., and Hoshi, T., Simulated weightlessness and bone metabolism: decrease
of protein and DNA synthesis in the femoral diaphysis
of rats, Res. Exp. Med., 189, 331, 1989.
112. Glucksmann, A., The role of mechanical stresses in
bone formation in vitro, J. Anat., 76, 231, 1942.
113. Functional Adaptation in Bone Tissue, A Kroc Foundation Conference, July 1983, Cowin, S. C., Lanyon, L. E., and Rodan, G., Eds., Calcified Tissue
Int., 36(Suppl. 1), 1, 1984.
114. Pienkowski, D. and Pollack, S. R., The origin of
stress-generated potentials in fluid saturated bone, J.
Orthoped. Res., 1, 30, 1983.
115. Pollack, S. R., Salzstein, R., and Pienkowski, D.,
The electric double layer in bone and its influence
on stress-generated potentials, Calcified Tissue Int.,
36, S77, 1984.
116. Marino, A. A., Spadaro, J. A., Fukada, E., Kah,
L. D., and Becker, R. O., Piezoelectricity in collagen films, Calcified Tissue Int., 31, 257, 1970.
117. Fukada, E. and Yasuda, I., Piezoelectric effects in
collagen, Jpn. J. Appl. Phys., 3, 117, 1964.
118. Marino, A. A. and Becker, R. O., Piezoelectricity
in hydrated frozen bone and tendon, Nature, 253,
627, 1975.
119. Marino, A. A. and Becker, R. O., The origin of
the piezoelectric effect in bone, Calcified Tissue Int.,
8, 177, 1971.
120. Bassett, C. A. L. and Becker, R. O., Generation
of electric potentials by bone in response to mechanical stress, Science, 137, 1063, 1962.
121. Cochran, G. V. B., Pawluk, R. J., and Bassett,
C. A. L., Stress generated electric potentials in the
mandible and teeth, Arch. Oral Biol., 12, 917, 1967.
122. Zengo, A. N., Pawluk, R. J., and Bassett, C. A. L.,
Stress-induced bioelectric potentials in the dentoalveolar complex, Am. J. Orthodont., 64, 17, 1973.
123. Zengo, A. N., Bassett, C. A. L., Pawluk, R. J.,
and Prountzos, G., In vivo bioelectric potentials in
the dentoalveolar complex, Am. J. Orthodont., 66,
130, 1974.
124. Marino, A. A. and Gross, B. D., Piezoelectricity
in cementum, dentine and bone, Arch. Oral Biol.,
34, 507, 1989.
125. Grimm, F. M., Bone bending, a feature of ortho-
dontic tooth movement, Am. J. Orthodont., 62, 384,

1972.
126. DeAngelis, V., Observations on the response of alveolar bone to orthodontic force, Am. J. Orthodont.,
58, 284, 1970.
127. Borgens, R. B., Endogenous ionic currents traverse
intact and damaged bone, Science, 225, 478, 1984.
128. Otter, M., Shoenung, J., and Williams, W. S.,
Evidence for different sources of stress-generated potentials in wet and dry bone, J. Orthoped. Res., 3,
321, 1985.
129. Friedenberg, Z. B. and Kohanim, M., The effect
of direct current on bone, Surg. Gynecol. Obstet.,
127, 97, 1968.
130. Friedenberg, Z. B., Harlow, M. C., and Brighton,
C. T., Healing of nonunion of the medial malleolus
by means of direct current: a case report, J. Trauma,
11, 863, 1971.
131. Bassett, C. A. L., Pilla, A. A., and Pawluk, R. J.,
Non-operative salvage of surgically resistant pseudoarthroses and non-unions by pulsing electromagnetic fields, Clin. Orthoped., 124, 128, 1977.
132. Friedenberg, Z. B. and Brighton, C. T., Bioelectricity and fracture healing, Plastic Reconstr. Surg.,
68, 435, 1981.
133. Marino, A. A., Electrical stimulation in orthopaedics: past, present and future, J. Bioelect., 3, 235,
1984.
134. Beeson, D. C., Johnston, L. E., and Wisotzky, J.,
Effect of constant currents on orthodontic tooth
movement in the cat, J. Dent. Res., 54, 251, 1975.
135. Davidovitch, Z., Finkelson, M. D., Steigman, S.,
Shanfeld, J. L., Montgomery, P. C., and Korostoff,
E., Electric currents, bone remodeling, and orthodontic tooth movement. I. The effect of electric currents on periodontal cyclic nucleotides, Am. J. Orthodont., 77, 14, 1980.
136. Davidovitch, Z., Finkelson, M. D., Steigman, S.,
Shanfeld, J. L., Montgomery, P. C., and Korostoff,
E., Electric currents, bone remodeling, and orthodontic tooth movement. II. Increase in rate of tooth
movement and periodontal cyclic nucleotide levels
by combined force and electric current, Am. J. Orthodont., 77, 33, 1980.
137. Davidovitch, Z., Montgomery, P. C., and
Shanfeld, J. L., Cellular localization and concentration of bone cyclic nucleotides in response to acute
PTE administration, Calcified Tissue Int., 24, 81,
1977.
138. Murad, F., Brewer, B., Jr., and Vaughn, M.,

Effect of thyrocalcitonin on adenosine, 3',5'-cyclic
phosphate formation by rat kidney and bone, Proc.
Natl. Acad. Sci. U.S.A., 65, 446, 1970.
139. Rasmussen, H. and Bordier, P., Vitamin D and
bone, Metab. Bone Dis. Relat. Res., 1, 7, 1978.
140. Kornstein, R., Somjen, D., Danon, A., Fisher,
H., and Binderman, I., Pulsed capacitive electric
induction of cyclic AMP changes, Ca45 uptake and
444
DNA synthesis in bone cells, Trans. Bioelectr. Repair Growth, 1, 34, 1981.
141. Rodan, G. A., Bourret, L. A., Harvey, A., and
Mensi, T., Cyclic AMP and cyclic GMP: mediators
of the mechanical effects on bone remodeling, Sci-
ence, 189, 467, 1975.

142.
Tashjian, A. H., Jr., Voelkel, E. F., Levine, L.,

and Goldhaber, P., Evidence that the bone resorption-stimulating factor produced by mouse fibrosarcoma cells is prostaglandin E2, J. Exp. Med., 136,
1329, 172.
Gomes, B. C., Hausmann, E., Weinfeld, N., and

DeLuca, C., Prostaglandins: bone resorption stimulating factors released from monkey gingiva, Calcified Tissue Int., 19, 285, 1976.
144. Shiozawa, S., Williams, R. C., Jr., and Ziff, M.,
Immunoelectron microscopic demonstration of prostaglandin E in rheumatoid synovium, Arthr. Rheum.,
25, 685, 1982.
145. Dekel, S., Lenthal, G., and Francis, M. J. 0.,
Release of prostaglandins from bone and muscle after
tibial fracture, Bone Jt. Surg., 63B, 185, 1981.
146. Uchida, A., Yamashita, K., Hashimoto, K., and
Shimomura, Y., The effects of mechanical stress on
cultured growth cartilage cells, Connect. Tissue Res.,
143.
17, 305, 1988.
147.
Somjen, D., Binderman, I., Berger, E., and Harell,

A., Bone remodeling induced by physical stress is
prostaglandin E2 mediated, Biochim. Biophys. Acta,
627, 91, 1980.
148.
Shen, V., Rifas, L., Kohler, G., and Peck, W. A.,

Prostaglandins change cell shape and increase intercellular gap junctions in osteoblasts cultured frdm rat
fetal calvaria, J. Bone Min. Res., 1, 243, 1986.
149.
Hasegawa, S., Sato, S., Saito, S., Suzuki, Y., and

Brunette, D. M., Mechanical stretching increases
the number of cultured bone cells synthesizing DNA
and alters their pattern of protein synthesis, Calcified
Tissue Int., 37, 431, 1985.
Buckley, M. J., Banes, A. J., Levin, L. G., Sumpio,

B. E., Sato, M., Jordan, R., Gilbert, J., Link,
G. W., and Tran Son Tay, R., Osteoblasts increase
their rate of division and align in response to cyclic,
mechanical tension in vitro, Bone Min., 4, 225, 1988.
151. Klein-Nutlend, J., Velhuijzen, J. P., de Jong, M.,
and Burger, E. H., Increased bone formation and
decreased bone resorption in fetal mouse calvaria as
a result of intermittent compressive force, in vitro,
150.
2, 441, 1987.
E.
Burger, H., Velhuijzen, J. P., Klein-Nulend,
J., and Van Loon, J. J. W. A., Osteoclastic invasion and mineral resorption of fetal mouse long
bone rudiments are inhibitied by culture under intermittent compressive force, Connect. Tissue Res., 20,
131, 1989.
Bone Min.,
152.
153.
Ozawa, H., Imamura, K., Etsuko, A., Takahashi,

N., Hiraide, T., Shibasaki, Y., Fukuhara, T., and
Suda, T., Effect of continuously applied compressive
pressure on mouse osteoblast-like cells (MC3T3-E1)

in vitro, J. Cell Physiol., 142, 177, 1990.
154. Duncan, G. W., Yen, E. H. K., Pritchard, E. T.,
and Suga, D. M., Collagen and prostaglandin synthesis in force-stressed periodontal ligament, in vitro,
J. Dent. Res., 63, 665, 1984.

Meikle, M. C., Heath, J. K., Atkinson, S. J.,
Hembry, R. M., and Reynolds, J. J., Molecular
biology of stressed connective tissues at sutures and
hard tissues in vitro, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC
Press, Boca Raton, FL, 1989, 71.
156. Curtis, A. S. G. and Sechar, G. M., The control
of cell division by tension or diffusion, Nature, 274,
52, 1978.
157. Norton, L. A., Anderson, K. L., Melsen, B.,
Arenholt Bindslev, D., and Celis, J. E., Buccal
mucosa fibroblasts and periodontal ligament cells
perturbed by tensile stimuli in vitro, Scand, J. Dent.
Res., 98, 36, 1990.
158. Ngan, P. W., Crock, B., Varghese, J., Lanese,
R., Shanfeld, J. L., and Davidovitch, Z., Immunohistochemical assessment of the effect of chemical
and mechanical stimuli on cAMP and prostaglandin
E levels in human gingival fibroblasts, in vitro, Arch.
Oral Biol., 33, 163, 1988.
159. Ngan, P., Saito, S., Saito, M., Shanfeld, J., and
Davidovitch, Z., The interactive effects of mechanical stress and interleukin 1 on prostaglandin E and
cyclic AMP production in human periodontal ligament fiborblasts in vitro: comparison with cloned
osteoblastic cells of mouse (MC3T3-E1), Arch. Oral
Biol., 35, 717, 1990.
160. Ragnarsson, B., Carr, G., and Daniel, J. C., Isolation and growth of human periodontal ligament cells
in vitro, J. Dent. Res., 64, 1026, 1985.
161. Brunette, D. M., Kanoza, R. J., Marmary, Y.,
Chan, J., and Melcher, A. H., Interactions between
epithelial and fibroblast-like cells in cultures derived
from monkey periodontal ligament, J. Cell Sci., 27,
127, 1977.
162. Brunette, D. M., Heersche, J. N. M., Purdon,
A. D., Sodek, J., Moe, H. K., and Assuras, J. N.,
In vitro cultured parameters and protein and prostaglandin secretion of epithelial cells derived from porcine rests of Malassez, Arch. Oral Biol., 24, 199,
1979.
163. Merrilees, M. J., Sodek, J., and Aubin, J. E.,
Effect of cells of epithelial rests of Malassez and
endothelial cells on synthesis of glycosaminoglycans
by periodontal ligament fibroblasts in vitro, Dev. Biol.,
97, 146, 1983.
164. Birek, C., Heersche, J. N. M., Jez, D., and
Brunette, D. M., Secretion of a bone resorbing factor by epithelial cells cultured from porcine rests of
Malassez, J. Periodont. Res., 18, 75, 1983.
165. Brunette, D. M., Mechanical stretching increases
155.
445
the number of epithelial cells
synthesizing DNA in
culture, J. Cell Sci., 69, 35, 1984.

166. Klein, D. C. and Raisz, L. G., Prostaglandins: stim-
ulation of bone resorption in tissue culture, Endocrinology, 86, 1436, 1970.

167. Rodan, G. A. and Martin, T. J., Role of osteoblasts
in hormonal control of bone resorption a hypothesis, Calcified Tissue Int., 33, 349, 1981.
168. Bindeman, I., Zor, U., Kaye, A. M., Shimshoni,
Z., Harell, A., and Somjen, D., The transduction
of mechanical force into biochemical events in bone

cells may involve activation of phospholipase A2,
Calcified Tissue Int., 42, 261, 1988.
169. Davidovitch, Z., Shanfeld, J. L., Montgomery,

P. C., Lally, E., Laster, L., Furst, L., and
Korostoff, E., Biochemical mediators of the effects
of mechanical forces and electric currents on mineralized tissues, Calcified Tissue Int., 36, S86, 1984.
170. Davidovitch, Z., Nicolay, O., Alley, K., Zwilling,
B., Lanese, R., and Shanfeld, J. L., First and second messenger interactions in stressed connective tissues in vivo, in The Biology of Tooth Movement,
Norton, L. A. and Burstone, C. J., Eds., CRC Press,
Boca Raton, FL, 1989, 97.
171. Yamasaki, K., Miura, F., and Suda, T., Prostaglandin as a mediator of bone resorption induced by
experimental tooth movement in rats, J. Dent. Res.,
59, 1635, 1980.
172. Yamasaki, K., Shibata, Y., and Fukuhara, T.,
The effects of prostaglandins on experimental tooth
movement in monkeys (Macaca fuscata), J. Dent.
Res., 61, 1444, 1982.
173. Yamasaki, K., Shibata, Y., Imai, S., Tani, Y.,
Shibasaki, Y., and Fukuhara, T., Clinical application of prostaglandin E, (PGE,) upon orthodontic
tooth movement, Am. J. Orthodont., 85, 508, 1984.
174. Yamasaki, K., Pharmacological control of tooth
movement, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press,
Boca Raton, FL, 1989, 287.
175. Jee, W. S. S., Ueno, K., Deng. Y. P., and
Woodbury, D. M., The effect of prostaglandin E2
in growing rats: increased metaphyseal hard tissue
and cortico-endosteal bone formation, Calcified Tissue Int., 37, 148, 1985.
176. Marks, S. C., Jr. and Miller, S. C., Local infusion
of prostaglandin E, stimulates mandibular bone formation in vivo, J. Oral Pathol., 17, 500, 1988.
177. Nefussi, J.-R. and Baron, R., PGE2 stimulates both
resorption and formation of bone in vitro: differential
responses of the periosteum and the endosteum in
fetal rat long bone cultures, Anat. Rec., 211,9, 1985.
178. Lee, W., Experimental study of the effect of prostaglandin administration on tooth movement with
particular emphasis on the relationship to the method
of PGE, administration, Am. J. Orthodont. Dentofacial Orthoped., 98, 231, 1990.
179.
Decker, J. L., Malone, D. G., Haroui, B., Wahl,

S., Schrieber, L., Klippel, J., Steinberg, A. G.,
and Wilder, R., Rheumatoid arthritis: evolving concepts of pathogenesis and treatment, Ann. Intern.
Med., 101, 810, 1984.
180.. Brandt, K. D. and Slowman-Kovacs, S., Nonsteroidal antiinflammatory drugs in treatment of osteoarthritis, Clin. Orthoped., 213, 84, 1986.
181. Torbinejad, M., Clagett, J., and Engel, D., A cat
model for the evaluation of mechanisms of bone resorption: induction of bone loss by stimulated immune complexes and inhibition by indomethacin,
Calcified Tissue Int., 29, 207, 1979.
182. Waite, I. M., Saxton, C. A., Young, A., Wagg,
B. J., and Corbett, M., The periodontal status of
subjects receiving non-steroidal anti-inflammatory
drugs, J. Periodont. Res., 16, 100, 1981.
183. Saffar, J. L. and Lasfargues, J. J., A histometric
study of the effect of indomethacin and calcitonin on
bone remodeling in hamster periodontitis, Arch. Oral
Biol., 29, 555, 1984.
184. Williams, R. C., Jeffcoat, M. K., Kaplan, M. L.,
Goldhaber, P., Johnson, H. G., and Wechter,
W. J., Flurbiprofen: a potent inhibitor of alveolar
bone resorption in beagles, Science, 277, 640, 1985.
185. Melcher, A. H., An overview of the anatomy and
physiology of the periodontal ligament, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL,
1989, 1.
186. van Steenberghe, D., The structure and function of
periodontal innervations, J. Periodont. Res., 14, 185,
1979.
187. Pfaffman, C., Afferent impulses from the teeth due
to pressure and noxious stimulation, J. Physiol., 97,
207, 1939.
188. Cash, R. M. and Linden, R. W. A., The distribution of mechanoreceptors in the periodontal ligament of the mandibular canine teeth of the cat, J.
Physiol., 330, 439, 1982.
189. Jyvisjiirvi, E., Kniffki, K.-D., and Mengel,
M. K. C., Functional characteristics of afferent C
fibers from tooth pulp and periodontal ligament, Prog.
Brain Res., 74, 237, 1988.
190. Freezer, S. R. and Sims, M. R., A transmission
electron-microscopy stereological study of the blood
vessels, oxytalan fibres and nerves of mouse-molar
periodontal ligament, Arch. Oral Biol., 32, 407, 1987.
191. Gould, T. R. L., Melcher, A. H., and Brunette,
D. M., Location of progenitor cells of periodontal
ligament of mouse molar stimulated by wounding,
Anat. Rec., 188, 133, 1977.
192. McCulloch, C. A. G. and Melcher, A. H., Cell
density and cell generation in the periodontal ligament of mice, Am. J. Anat., 167, 43, 1983.
193. Khouw, F. E. and Goldhaber, P., Changes in vasculature of the periodontium associated with tooth
movement in the rhesus monkey and dog, Arch. Oral
Biol., 15, 1125, 1970.
194. Rygh, P., Ultrastructural vascular changes in pressure zones of rat molar periodontium incident to or-
446
195.
196.
197.
198.
thodontic movement, Scand. J. Dent. Res., 80, 307,

1972.
Jeffcoat, M. K., Kaplan, M. L., Rumbaugh, C. L.,
and Goldhaber, P., Magnification angiography in
beagles with periodontal disease, J. Periodont. Res.,
17, 294, 1982.
Bien, S. M., Fluid dynamic mechanisms which regulate tooth movement, Adv. Oral Biol., 2, 173, 1966.
Lotz, M., Vaughan, J. H., and Carson, D. A.,
Effect of neuropeptides on production of inflammatory cytokines by human monocytes, Science, 241,
1218, 1988.
Laurenzi, M. A., Persson, M. A. A., Dalsgaard,
C.-J., and Ringden, O., Stimulation of human B
lymphocyte differentiation by the neuropeptides substance P and neurokinin A, Scand. J. Immunol., 30,
695, 1989.
199. Hafstrom, I., Gyllenhammar, H., Palmblad, J.,

and Ringertz, B., Substance P activates and modulates neutrophil oxidative metabolism and aggregation, J. Rhemuatol., 16, 1033, 1989.
200. Roscetti, G., Ausiello, C. M., Palma, C., Gulla,
P., and Roda, G., Enkephalin activity on antigeninduced proliferation of human peripheral blood mononucleate cells, Int. J. Immunopharm., 10, 819, 1988.
201. Marotti, T., Sverko, V., and Hrsak, I., Modulation of superoxide anion release from human polymorphonuclear cells by Met- and Leu-enkephalin,
Brain, Behav. Immun., 4, 13, 1990.
202. Weinstock, J. V., Blum, A., Walder, J., and
Walder, R., Eosinophils from granulomas in Schistosomiasis mansoni produce substance P, J. Immunol., 141, 961, 1988.
203. Weinstock, J. V. and Blum, A. M., Release of
substance P by granuloma eosinophils in response to
secretagogues in murine Schistosomiasis mansoni, Cell
Immunol., 125, 380, 1990.
204. Roth, K. A., Lorenz, R. G., Unanue, R. A., and
Weaver, C. T., Nonopiate active proenkephalin-derived peptides are secreted by T helper cells, FASEB
J., 3, 2402, 1989.
205. Fagarasan, M. O., Bishop, J. F., Rinaudo, M. S.,
and Axelrod, J., Interleukin 1 induces early protein
phosphorylation and requires only a short exposure
for late induced secretion of P-endorphin in a mouse
pituitary cell line, Proc. Natl. Acad. Sci. U.S.A., 87,
2555, 1990.
206. Fagarasan, M. 0. and Axelrod, J., Interleukin-1
amplifies the action of pituitary secretagogues via
protein kinases, Int. J. Neurosci., 51, 311, 1990.
207. Su, T.-P., London, E. D., and Jaffe, J. H., Steroid
binding at a receptors suggests a link between endocrine, nervous, and immune systems, Science, 240,
219, 1988.
208. Levine, J. D., Clark, R., Devor, M., Helms, C.,
Moskowitz, M. A., and Basbaum, A. I., Intraneuronal substance P contributes to the severity of experimental arthritis, Science, 226, 547, 1984.
209. Lotz, M., Carson, D. A., and Vaughan, J. H.,
210.
211.
212.
213.
214.
215.
Substance P activation of rheumatoid synoviocytes:

neural pathway in pathogenisis of arthritis, Science,
235, 893, 1987.
Yokoyama, M. M. and Fujimoto, K., Role of lymphocyte activation by substance P in rheumatoid arthritis, Int. J. Tissue React., 12, 1, 1990.
O'Bryne, E. M., Blancuzzi, V. J., Wilson, D. E.,
Wong, M., Peppard, J., Simke, J. P., and Jeng,
A. Y., Increased intra-articular substance P and prostaglandin E2 following injection of interleukin-1 in
rabbits, Int. J. Tissue React., 12, 11, 1990.
Lam, F. Y. and Ferrell, W. R., Inhibition of carrageenan-induced joint inflammation by substnace P
antagonist, Ann. Rheum. Dis., 48, 928, 1989.
Lam, F. Y. and Ferrell, W. R., Capsaicin suppresses substance P-induced joint inflammation in the
rat, Neurosci. Lett., 105, 155, 1989.
Olgart, L., Gazelius, B., Brodin, E., and Pernow,
B., Localization of substance P-like immunoreactivity in nerves in the tooth pulp, Pain, 4, 153, 1977.
Olgart, L., Gazelius, B., Brodin, E., and Nilsson,
G., Release of substance P-like immunoreactivity from
the dental pulp, Acta Physiol. Scand., 101, 510, 1977.
216. Wakisaka,
S., Nishikawa, S., Ichikawa,H.,
Matsuo, S., Takano, Y., and Akai, M., The distribution and origin of substance P-like immunoreactivity in the rat molar pulp and periodontal tissues,
Arch. Oral Biol., 30, 813, 1985.
217.
Wakisaka, S., Ichikawa, H., Nishikawa, S.,

Uchiyama, T., Matsuo, S., Takano, Y., and Akai,
M., Immunohistochemical study on regeneration of
substance P-like immunoreactivity in rat molar pulp
and periodontal ligament following resection of the
inferior alveolar nerve, Arch. Oral Biol., 32, 225,
1987.
218.
Wakisaka, S., Ichikawa, H., Nishikawa, S.,

Matsuo, Y., Takano, Y., and Akai, M., Immu-
nohistochemical observation on the correlation between substance P- and vasoactive intestinal polypeptide-like immunoreactivities in the feline dental
pulp, Arch. Oral Biol., 32, 449, 1987.
219. Davidovitch, Z., Nicolay, O. F., Ngan, P. W., and
Shanfeld, J. L., Neurotransmitters, cytokines, and
the control of alveolar bone remodeling in orthodontics, Dent. Clin. N. Am., 31, 411, 1988.
220. Said, S. I. and Mutt, V., Polypeptide with broad
biological activity: isolation from small intestine, Science, 169, 1217, 1970.
221. Hohmann, E., Levine, L., and Tashjian, A. H.,
Jr., Vasoactive intestinal peptide stimulates bone resorption via a cyclic adenosine 3',5'-monophosphate-
dependent mechanism, Endocrinology, 112, 1233,

1983.
222.
Hohmann, E. L. and Tashjian, A. H., Jr., Functional receptors for vasoactive intestinal peptide of
human osteosarcoma cells, Endocrinology, 114, 1321,
223.
Hohmann, E. L., Elde, R. P., Rysavy, J. A.,

Einzig, S., and Gebhard, R. L., Innervation of
1984.
447
periosteum and bone by sympathetic vasoactive intestinal peptide-containing nerve fibers, Science, 232,
868, 1986.
Herness, M. S., Vasoactive intestinal peptide-like
immunoreactivity in rodent taste cells, Neuroscience,
33, 411, 1989.
225. Motakef, M., Shanfeld, J., and Davidovitch, Z.,
224.
Localization of VIP at bone resorption sites in vivo,

J. Dent. Res., 69, 253, 1990.
226. Rosenfeld, M. G., Mermod, J.-J., Amara, S. G.,
Swanson, L. W., Sawchenko, P. E., Rivier, J.,
Vale, W. W., and Evans, R. M., Production of a
novel neuropeptide encoded by the calcitonin gene
via tissue-specific RNA processing, Nature, 304, 129,
1983.
227. Stjarne, P., Lundblad, L., Angard, A., Hokfelt,
T., and Lundberg, J. M., Tachykinins and calcitonin gene-related peptide: co-existence in sensory
nerves of the nasal mucosa and effects on blood flow,
Cell Tissue Res., 256, 439, 1989.
228. Fujimori, A., Saito, A., Kimura, S., Watanabe,
T., Uchiyama, Y., Kawasaki, H., and Goto, K.,
Neurogenic vasodilation and release of calcitonin generelated peptide (CGRP) from perivascular nerves in
the rat mesenteric artery, Biochem. Biophys. Res.
Commun., 165, 1391, 1989.
229. Larsson, J., Ekblom, A., Henriksson, K.,
Lundeberg, T., and Theordorsson, E., Immunoreactive tachykinins, calcitonin gene-related peptide
and neuropeptide Y in human synovial fluid from
inflamed knee joints, Neurosci. Lett., 100, 326, 1989.
230. Silverman, J. D. and Kruger, L., Calcitonin-generelated-peptide immunoreactive innervation of the rat
head with emphasis on specialized sensory structures,
J. Comp. Neurol., 280, 303, 1989.
231. Wakisaka, S., Ichikawa, H., Nishikawa, S.,
Matsuo, S., Takano, Y., and Akai, M., The distribution and origin of calcitonin gene-related peptide-containing nerve fibers in feline dental pulp, Histochemistry, 86, 585, 1987.
232. Taylor, P. E., Byers, M. R., and Redd, P. E.,
Sprouting of CGRP nerve fibers in response to dentin
injury in rat molars, Brain Res., 461, 371, 1988.
233. Kata, J., Ichikawa, H., Wakisaka, S., Matsuo,
S., Sakuda, M., and Akai, M., The distribution of
vasoactive intestinal polypeptides and calcitonin generelated peptide in the periodontal ligament of mouse
molar teeth, Arch. Oral Biol., 35, 63, 1990.
234. Hill, E. L. and Elde, R., Calcitonin gene-related
peptide-immunoreactive nerve fibers in mandibular
periosteum of rat: evidence for primary afferent origin, Neurosci. Lett., 85, 172, 1988.
235. Gazelius, B., Edwall, B., Olgart, L., Lundberg,
J. M., Hokfelt, T., and Fischer, J. A., Vasodilatory
effects and coexistence of calcitonin gene-related
peptide (CGRP) and substance P in sensory nerves
of cat dental pulp, Acta Physiol. Scand., 130, 33,
1987.
236. Takahashi, O., Traub, R. J., and Ruda, M. A.,
Demonstration of calcitonin gene-related peptide immunoreactive axons containing dynorphin A (1-8)

immunoreactive spinal neurons in a rat model of peripheral inflammation and hyperalgesia, Brain Res.,
475, 168, 1988.
237. Kvinnsland, I. and Kvinnsland, S., Changes in
CGRP-immunoreactive nerve fibers during experimental tooth movement in rats, Eur. J. Orthodont.,
12, 320, 1990.
238. Okamoto, Y., Davidovitch, Z., and Shanfeld, J.,
CGRP in dental pulp and periodontal ligament during
tooth movement (abstract), J. Dent. Res., submitted.
239. Tashiro, T., Takahashi, O., Satoda, T.,
Matsushima, R., and Mizuno, N., Immunohistochemical demonstration of coexistence of enkephalinand substance P-like immunoreactivities in axonal
components in the lumbar segments of cat spinal cord,
Brain Res., 424, 391, 1987.
240. Jessell, T. M. and Iversen, L. L., Opiate analgesics
inhibit substance P release in rat trigeminal nucleus,
Nature, 268, 549, 1977.
241. Walker, J. A., Tanzer, F. S., Harris, E. F.,
Wakelyn, C., and Desiderio, D. M., The enkephalin response in human tooth pulp to orthodontic
force, Am. J. Orthodont. Dentofacial Orthoped., 92,
9, 1987.
242. Tanzer, F. S., Tolun, E., Fridland, G. H., Dass,
C., Killmar, J., Tinsley, P. W., and Desiderio,
D. M., Methionine-enkephalin peptides in human
teeth, Int. J. Peptide Protein Res., 32, 117, 1988.
243. Robinson, Q. C., Killmar, J. T., Desiderio, D. M.,
Harris, E. F., and Fridland, G., Immunoreactive
evidence of beta-endorphin and methionine-arg-glyleu in human tooth pulp, Life Sci., 45, 987, 1989.
244. Hanazawa, S., Ohmori, Y., Amano, S., Miyoshi,
T., Kumegawa, M., and Kitano, S., Spontaneous
production of interleukin-1-like cytokine from mouse
osteoblastic cell line (MC3T3-E1), Biochem. Biophys. Res. Commun., 131, 774, 1985.
245. Hanazawa, S., Amano, S., Nakada, K., Ohmori,
Y., Miyoshi, T., Hirose, K., and Kitano, S., Biological characterization of interleukin-1-like cytokine
produced by cultured bone cells from newborn mouse
calvaria, Calcified Tissue Int., 41, 31, 1987.
246. Rath, N. C., Oronsky, A. L., and Kerwar, S. S.,
Synthesis of interleukin-1-like activity by normal rat
chondrocytes in culture, Clin. Immunol. Immunopathol., 47, 39, 1988.
247. Kaplan, E., Dinarello, C. A., and Gelfand, J. A.,
Interleukin-1 and the response to injury, Immunol.
Res., 8, 118, 1989.
248. Gowen, M., Wood, D. D., and Russell, G. G.,
Stimulation of the proliferation of human bone cells
in vitro by human monocyte products with interleukin-1 activity, J. Clin. Invest., 75, 1223, 1985.
249. Boyce, B. F., Yates, A. J. P., and Mundy, G. R.,
Bolus injections of recombinant human interleukin1 cause transient hypocalcemia in normal mice, Endocrinology, 125, 2780, 1989.
448
Boyce, B., Aufdemorte, T.,

Bonewald, L., and Mundy, G. R., Infusions of
250. Sabatini, M.,
recombinant human interleukins la and 1p cause

hypercalcemia in normal mice, Proc. Natl. Acad. Sci.
U.S.A., 85, 5235, 1988.
251. Feige, U., Karbowski, A., Rordorf-Adam, C., and
Pataki, A., Arthritis induced by continuous infusion
of hr-interleukin-la into the rabbit knee joint, Int. J.
Tissue React., 11, 225, 1989.
252. Mizel, S. B., Dayer, J. M., Krane, S. M., and
Mergenhagen, S. E., Stimulation of rheumatoid synovial cell collagenase and prostaglandin production
by partially purified lymphocyte-activating factor (interleukin 1), Proc. Natl. Acad. Sci. U.S.A., 78, 2474,
1981.
253. Rafter, G. W., Interleukin 1 and rheumatoid arthritis, Med. Hypoth., 27, 221, 1988.
254. Johnson, W. J., Breton, J., Newman-Tarr, T.,
Connor, J. R., Meunier, P. C., and Dalton, B. J.,
Interleukin-1 release by rat synovial cells is dependent on sequential treatment with r-interferon and
lipopolysaccharide, Arthr. Rheum., 33, 261, 1990.
255. Masada, M. P., Persson, R., Kenney, J. S., Lee,
S. W., Page, R. C., and Allison, A. C., Measurement of interleukin-la and - 1 in gingival cervicular
fluid: implications for the pathogenesis of periodontal
disease, J. Periodont. Res., 25, 156, 1990.
256. Thomson, B. M., Saklatvala, J., and Chambers,
T. J., Osteoblasts mediate interleukin 1 stimulation
of bone resorption by rat osteoclasts, J. Exp. Med.,
150, 104, 1986.
257. Dewhirst, F. E., Ago, J. M., Peros, W. J., and
Stashenko, P., Synergism between parathyroid hormone and interleukin 1 in stimulating bone resorption
in organ culture, J. Bone Min. Res., 2, 127, 1987.
258. Gowen, M. and Mundy, G. R., Actions of recombinant interleukin 1, interleukin 2 and interferongamma on bone resorption in vitro, J. Immunol., 136,
2478, 1986.
259. Tatakis, D. N., Schneebergy, G., and Dziak, R.,
Recombinant interleukin-l stimulates prostaglandin
E2 production by osteoblastic cells: synergy with
parathyroid hormone, Calcified Tissue Int., 42, 358,
1988.
260. Ries, W. L., Seeds, M. C., and Key, L. L., Interleukin-2 stimulates osteoclastic activity: increased
acid production and radioactive calcium release, J.
Periodont. Res., 24, 242, 1989.
261. Davidovitch, Z., Lynch, P., and Shanfeld, J. L.,
Interleukin 2 localization in the mechanically-stressed
periodontal ligament, J. Dent. Res., 68, 332, 1989.
262. Nieto-Sampedro, M. and Chandy, K. G., Interleukin-2-like activity in injured rat brain, Neurochem.
Res., 12, 723, 1987.
263. Bevilacqua, M. P., Pober, J. S., Majeau, G. R.,
Fiers, W., Cotran, R. S., and Gimbrone, Jr., Recombinant tumor necrosis factor induces procoagulant activity in cultured human vascular endothelium:
characterization and comparison with the actions of
interleukin 1, Proc. Natl. Acad. Sci. U.S.A., 83,

4533, 1986.
264. Gamble, J. R., Harlan, J. M., Klebanoff, S. J.,
and Vadas, M. A., Stimulation of the adherence of
neutrophils to umbilical vein endothelium by human

recombinant tumor necrosis factor, Proc. Natl. Acad.
Sci. U.S.A., 82, 8667, 1985.
265. Bertolini, D. R., Nedwin, G. E., Bringman, T. S.,
Smith, D. D., and Mundy, G. R., Stimulation of
bone resorption and inhibition of bone formation in
vitro by human tumor necrosis factors, Nature, 319,
516, 1986.
266. Delaisse, J.-M., Eeckhout, Y., and Vaes, G., Boneresorbing agents affect the production and distribution
of procollagenase as well as the activity of collagenase in bone tissue, Endocrinology, 123, 264, 1988.
267. Yoshihara, R., Shiozawa, S., Imai, Y., and Fujita,
T., Tumor necrosis factor a and interferon T inhibit
proliferation and alkaline phosphatase activity of human osteoblastic SaOS-2 cell line, Lymphokine Res.,
9, 59, 1990.
268. Shapiro, S., Tatakis, D. N., and Dziak, R., Effect
of tumor necrosis factor alpha on parathyroid hormone-induced increase in osteoblastic cell cyclic
AMP, Calcified Tissue Int., 46, 60, 1990.
269. Stashenko, P., Dewhirst, F. E., Peros, W. J., Kent,
R. L., and Ago, J. M., Synergistic interactions between interleukin 1, tumor necrosis factor, and lymphotoxin in bone resorption, J. Immunol., 138, 1464,
1987.
270. Pfeilschifter, J., Chenu, C., Bird, A., Mundy,
G. R., and Roodman, G. D., Interleukin-1 and tumor necrosis factor stimulate the formation of human
osteoclast-like cells in vitro, J. Bone Min. Res., 4,
113, 1989.
271. Hashimoto, J., Yoshikawa, H., Takaoka, K.,
Shimizu, N., Mashuhara, K., Tsuda, T.,
Miyamoto, S., and Ono, K., Inhibitory effects of

tumor necrosis factor alpha on fracture healing in rats,
Bone, 10, 453, 1989.
272.
Johnson, R. A., Boyce, B. F., Mundy, G. R., and

Roodman, G. D., Tumors producing human tumor
necrosis factor induce hypercalcemia and osteoclastic
bone resorption in nude mice, Endocrinology, 124,
1424, 1989.
Hopkins, S. J. and Meager, A., Cytokines in synovial fluid. II. The presence of tumor necrosis factor
and interferon, Clin. Exp. Immunol., 73, 88, 1988.
274. Maury, C. P. J. and Teppo, A.-M., Cachectin/
tumor necrosis factor-a in the circulation of patients
273.
with rheumatic disease, Int. J. Tissue React., 11,

189, 1989.
275. Yocum, D. E., Esparza, L., Dubry, S., Benjamin,
J. B., Volz, R., and Scuder, P., Characteristics of
tumor necrosis factor production in rheumatoid arthritis, Cell Immunol., 122, 131, 1989.
276. Gowen, M., Nedwin, G. E., and Mundy, G. R.,
Preferential inhibition of cytokine-stimulated bone re-
449
277.
278.
279.
280.
281.
282.
283.
sorption by recombinant interferon gamma, J. Bone

Min. Res., 1, 469, 1986.
Gowen, M., MacDonald, B. R., and Russell, G.,
Actions ofrecombinant human T-interferon and tumor
necrosis factor a on the proliferation and osteoblastic
characteristics of human trabecular bone cells in vitro, Arthr. Rheum., 31, 1500, 1988.
Nanes, M. S., McKoy, W. M., and Marx, S. J.,
Inhibitory effects of tumor necrosis factor-a and interferon-T on deoxyribonucleic acid and collagen synthesis by rat osteosarcoma cells (ROS 17/2.8), Endocrinology, 124, 339, 1989.
Vignery, A., Niven-Fairchild, T., and Shepard,
M. H., Recombinant murine interferon-r inhibits the
fusion of mouse alveolar macrophages in vitro, but
stimulates the formation of osteoclast-like cells in
implanted syngeneic bone particles in mice in vivo,
J. Bone Min. Res., 5, 637, 1990.
Melin, M., Hartmann, D. J., Magloire, H.,
Falcoffe, E., Auriault, C., and Grimaud, J. A.,
Human recombinant gamma-interferon stimulates
proliferation and inhibits collagen and fibronectin
production by human dental pulp fibroblasts, Cell
Mol. Biol., 35, 97, 1989.
Judis, G., Davidovitch, Z., and Shanfeld, J., Identification of IFN at in vivo bone resorption sites, J.
Dent. Res., 69, 244, 1990.
Rosol, T. J., Shanfeld, J. L., and Davidovitch,
Z., Biochemical aspects of orthodontic tooth movement. II. Effects of neurotransmitters and cytokines
on bone resorption and formation in vitro, J. Dent.
Res., submitted.
Saito, S., Ngan, P., Saito, M., Kim, K., Lanese,
R., Shanfeld, J., and Davidovitch, Z., Effects of
cytokines on prostaglandin E and cAMP levels in
human periodontal ligament fiborblasts in vitro, Arch.
Oral Biol., 35, 387, 1990.

Saito, S., Ngan, P., Saito, M., Lanese, R.,
Shanfeld, J., and Davidovitch, Z., Interactive effects between cytokines on PGE production by human
periodontal ligament fibroblasts, in vitro, J. Dent.
Res., 69, 1456, 1990.
285. Saito, S., Rosol, T. J., Saito, M., Ngan, P. W.,
Shanfeld, J., and Davidovitch, Z., Bone-resorbing
activity and prostaglandin E produced by human per-
284.
iodontal
ligament cells in vitro, J. Bone Min. Res.,
5, 1013, 1990.
286. Saito, S., Saito, M., Ngan, P., Lanese, R.,
Shanfeld, J., and Davidovitch, Z., Effects of para-
thyroid hormone and cytokines on prostaglandin E

synthesis and bone resorption by human periodontal
ligament fibroblasts, Arch. Oral Biol., in press.
287. Saito, S., Saito, M., Ngan, P. W., Shanfeld, J.,
and Davidovitch, Z., Interleukin 1 beta and prostaglandin E are involved in the response of periodontal cells to mechanical stress in vivo and in vitro,
Am. J. Orthodont. Dentofacial Orthoped., in press.
450

Tooth Movement

Încărcat de

Informații document

Drepturi de autor

Formate disponibile

Partajați acest document

Partajați sau inserați document

Opțiuni de partajare

Vi se pare util acest document?

Este necorespunzător acest conținut?

Drepturi de autor:

Formate disponibile

Tooth Movement

Încărcat de

Drepturi de autor:

Formate disponibile

Critical Reviews http://cro.sagepub.

>> Version of Record - Jan 1, 1991

Critical Reviews in Oral Biology and Medicine,

Throughout their natural history, teeth move

neurotransmitters, cytokines, synergism.

dictated by the forces of the surrounding muscles

II. HISTORICAL PATHFINDERS

dontic tooth movement: "To extract an irregular

aries as misinterpretations. He was also criticized

III. HISTOMORPHOLOGY OF TOOTH

evaluation. Moreover, Reitan studied paradental

activity.26 Based on these observations, Storey,27

erative and remodeling reactions. Prolonged

B. Histochemical Changes Associated

activity was found in alveolar bone marrow cells

resorption, reattachment of the PDL seemed to

trast, Martinez and Johnson35 were better able to

a spring for periods of time ranging from 15 min

tochondria of osteoclasts by electron microscopy.

C. Ultrastructural Changes in Paradental

Despite Reitan's reported observation'9 that

amined stretched cranial sutures in rats.32 In both

PDL. In the latter site, Rygh was undecided on

pansion device, and the teeth were then retained

D. Autoradiographic Examination of the

of actively migrating and matrix-secreting cells.

E. Tooth Movement: Visible Tissue

ing recommendations. Schwarz warned against

IV. MECHANISMS OF CELLULAR

cle cells,84 osteosarcoma cells,85 and endothelial

crease in nuclear size is necessary for S phase

B. The Effect of Mechanical Stress on

Typically, orthodontic forces are applied

FIGURE 1. "Flat" alveolar bone osteoblasts (arrows)

periodontal ligament. (Magnification

(10,000 to 12,000 microstrain, 0.5 Hz), for 42

d. When the number of cycles was increased to

FIGURE 2. Alveolar bone osteoblasts (arrows) at PDL

FIGURE 3. Periodontal cells in PDL compression zone

teoregulatory stimulus, suggesting that functional

candidates to sense the distribution, rate of

that have incorporated 3H-uridine into their

ingly critical involvement of bone matrix in the

2. Bioelectric Phenomena in Bone

boosted by Fukada and Yasuda's" report in 1957

in their vicinity. Borgens,127 examining intact and

Bioelectric measurements in alveolar

pressed (concave) side of the orthodontically

(i.e., its pathway and density) remains unknown

a current of 20 ILA to gingiva near orthodontically

3. Biochemical Events in Cells Affected

or cGMP) have been identified as mediators of

the level of tissue inhibitors of metalloproteinases

frequency and in the proportion of cells in S

PDL fibroblasts. They observed no junctional

ever, one of the main foci of attention in this

the staining for PGE in PDL and alveolar bone

V. REGULATION OF TOOTH MOVEMENT

PDL, while cementoblasts cover the dental root

(1 to 7 d) on the PDL vascular bed, Khouw and

tion, respectively. Examination by TEM of se-

vere PDL compression sites in rats94 revealed

4): (1) movement of fluids within the PDL; (2)

B. Interactions between Cells and