Transcribed by Jazmin Lui May 5, 2014

Craniofacial Biology: Periodontal Tissues: PDL Dr. Craig

1. Craniofacial Biology Periodontal tissues: Periodontal ligament

Good morning everyone, welcome back. So last week we started a discussion
of the development of periodontal tissues and I think we tried to impress
upon you was the development of the periodontal connective tissue
attachment apparatus cementum, PDL, alveolar bone, its really an extension
of the mesothelial interactions that characterize odontogenesis. Gingiva
doesnt participate in it. So gingivas there even if teeth dont form. And then
what else did we talk about? We talked about how the different tissues
develop. We talked a little bit about cementum and since we didnt have the
cementum lecture and kind of harshly let you listen to last years podcast, Ive
never even listened to my own podcast and I make it a point to because I
dont want to be embarrassed. So anyways you listened to the podcast, um
and thats kind of unfortunate the way the schedule worked out because the
cementum lecture is my favourite lecture. I kinda look at cementum as the
last calcified frontier in the human body. It is the last mineralized tissue to
evolve and it has some unique characteristics. But what you should know
from the cementum lecture is ways of characterizing cementum. Most of
which are foolish. Time of formation: primary, secondary. What were going to
use is whether it has cells in it and whether it has fibers in it, so you should
know what kind of cementum will be found whether its in the coronal part of
the tooth, right? If its in the oral cavity its not attached to anything so
theres no fibres in it. If its in the oral cavity theres no blood supply there
right? So it cant be cellular so it has to be acellular, afibrillar, those kinds of
things. The first cementum laid down during development is fibrillar acellular
cementum. And then sometimes on the apical part of the teeth, the lower
third, theres another kind of cementum that gets laid down thats cellular
cementum. It has cementocytes in it, and its very analogous to bone, except
all the cell processes, or the canaliculi point out to the PDL because the PDL
Transcribed by Jazmin Lui May 5, 2014

is the only source of nutrients. What else did I tell you about cementum?
There is cellular cementum and theres acellular cementum and there are 2
forms of cementum which are misnomers, alright. So that intermediate
cementum layer sometimes called the hyaline layer of Hopewell smith, thats
synthesized by the inner cellular layer of Hertzwigs epithelial root sheath. It
has basement membrane components, but more importantly, if youre in the
pharmaceutical industry it has amelogenin or amelogenin-like peptides. And
those amelogenin-like peptides we think are inductive for cellular cementum
formation and were going to use that this week when we talk about
periodontal regeneration. The fourth type of cementum which is a misnomer;
so intermediate cementum is not a cementum because its not made
mesenchymal cells, alright? Cementum is a mesenchymal mineral tissue.
Sometimes in a clinic youll hear the words, especially from your
neighbourhood periodonttist that the cementum layer thats been lined up
against the periodontal pocket has absorbed bacterial products so we call
that affected cementum. So one of the goals of scaling and root planing,
which youll learn very soon if you havent learned already, is to remove the
affected cementum layer. And I think thats just about eveyrthing I kind of
wanted to fill you in on. And were going to kind of put some points from
cementum lecture in these lectures. So this is the longest lecture of the unit,
the PDL. So we talked last walk that it shouldnt be called a ligament, it
should be called a suture, but even then its a very specialized suture. It has
cementum on one side, alveolar bone on the other side, and we have these
fibers that go across. But then you also have these little guys, you have these
little cells, and these epithelial cells, I cant think of another ligament suture
that has epithelial cells, pathologically imbedded in it, can you? I cant
remember. You guys took anatomy so, youre better experts than I am. So we
have these epithelial cells in there. This PDL can do wonderful things for our
patients. And Im going to try to underline that in our discussion of the PDL.

2. Periodontal Ligament (Why not ankylosis?)
Transcribed by Jazmin Lui May 5, 2014


OK so, why do we have this gomphosis? I have no idea why we call it that,
anatomically that kind of suture is called a gomphosis. Why not just an
ankylosis? Like a dental implant? Why not just have a bone ankylose right to
the tooth surface? So we talked a little bit last week about dynamic support
and attachment to dentition. Youll want to underline this dynamic, when you
get into the clinic youll find out how dynamic that PDL is in some of your
patients, orthodontics, those sorts of things. It provides for eruption. So
humans continue to development post utero, so we have 2 sets of teeth, and
this idea of eruption allows us to have 2 sets of teeth to develop post utero.
A lot of patients that Ive placed implants in, and this is what they complain
about: Dr. Craig when I bite down I cant feel it, you know? so theres lots of
proprioceptive receptors on PDLs. So you get this real feedback when you
chew on bolus of food how hard it is, the consistency, it gives you an
essential feeling you dont get with dental implants. So as we go through this
unit were gonna kinda compare osteointegration, which is nothing more than
a functional ankylosis right on the titanium oxide surface on implants, and this
wonderous thing called PDL. Were going to compare and contrast them,
because up until recently, and I think the pendulum has swung back a bit, but
this school used to be really into putting in implants. Pull out teeth, put in
implants, they wont get tooth decay. And I think dental implants, dont get
me wrong, I think that implants are wonderful for the replacement of missing
teeth, but I dont think they are a replacement for teeth. And Ill kind of give
you my bias as we go through this. So were going to compare teeth a little
bit with dental implants as we go along.

3. PDL macroscopic features: Dimensions
Ok so this is, this falls underneath the name of stuff you have to memorize.
Why I have no idea why but this stuff shows up on standardized exams. When
I took my perio boards years ago, it was on my perio boards. For some
reason people who make the standardized exam love to ask you the width of
Transcribed by Jazmin Lui May 5, 2014

the human PDL. And what you have to remember is the dimension is 0.2, 0.2
mm. The average width of the PDL. Why this is important I really dont know,
there are some things in our profession are sort of, interwoven and become
almost like folk history. So one of those things you have to remember is its
0.2 mm for the PDL. Because the tooth has some motion, its gong to be
narrowest at mid root and widest coronally and apically. The PDL is the widest
in young folks and its one of the few things that get thinner as you age is
the PDL. Ok? In general the PDL width will increase with function. So heres
some average data taken from literature. So heavy occlusion can mean, in this
series of experiments can mean 0.2 or so, without an antagonist this kind of
goes down. And when its impacted it goes down even further. This is isnt
anything thats kind of earth shattering but its stuff you have to know to go
through.

3. PDL macroscopic features: Surface area

If you kind of think about the root surfaces in the human adult dentition,
theres a lot of root surface. And I think thats one of the reasons why theres
these associations with periodontitis and systemic effects, which well talk
about in general pathology. Theres a lot of root surface area in there; the
most root surface is on the maxillary molars, about a third of the root surface
is on the maxillary molars. Why? When you start placing implants all of
sudden this will come back to you. Why do maxillary molars have 3 roots and
mandibular molars have 2 roots? Ever think about stuff like this? Does this
ever keep you up at night? It keeps me up at night. So there is a great
thinker who happened to be in Jamaican, and his name is Bob Marley, so Bob
Marley, when he was coherent had this song, one of the lyrics that went like
for every action there is a reaction. So if you think about it, mandibular bone
is really dense. Big cortical plate, lots of trabeculae. Maxillary bone, posterior,
got the sinus up there, and its really thin, waspy bone, right? Or wispy bone.
So you need more surface area to contact more bone, so that you can act
Transcribed by Jazmin Lui May 5, 2014

equally against the force of the mandible. This will come back to you when
you start placing implants because implants placed on the maxillary posterior,
thats the hardest place to place an implant. Maxillary molars are about a
third of the total root surface area.

5. PDL composition

So heres a cross-section of our PDL. So heres cementum over here, theres
alveolar bone over here, heres Sharpeys Fibers coursing out of the bone, you
cant really see them. Theres fibers coursing out of the cementum also,
youve got enormous neurovascular bundles, so this is obviously a tissue
turning over really quickly. And if we kinda look, if we do some histo-
morphology, oh they used to love doing this in the 1960s, they would take
come sections of tissue and figure out what the surface area or volume was.
So by far, where are my fibres, so by far the PDL fibres take up half the
volume, if you will, of the PDL. So lets talk a little bit about PDL fibres.

7. Periodontal ligament fibers

So if you take an SEM of the PDL, heres some PDL fibers cut in longitudinal
section and here theyre cut in cross-section and you kind of get this idea
they are cable, right? So this is Type I collagen that we talked about in Basic
Tissues. So most of the collagen is going to be Type I. And associated with
Type I is a little bit of Type III. And then this FACIT collagen, Type XII that we
talked about in Basic Tissues is in the PDL also.

8. Periodontal ligament fibers

So as it turns out for the purposes of this course were gonna divide these
fibres into two groups. Principal fibers and gingival fibres, alright? And
principal fibres, these are going to be the guys that attach the tooth to the
Transcribed by Jazmin Lui May 5, 2014

alveolus, alright? In a dental implant do you have principal fibres? Nope.
Dont have cementum. Dont have principal fibres. So all these guys are
knocked out in a dental implant. And then gingival fibers are those that
attach the gingiva to the tooth surface. And theyre important to make
biological seal, the function of our gingiva. So there are 5 principal fibres and
5 gingival fibres and you gotta know this stuff.

9. Periodontal ligament fibers (Principal Fibers)

This also falls under the heading of stuff Ive got to know. Ok so heres some,
lets talk about principal fibres. And since the principal fibres are in the
developing tooth they develop in a coronal, from a coronal to apical direction.
So the first fibres we can see apically on these are the alveolar crest group
they go from cementum just below the epithelial attachment and course to
the very apical descent of the alveolar crest. And then theres some fibres that
appear histologically to be horizontally located and theyre called the
horizontal group. And then theres these fibres that do most of the work, they
look like the suspension of the Mercedes Benz and theyre obliquely oriented,
kind of suspending the tooth from the alveolus, oriented ?... theyre called
the oblique group. And then around the apex there are some fibers that
radiate out, and theyre called the inter-radicular, out of the apical group. And
on multi rooted teeth, there are some fibres that go from the furcation area
to the interdental septum, and theyre called interradicular. So these are five
guys, the five principal fibres you have to know.

10. Periodontal ligament fibers (Principal fibers: apical group)

And if you actually look for these in histology, so this isso youre looking at
the apex of the tooth, right? Cut in cross-section. And so heres alveolar bone,
and notice this is 0.2 mm. Neurovascular bundles all over the place. And then
you see these orientations of cells and fibres. So these would be the apical
Transcribed by Jazmin Lui May 5, 2014

group. You have to have a little bit of imagination to see them. So heres
probably one over here, heres probably one over here. Theyre not that easy
to discern but early histologists thought they were quite neat so they named
them, we need to know these guys.


11. Periodontal ligament fibers (Gingival fibers)

So how about gingival fibres? So these are not participating in attaching the
tooth to the alveolus. But they are participating in attaching the gingiva to
the tooth. And theyre named by origin and insertion. Right? Dentogingval
group. So that starts off in the cementum and goes off into the gingiva, so
thats the dentogingival group. Dentoperiosteal, so they start off in the
cementum and they go off and insert in the alveolar bone on the external
aspect. Dentoperiosteal group. And then the alveoloar gingival group, right?
They go from the alveolar gingival crest and go up to gingiva. So theyre not
actually attached to the tooth but theyre in some substance, if you will, like
the gingiva, and attaching it to the underlying bone. And then we have the
circular group. So the circular group goes around the tooth sort of like the
steel bands on a steel banded tire. And so heres some circular guys cut in
cross-section. And finally the trans-septal group, they go from tooth on one
side, go across the interdental septum and go to the tooth next door. So they
kind of attach teeth together and give continuity to the gingiva. So which are
going to be present in a dental implant? Weve evolved these five gingival
groups so you think theyre important. Well anything that requires cementum,
the implants we have now, dont have cementum on them. Theyre ankylose.
So we dont have dentogingival, we dont have dentoperiosteal and we dont
have trans-septal. All we have are circular and alveolar gingival on dental
implants. So what were really putting our money on as far as sealing all the
bad things in our oral cavity, with regards to dental implant, is that epithelial
attachment and these two gingival fibres.
Transcribed by Jazmin Lui May 5, 2014


12. Periodontal ligament fibers (section just apical)

OK so heres a forming PDL. So heres enamel up here and heres the end of
epithelial attachment that well talk about tomorrow. And heres some fibres
that are attaching to the tooth surface and going out into the gingiva. So this
must be the dentogingival group. And in humans now, not in dogs, but in
humans, theres always a space between the alveolar crest and the
cementoenamel junction or a cavosurface margin if you put a restoration that
goes further down on the root surface, further apically. And that width is
about 2 mm depending on who you read. The first person who actually came
up with this is Garicio (I think Im pronouncing his name correctly) and he
came up with about 1.5 to 2 mm. So this space, this space in humans now, is
required for some of these gingival groups. So this is called the concept of
biological width. So why is that important Dr. Craig? So in your clinical
lifetime if youre a restoring dentist, youll have caries that will come down
and youll place your cavosurface margin down here and your alveolar crest is
up here. And hopefully its not an anterior, if its an anterior tooth, you look in
there and the gingiva is always inflamed and red and very unaesthetic. So you
scratch your head and send them to your favourite periodontist, who makes
the gingiva healthy again. And he/she tells you Hey Doctor, you violated
biological width and theres not enough room for these fibres to attach, so
consequently, theyre not doing their function, and their function is to seal all
that bad microbial stuff from the oral cavity from getting into the periodontal
tissue attachment apparatus and thats happening so were going to have to
somehow recreate this 2 mm for you. And theres two ways of doing it: one
is called, its a resective surgical technique, its called a crown lengthening
procedure, where you move the gingiva all the way, take you chisels and your
burs and cut off the bone, do an osteoectomy, and you reattach the gingiva
further apically down the surface. Oh thats, especially in the anterior, thats
especially not a good thing. If youre worried about aesthetics. Is there
Transcribed by Jazmin Lui May 5, 2014

another way of gaining this 2 mm? Without picking up a scalpel blade and a
hand-piece and a chisel? The name of this lecture is PDL. PDL allows for
eruption. Wow. What if I put an orthodontic bracket on that tooth and use
the adjacent teeth for anchorage, and forcibly erupt that tooth? And if Im
doing that orthodontic tooth development in a relatively inflammation free
environment, everything is going to come with that tooth. Now Ive created a
more clinical crown. So thats called forced eruption. So thats our two bags
of tricks in your arsenal, if you need to generate additional sound tooth
structure from biological...if you need to do it by picking up a Parker blade,
doing your resective procedure. Or you can pull that tooth by forced eruption.
Questions about biologic width? This will come back over and over again,
especially if youre a restorative dentist (Student: wouldnt that just make your
tooth bigger?) but youre going to cut that down and put a crown on it,
right? Yeah you can put a crown on it. (cutting down a tooth is better than
cutting bone?). Well it depends because you cant just cut bone away,
especially in the anterior, because the gingiva is following the bone contour.
So all of a sudden, you have this nice scalloping, then all of a sudden the
gingiva is way up here. Not a great thing for a smile. In that situation perhaps
you might consider forced eruption which brings the whole tooth down right,
with the package, and now youve generated that crown length
orthodontically rather than picking up a Parker blade. Any other questions?
Biologic width. Ok.

13. Periodontal ligament cell populations (periodontal ligament fibroblast)

So whats present in the PDL? A bunch of cells, very heterozygous population
of cells if you want, PDL fibroblasts were going to talk about, well talk a little
bit about the epithelial rests of mallesez. Kind of unique to have epithelium in
a suture. Osteoblasts and cementoblasts, those are part of the lecture. Were
not going to talk about the neurovascular elements. But we are going to talk
about these little guys. There turns out to be a periodontal ligament
Transcribed by Jazmin Lui May 5, 2014

pluripotential adult stem cell population. And youre going to be making use
of it when you get into the clinic. And were going to talk about it.

14. Periodontal ligament cell populations (periodontal ligament fibroblasts
both)

So lets talk about the PDL fibroblast population. All these guys in here that
are nursing these PDL fibres. As it turns out these fibers are both synthesized
and resorbed by PDL fibroblasts as opposed to alveolar bone.

15. Periodontal ligament cell populations (periodontal ligament fibroblasts
actively)

Osteoblasts make alveolar bone, and then cells derived from the immune
system come in and become osteoclasts. So bones a little different than non-
calcified tissue. So anyways here are happy PDL fibroblasts, saw this picture
already in basic tissues. Heres the nucleus that the DNA is really opened, kind
of get the feeling this is a biosynthetically a very active cell, lots of
endoplasmic reticulum, lots of golgi. Really making lots of protein and that
protein is going into these PDL fibres. Look at these cell processes curving
around these fibres. Kind of getting a feeling that theyre sensing, those fibres
are sensing, as far as stress as concerned and theyre also synthesizing these
PDL fibres. Couple more pictures.

16. Periodontal ligament cell population (note the intimate association)
Heres some more cell processes wrapped around these developing fibres.

17. Periodontal ligament cell population (note the intimate association)
More cells wrapped around fibres. Look how many collagen molecules there
are around here. Lots of ligament around here.

Transcribed by Jazmin Lui May 5, 2014

18. Periodontal ligament cell populations (cytoplasm of PDL)

And sometimes when you look at the cytoplasm all of a sudden you see these
little inclusions, here are some inclusions, almost as if type I collagen has
gotten inside the cell. But you know that type I collagen assembles
extracellularly, not inside the cell. It looks like these little guys are getting
eaten up or perhaps dissolved.

19. Periodontal ligament cell populations (section of PDL fibroblast)

Heres a better picture. Heres a long fibre thats been degraded. Here are
some more fibres that have been degraded. So PDL fibroblasts are not only
synthesize the PDL but are also responsible degradation and turnover. And we
talked in Basic Tissues that collagen really turned over in the PDL, one of the
fastest turning over extracellular matrices in the human body.

20. Periodontal ligament cell populations (cell rests of Malassez)

Cell rests of Malassez. So they express, theyre very primitive. Theyre derived
from Hertzwigs epithelial root sheath. And they form this sort of fishnet
stocking around tooth if you will. Here are some of the cords in cross-section,
almost on the cementum surface, real close to the cementum surface, some
neurovascular bundles. And if you stain them with cytokeratins, they stain with
very primitive cytokaritins, especially C-19 is usually indicative of a very
primitive epithelium. Also if you put these guys in culture and ask, if they also
stain with other cytoskeletal elements, they also stain with vimentin, which is a
marker of mesenchymal cells. So these cells are like AC/DC in a way, epithelial
but can also be induced to become mesenchyme. So thats kind of known
now in biology, its called an epithelial-mesenchymal transition. So we
talked last week that some papers suggest that cells derived from Hertzwigs
epithelial root sheath can also undergo an epithelial mesenchymal
Transcribed by Jazmin Lui May 5, 2014

transformation and become acellular cementoblasts. And as well see in a few
minutes we can regenerate PDL, we can regenerate cementum, PDL, alveolar
bone. So there are people who believe perhaps these cell rests of mallessez in
wound healing and development also undergo an epithelial mesenchymal
transition and become the source of cementoblasts. This is very controversial
at this point in time in the profession. These guys do supply epithelium for
various kinds of pathologic cysts. And there folks in the profession that
believe if youre going to get cementum formation you have to epithelial-
mesenchymal interactions in wound healing, and perhaps these cells are the
source of the epithelial signals that are essential for cementum formation. So
no one really knows what these guys do but theyre there.

21. Periodontal ligament cell populations (the cell rests of Malassez)

Sometimes we get these serendipitous cuts. So these are a long cord of
epithelial cells that kind of demonstrate that these cells are attached in a
syncitium I cant say that theyre all attached together.

22. Periodontal ligament cell populations (high power light)

Big neurovascular bundle, lots of axons that are present in PDL. Lots of
proprioception.

23. Vascular supply of the periodontal ligament

Blood supply to PDL. Stuff you got to know. Theres 3 sources of vascularity,
the PDL. One is apically through the dental artery. Two are through
fenestrations in the alveolar bone, sometimes the bundle bone of the alveolus
proper is another synonym for it, third is the cribiform plate. This bone has all
these perforations in it and through those perforations come vascular supply
for the PDL. And the third source is through the gingiva coming through the
Transcribed by Jazmin Lui May 5, 2014

apical portion of the PDL. So the apical artery, perforations through alveolar
bone, and through the gingiva. So PDL very very well supplied with
vasculature.

24. Periodontal ligament cells (studies of)

Ok so as it turns out there are cells that are probably associated just outside
the vasculature, which really arent fibroblasts, theyre sort of primitive cells
and these are kind of pointed out by Tony Mulcher at Toronto. And then
there was a person who came along called Stewart Nyman, he was a general
dentist in northern Sweden and Stewart once said I was a general dentist for
20 years and I had 20 questions so I decided to sell my practice and get a
PhD in histology with Jan Lindhe at the University of Gutenberg in Sweden.

25. Periodontal regeneration (conventional flap surgery)

And this was Stewarts PhD thesis I think was the first demonstration of
regeneration of 3 tissues together. So anyways, this is called periodontal
regeneration. So if you have a periodontal lesion and there is an
inflammatory response that is brought in, that is elicited by gram negative
bacteria, youll hear more about this next year, and it destroys the cementum
PDL alveolar bone in that site. And if you come through with a probe you can
probe this defect and its called a pocket, a periodontal pocket, and it bleeds,
and it subterates, and theres all sorts of terrible things. Source of
inflammation. So one of the things that we do in periodontal treatment is first
off we teach the patient how to do oral hygiene to get rid of gram negative
anaerobic bacterial load. But then we have this pocket afterwards, and we
know if the pocket measures more than 5 mm or so then its really rough. Its
really hard for the patient to clean. Its really for you to get an instrument into
this in a closed procedure. So this is called a modified wooden flap. Youll
learn how to do these. Come in with an inverse bevel incision, make a cut all
Transcribed by Jazmin Lui May 5, 2014

the way to here, you pull this whole piece of inflamed tissue out, right? You
look in here you can see the root surface, now you can get in and can clean
off the affected root cementum. Its absorbed bacterial products. Do it under
direct visualization so youre much more efficient. And then at the end you
take this little flap of tissue and you kind of suture it back. And now what
happens is youve got this space. And hopefully youve got nice bleeding. So
the blood is going to clot youve got nice fibrin protein scaffold thats going
to form. And onto this protein scaffold several different proteins can migrate.
So you might get gingival epithelium to migrate along that previously
exposed root surface, you might gingival connective tissue, fibroblasts might
get into that blood clot. You might get alveolar bone cells, and perhaps you
might even get PDL cells. So what happens with conventional surgery like this


26. Periodontal regeneration (conventional flap surgery)

- is the gingival epithelium really likes to migrate and really likes to cover
wounds. Thats its job. So heres the gingival epithelium and quick like a
bunny it migrates down along previously exposed surface, covers up this
gingival connective tissue and now this epithelium attach to this root surface
by hemidesmosomes. And youre not going to get PDL, cementum, alveolar
bone regeneration. Youre going to get a scar. Thats a scar. So thats what
surgeons up to recently had. And we were happy with this. Put a probe here.
Cant get the probe in. However this thing will open up if inflammation comes
back. So this is termed long junctional epithelial attachment, long junctional
epithelial attachment. So this is what usually happens in periodontal surgery.
So along comes Stewart and the Swedes at this point in time have a very, I
dont know why, um but they have a very clear thought process. So he looks
at this and he says Gee this is like a foot race. -

27. Periodontal regeneration (guided tissue regeneration)
Transcribed by Jazmin Lui May 5, 2014

- What if I put a barrier in here so I can select for the cells I want in that
blood clot, that fibrin scaffold, and exclude the cells I dont want?. So this is a
little experimental model here, heres a tooth with periodontitis on it and
theyve done a modified Whitman and theyve cleaned the root surface and
theyve put a little notch right at the residual crest of bone. And now instead
of suturing this back in position, Stewart came in and he put in a little filter
paper. Originally it was millifore (?) filter paper. And people didnt really know
what they were doing so they put a drop of cyano acryl to hold the filter
paper in place and then they sutured everything back together again. So now
we have blood flows into the space, the blood will clot into a fibrin scaffold.
But cells from epithelium and gingival connective tissue wont have access
because they cant get across that border, barrier. But cells from the PDL can.
So this is what happens.

28. Periodontal regeneration (result after conventional periodontal flap
surgery)

This is monkey not human. But this has been done with humans a lot. Heres
the enamel space and heres the sulcular epithelium and heres the previously
exposed root surface. And they made a little notch down here, they didnt put
a barrier on this side, this is the control side. So they sutured it back together
again. And this epithelium has migrated all the way down to about here. Long
junctional epithelial attachment. These were the kinds of results that gave
Stewart the idea that perhaps there was a cell population in PDL if you gave
them the right environment they could regenerate. Cause heres the notch,
youll notice down here some fibers attaching to previously exposed notch,
and theres bone that has reformed cause remember the notch was at the
height of alveolar bone. And heres some PDL way down there thats formed.
So hes thinking Well perhaps, If I put a barrier down here, I can
preferentially allow this PDL cells to get access and exclude cells from gingival
connective tissue.
Transcribed by Jazmin Lui May 5, 2014


29. Periodontal regeneration (result after guided tissue regeneration)

So thats what you have on this slide. So heres your notch and heres your
regenerated dental cementum. And heres PDL. And heres all regenerated
alveolar bone. So this one of the first in medicine, if I understand right, one of
the first demonstrations that you could regenerate 3 connective tissues that
have a meaningful relationship with one another to have function. Youve
regenerated cementum, PDL, alveolar bone. So when I was a little periodontal
resident, I was about that time, Stewart Nyman and Tor Karring came to
Connecticut and Harold Lowe was my dean, another Scandinavian. So we had
a lot of Scandinavians coming through our department. And I remember
sitting in the back and Stewart Nyman showed this picture. And everyone was
just blown away. You cant regenerate, well I was taught in perio you can stop
disease but you cant just regenerate cementum, PDL and alveolar bone!
Thats impossible! Well its not impossible. So I was the guy in the back that
said Nehhh, Dr. Nyman, when Im looking here, that cementum is not really
attached to the previously exposed root surface. And he said Well you know,
this is an artifact But way down here, this is cementum that formed during
development and thats attached, but this cementum doesnt appear to be
very well attached Well you know so he smoked a lot and drank. I liked
him! Yes, well I wont tell you that story. So anyways, if you look at high
power this is all cellular cementum. You dont see acellular cementum here.
You see cellular cementum. Ok? So this technique is called guided tissue
regeneration.

30. Periodontal regeneration

Because youre selecting for the tissue to repopulate the wound surface. Ill
just give you some gory pictures here. This is an old picture from a alady who
participated in a study to regenerate furcations at University of Pennsylvania,
Transcribed by Jazmin Lui May 5, 2014

back when I was at University of Pennsylvania. So heres a molar, this is a big
old class 2 or perhaps even a class 3 furcation. This is a real problem I mean,
this tooth is going downhill. So this probe goes about another 5 mm further
down there, theres no bone here at all.

31. Periodontal regeneration

This is when we used goretex. It used to be called raincoats for teeth, goretex.
We stopped using the cyano acrylate on the crown but we used to suture the
living daylights out of these things. So here is the little Goretex membrane
covering site.

32. Periodontal regeneration

And if you come back 6 months, all this is hard. Alright and if you took the
tooth out youd see regenerated cementum, PDL, alveolar bone.

33. Periodontal regeneration

Heres another case, heres an upper first molar, this is harder to do.

34. Periodontal regeneration

You see, you cant move the flap because its attached. Heres a big old class
2 furcation. Big old infra-bony defect. Very ugly defect. This tooth is out of
this patients mouth. Youll have to do something.

35. Periodontal regeneration

These are hard to place up here. But they are placed.

Transcribed by Jazmin Lui May 5, 2014

36. Periodontal regeneration

So theres suturing. If you really want to get your patient upset put the knots
for your sutures on the tongue side. It will drive them nuts.

37. Periodontal regeneration

And 6 months re-entry and that furcation has filled in. Pretty amazing, so you
can actually regenerate lost PDL connective tissue attachment apparatus.

38. Periodontal regeneration (what organizes)

At this point cellular cementum, well thats pretty good! Thats a lot better
than long junctional epithelium. Can you do this in every situation Dr. Craig?
No. Very technique sensitive, it helps if you have nice surrounding walls, so
called free wall of bone. So the questions becomes, so now I know that this
population of cells in the PDL that have this ability to wander out, to migrate
out into this fibrin scaffold after surgery and differentiate into PDL alveolar
bone. This is not a tumor, this is very organized. So whats organizing this? So
people on this side of the ocean were very interested in dentin because we
felt that there must have been something in dentin that were telling these
cells Well Im dentin so you have to lay down some cementum and once
you get cementum you can get PDL. And finally you get alveolar bone, once
you get the PDL, very similar to how pelvis develops initially. Second question
is, do separate stem cell populations exist for cementum, PDL and alveolar
bone? Or is there one cell population that takes signals from the wound
healing environment and differentiates down the appropriate pathway? Or is
there a single pluripotential stem cell population and at this point now we
dont have the complete answers but Ill give you some inference.

39. Periodontal regeneration (dentin is not necessary)
Transcribed by Jazmin Lui May 5, 2014


So now another Swedish person comes in, Torr Karring and hes working with
Daniel Buser and Karen Warrer in University of Bern in Switzerland. And the
story goes that theyre studying implants and these are pressfit implants
called ITI implants, they were very big years ago. 10 years ago. 12 years ago.
He had a bunch of dental students working for him, working with him. And
the first step was to take teeth out of dogs and then heal, and put in implants
and study how implants osteointegrate. And its really hard to extract dog
teeth. They have long, spindly litte roots, and really, well theyre carnivores so
they have this really strong maxillary bone. Instead of luxating them out they
fracture them so that the roots had stayed in bone. So Karring, whos a very
interesting guy, said Well these dogs are expensive, is there some way we
make lemonade out of lemons? What well do is well take a Pressfit implant
and well make an osteotomy, to these bones that have these retained tooth
roots. Well see whats the interface on this pressfit implant. So this is the
control side where there is no extracted teeth. What youll see is areas of
bone that have fused right up to the titanium oxide implant surface. So this is
called osseous integration.

40. Periodontal regeneration (dentin is not necessary)

On the treatment side, now Torr hes supplying a different stem cell
population to this wound healing environment. So heres the roots, they really
knocked these animals really badly, and heres the pressfit implant and what
were going to do is focus down on that little area down there.

41. Periodontal regeneration (higher power view)

Heres a higher power. So heres a little root chip. And you can kind of see
this cellularity of this tissue, heres the PDL down here and it seems to be
confluent with tissue thats inbetween the implant side and the osteotomy
Transcribed by Jazmin Lui May 5, 2014

bone side. This bone is staining kind of blue-ish because its newly formed
bone. Wont go into the staining.

42. Periodontal regeneration (higher power view of implant connective tissue
interface. Note)

Heres a higher power. Heres the retained root chip. And you can kind of see
the cellularity of this tissue. And here this green stuff is the cementum from
the root and It looks like its in continuity with implant surface. And heres the
alveolar bone site.

43. Periodontal regeneration (bifringence microscopy)

So were going to jump into something called bifringence microscopy. You
can actually see fibres this way. So heres the implant surface and this
mineralized tissue and it has fibres which seem to be inserting into the tissue.
And heres the bone side. So this looks very very much like a PDL connection
apparatus. So if you supply the correct cell population to this wound healing
environment you can actually come up with dental implants that have a PDL.
Which is kind of neat. So I predict, and I always predict these things but they
never come out, so take it with a grain of salt, this is going to be the fourth
generation of dental implants. So the first implant was commercially done by
Branemark and that was commercially pure titanium and it was very fragile.
The second generation were titanium alloys to make the titanium implant a
little bit stronger. The third generation is micro-texturing of the implant
surface so you have more surface area so you have better support for osteo-
integration. Because osteo-integration is a surface phenomenon not a
biological phenomenon. And I think that the fourth generation will be folks,
perhaps, in your group, who will figure out how to get that stem cell
population out of there up onto the dental implant so you can have a dental
Transcribed by Jazmin Lui May 5, 2014

implant that actually has a PDL like natural teeth. And you can do all the
things you do with in implant that you do with natural teeth.

44. Postnatal stem cells in the PDL

So one final idea and then well take a break. So as it turns out there is a
group that has isolated human PDL stem cells and theyre working at NIDCR
intramural and they published this in the Lancet 10 years ago so this is old
history. So what they did was they took freshly extracted third molars, they
peeled off PDL, and dissociated with enzymes and put them into culture.
Some of these cells started to light up with STRO-1 and this is
unpronounceable antigen and these are associated with stem cells and
ligaments. And they started to compare these cells which they called (PDL
stem cells) against two other stem cell populations that had been
characterized. Dental pulp stem cells and bone stem cells. And theyre going
to show you that theres some similarity between these stem cells but its the
PDL stem cell that seems to make a PDL connective tissue apparatus.

45. Postnatal stem cells in the PDL

Were not going to go through everything. Theyre showing you that they can
induce these cells into different phenotypes, different differentiation pathways
depending upon how they culture them. So they cultured these cells in
corticosteroids, in pyrophosphates the dentin stem cells form these
mineralization nodules, PDL does but not nearly as well, and that kind of
makes sense cause the PDL doesnt mineralize does it?

46. Postnatal stem cells in the PDL (PDLSC can be induced)

What if you put it in an adipogenic medium? You can actually make these
PDL stem cells, if you put them into an adipogenic medium, actually
Transcribed by Jazmin Lui May 5, 2014

differentiate into adipocytes. And heres 2 markers, PPAR-gamma-2, and
lipoprotein lipoxase, these are both markers for adipocytes, and you can
actually induce these PDL stem cell populations to do this if you put them
into the right culture environment. So theyre showing you that they are
pluripotential.

47. Postnatal stem cells in the PDL (transplantation of PDLSC)

Last 2 slides then well take a break. So you can also put these human stem
cells on little hydroxyapatite particles and put into a nude mouse that lacks
ability to reject grafts. Heres the HAP particle and heres something that they
label as cementum-like and these cells seem to be inserting fibres into the
mineralized matrix. Heres the bone stem cell population, heres the
hydroxyapatite and this bone material and theyre trying to convince you
theres a periosteum forming. This is the best one, this is the dental pulp stem
cells. Heres the HAP and heres this material thats mineralizing and you can
almost see these odontogenic processes going in, looks like dentin. And since
this is derived from human, mitochondria are different in a human than in a
mouse. So if you have antibody against human mitochondrial proteins you
can this is the immune precipitate you can demonstrate that these cells
are forming cementum here because brown stain are derived from human
and not mouse.

48. Postnatal stem cells in the PDL

And finally when you create a periodontal defect like the ones were talking
about youll put in these human PDL stem cells, some of these cells will form
PDL, so heres the reaction product, showing that indeed these are from the
human and not the mouse. Some are forming cementum on the previously
exposed surface and the reaction product, the cementoblasts, showing its
from the human and not the mouse, and finally also they contribute to the
Transcribed by Jazmin Lui May 5, 2014

alveolar bone, again the reaction product is shown from human and not the
mouse. So at this point in time we think there is a single pluripotential cell
population that resides in the PDL perhaps perio-vascularly and if you treated
that PDL appropriately you can generate the cementum, PDL and alveolar
bone.

49. Summary

So I already said this. You can induce them to differentiate depending upon
what you do to these cells. Dentin surface is not essential for periodontal
connective tissue attachment regeneration, it happens on titanium.

50. Summary: PDL

What do you have to know from our discussion? The PDL is a heterogenous
tissue, kind of unique in the body. Specialized suture. It has 5 principle fiber
groups, you gotta learn them, 5 gingival fiber groups, you gotta learn them
and health PDL fibroblasts are responsible for both synthesis and degradation
of the PDL. Its highly metabolic and a multipotential stem stell population
exists. So lets take a 5 minute break and allow my voice to come and well
talk a little bit about alveolar bone in the second session.