Breast Augmentation and Reconstruction

from a Regenerative Medicine Point of View:

State of the Art and Future Perspectives

Breast reconstruction and augmentation are very common procedures, yet the prevailing current methods
utilize silicone implants that may have significant local complications requiring reoperation. Lipofillling is
increas-ingly used to contour and is considered safe, however, its utility is limited by significant volume loss.
A new approach could offer an alternative and increase the scope of patient choice. A small number of teams
around the world are investigating a breast tissue engineering (TE) paradigm. Conventional breast TE concepts
are based on seeding a scaffold with the patients’ own stem cells. However, the clinical viability of many of
these approaches is limited by their costs in relevant volumes. In this article the state of the art of tissue-
engineered breast reconstruction is reviewed and future perspectives are presented and discussed.

Introduction Breast Reconstruction Methods

Breast cancer is the most common cancer in women
around the world, making up a quarter of all female
1
cancers. According to the latest World Cancer Report
(2014) in 2012 there were 1,700,000 new cases diagnosed.
Improved survival rates make breast cancer the most prev-
alent out of any cancer, with over 6,300,000 survivors in the
1
world at any time. The lifetime risk in many developed
2
countries reaches 1 in 8. The majority of these patients will
undergo breast conserving surgery though some will require
mastectomy (with or without chemotherapy/radiation).
Some women known to be at high risk also elect to undergo
3
prophylactic mastectomy. These procedures may have a
significant psychological impact on women with concerns
4
over body image leading to anxiety and depression. Breast
reconstruction can contribute to improved psychological,
5
physical, and sexual well-being. In addition, breast aug-
mentation for women seeking to improve their body image
is one of the most popular cosmetic surgical procedures in
the United States, South-America, Europe, and Asia.
The predominant current options for breast reconstruction
postmastectomy are either autologous tissue flaps or implant
based. Flap-based surgery allows reconstruction with the
patient’s own tissue, which is moved to the defect site
producing a good aesthetic outcome and a normal feeling
breast. Flaps can be musculocutaneous such as the lattisu-
mus dorsi flap and the transverse rectus abdoninus muscle
flaps. These come with the potential of donor site morbidity
in terms of local muscle weakness and potential hernia
6
formation. These flaps are commonly pedicled, where the
blood vessels remain attached as opposed to free where the
flap is separated and attached to local blood supply via
microsurgery. The free flap is the approach taken for deep/
superficial inferior epigastric artery perforator flaps where
adipose tissue and skin is taken from the abdomen and
transplanted to the breast. There is reduced donor site
morbidity with less loss of abdominal strength, but an in-
6
creased risk of flap loss with microsurgery. Other less
common flaps such as gluteal flaps might be considered in

1
Centre in Regenerative Medicine, Institute of Health and Biomedical Innovation, Queensland University of Technology, Australia.
2
School of Medicine, University of Queensland, Brisbane, Australia.
3
Plastic and Reconstructive Surgery Unit, Princess Alexandra Hospital, Woolloongabba, Australia.
4
Department of Plastic and Hand Surgery, Klinikum rechts der Isar, Technische Universita¨t Mu¨nchen, Mu¨nchen, Germany.
5
Surg 1, Breast Endocrine Unit, Royal Brisbane and Women’s Hospital, Herston, Brisbane, Australia.
6
ARC Centre in Additive Biomanufacturing, Queensland University of Technology, Brisbane, Australia.

281
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patients with previous liposuction or abdominoplasty,
which preclude abdominal donor sites. Autologous tissue
flaps produce a more texturally natural aesthetic outcome
but require greater surgical complexity, time, and expense.
As such, this type of treatment usually requires access to
3
spe-cialized plastic surgery services.
An implant-based approach is therefore the most common
method used in breast reconstructive surgery and augmen-
tation. The International Society of Aesthetic Plastic Sur-
geons (ISAPS) provides data estimating that in 2014 there
were over 200,000 implant-based breast reconstructions and
7
1,300,000 breast augmentations performed worldwide. The
top countries for breast augmentation were the United States
and Brazil with over 300,000 and 200,000 procedures re-
spectively with other South American and European coun-tries
including Germany, Mexico, France, and Colombia each
7
performing around 50,000 such operations that year. There
are both silicone and saline implants available but silicone
implants are by far the most popular as they more closely
8
mimic the natural feel of the breast. In the United States,
around 80% of the implants used in both augmen-tation and
9 22,26
reconstruction consist of silicone. These im-plants provide
satisfying results for many of these women in terms of cost
and aesthetic outcome, yet they come with inherent mid and
long-terms risks.
Today, silicone implants are still marked by high rates of
local complications and reoperation rates that have affected
the devices since inception. These include capsular con-
tracture, implant rupture, infection, and seroma formation. The
most common complication is capsular contracture, which
occurs due to a chronic foreign body reaction against the
implant material leading to scar tissue formation, pain, and
tissue deformities. Incidence rates at 5 years with some of the
currently available implants available in the United States are
10
as high as 14% for both augmentation and re-construction.
The reoperation rate of breast surgery re-flects the frequency
of local and perioperative complications and the overall
success of the initial procedure, and around 40% of
reconstructive patients will require additional sur-gery within
11
5 years. The Kaplan–Meier cumulative inci-dence rates of
the 5 available implants in the United States are shown in
12–16
Figure 1. Interestingly, the manufactur-ers concede that
these implants should not be considered
lifetime devices, and reoperation exposes patients to un-
17,18
necessary risks and cost. Incidence of complications is
greatly elevated in patients requiring radiation therapy,
which is recommended for those with higher stage breast
10,19
cancers. The level of risk that is acceptable for a
procedure is a subject that warrants discussion between
surgeon and patient and ideally should be decided in
consultation, taking into account both the incidence and
severity of complications.
Considering the complications and reoperation rates with
silicone implants, different approaches increase the scope
of patient choice for breast augmentation and re-
construction. Other approaches include lipofilling and a
tissue engineering & regnerative medicine (TE&RM)
paradigm—each has their own downsides but a combina-
tion of these two techniques could offer a promising new
alternative. To be clinically viable it is essential to examine
the fabrication capabilities, costs, surgical considerations,
and above all safety.
VISSCHER ET AL.
Alternative Methods in Breast Reconstruction
and Augmentation
Lipofilling
Lipofilling is a technique used in breast surgery whereby
autologous fat is collected via blunt needle aspiration
(liposuction) and then injected into the area of a defect. Li-
pofilling was first perceived to have significant problems such
as trauma to cells resulting in decreased viability, calcium
deposition from fat necrosis obscuring mammography, and
20
volume loss of up to 70%. The American Society of Plastic
Surgery (ASPS) published a position article in 1987 deploring
21 21,22 23
its use due to safety concerns. Coleman and Spear
were among the first to refine the harvesting techniques and
numerous protocols and specialized equipment have now been
24
developed to ensure cellular viability with harvesting. It was
not until 2009 that the ASPS reversed their stand-point.
25
Lipofilling is inexpensive, simple, and relatively noninvasive
compared to alternatives. The most widespread use has been
to correct contour abnormalities and small de-fects but
Coleman has shown that with a number of sessions a
significant volume of tissue can be produced.
Fat grafts contain two cell groups (1) terminally differen-
tiated mature adipocytes and (2) stromal vascular fraction
(SVF) (including preadipocytes and multipotent adipose-
27
derived stem cells [ADSCs]). The proliferation and differ-
entiation of preadipocytes and ADSCs are responsible for fat
27,28
graft survival. ADSCs and preadipocytes synergistically
encourage angiogenesis and adipogenesis through the release
of growth factors and differentiation into mature fat cells.
29
A limitation of this technique remains graft volume loss of
around 30% post transplantation, which is thought to be due to
30
stress from mechanical forces. There is a maximum volume
that can be viably accommodated in the tissue space before
internal pressure causes interstitial hypertension cut-ting off
31,32
vascular supply. Using lipofilling singularly around 300
cc of lipoaspirate can produce an increase of only around one-
cup size per session (for medium size breasts), requiring
multiple sessions for greater enlargement. Increas-ing
interstitial volume, mechanical support, and vascular supply
could vastly increase the utility of this procedure.
The Yoshimura group has provided invaluable insights
33
into the fate of adipocytes after fat grafting. The fate of
adipocytes can be categorized into three zones depending
on distance from blood supply, the survival zone where
both cell types survive, regeneration zone where ADSCs
regen-erate, and the necrotic zone where both cell types
die. They acknowledge the great potential of fat cell
grafting but warn against formation of fat necrosis formed
by large grafts. In a retrospective human study they showed
that such fat ne-crosis led to long-term inflammation,
34
progressive calcifica-tion, and undesirable symptoms.
Tissue engineering and regenerative medicine
Acknowledged as one of the most promising pivotal
modules on the agenda of medical and surgical practice in
the 21st century, regenerative medicine is thought of to be
transformative in scope, equipped to add value to and ex-
pand the scope of existing models of care. By exploiting a
growing comprehension of the innate mechanisms of not
only repair but true regeneration, the emergent model of
283

FIG. 1. Kaplan–Meier Curves for cumulative risk of capsular contractures and reoperation for available implants in the United States using data compiled from the FDA,
12–16
Health Canada, and published articles (a) primary augmentation, (b) augmentation reoperation, (c) primary reconstruction, (d) reconstruction reoperation.
284
regenerative healthcare encompasses the discovery, devel-
opment, and delivery of next-generation of holistic and
evidence-based treatment concepts. Central from a transla-
tional point of view is that patient-centric regenerative
paradigms aspire not just to repair and restore the normal
structure and function of damaged, dysfunctional, or dis-
35
eased tissues but to replace them.
Under the TE paradigm a scaffold is used to support
cellular growth—often stem cells are combined with growth
factors and cultured in vitro before implantation. The con-cept
of generating new autologous breast tissue may be appealing
to the patient over insertion of a foreign object, and may
negate some common disadvantages of other im-plants. The
central problem with all implanted biomaterials is that of the
local tissue foreign body reaction, as seen with capsular
36
contracture in silicone implants. This may be addressed in
TE with the use of scaffolds that integrate with host tissue or
biodegradable scaffolds that can support growth until a stable
tissue has formed and subsequently resorb leaving only the
37
host cells.
Scaffolds that have been used in TE adipose tissue can be
divided by origin into natural or synthetic and by structure
into solid scaffolds or hydrogels. Natural scaffolds consist
largely of hydrogels, which are easily deformable with little
structural integrity, or in solid form mesh and sponges.
Collagen is a natural biopolymer providing structure to most
of the tissues in the body; it has been used in sponge form for
adipose TE showing differentiation of seeded pre-adipocytes
38,39
into viable adipocytes in vivo. However, its use has been
limited by batch-to-batch variation, cost, and immune
compatibility. Fibrin glues are hydrogels composed of natural
blood products, and they have proven effective and are already
40
FDA approved but remain costly. A common approach in
adipose TE is the use of hydrogels that mimic the extracellular
matrix (ECM) providing mechanical and chemical cues for
cellular attachment and growth. Matrigel is a basement
membrane preparation extracted from a mouse sarcoma cell
line consisting of collagen, la-minin, and perclan and Myogel
41,42
is a skeletal muscle ex-tract. They have both demonstrated
adipocytic potential in vitro and in vivo. A recent
advancement is the use of decellularized human or porcine
ECM for adipose TE. Porcine adipose tissue is readily
available in large quantities from food waste and has been
successfully decellularized, preserving structure and removing
xenogenic epitopes. It has demonstrated high stability and
43
tissue compatibility. Human adipose tissue is also available
after surgery such as liposuction and abdominoplasty and
protocols have been developed to decellurize the tissue
retaining key ECM components with the potential to offer an
44
off-the-shelf product. Han et al. demonstrated that adipose
ECM pro-duces a conducive microenvironment allowing
ADSC to recruit host cells for angiogenesis and
44
adipogenesis. Al-ternatively, the ECM products can be
delivered with a cross-
linkable vehicle to produce highly hydrated gels supporting
45,46
cell encapsulation and viability. The local biomechani-
cal and biochemical environment has been shown to be
important for adipogenesis with gels matching the stiffness
of normal fat tissue causing the upregulation of adipogenic
47
growth factors.
Hyalaronic acid is a component of ECM that can be
synthetically created and properties modified via cross-
VISSCHER ET AL.
linking. It supports development of precursor cells and has
been used to deliver adipocyte precursors supporting adi-pose
48
tissue formation. Polyethylene glycol is another commonly
used synthetic hydrogel that demonstrates great cell
49
encapsulation and adipocyte growth. Aliphatic poly-esters
are synthetic biocompatible degradable polymers that have
proven popular in a number engineering appli-cations and
allow the production of specific shaped struc-tural scaffolds
via additive manufacturing. Polyglycolactide (PGA) and
polylactide and their copolymer have been in-vestigated with
degradation times of 6–18 months consistent with past views
that scaffolds should degrade with tissue ingrowth.
Polycaprolactone (PCL) resists degradation for slightly longer
persisting for >2 years, which is in keeping with more recent
concepts that scaffold support is required until mature tissue is
50
formed.
Hydrogels have proven successful in adipose TE and show
great promise for translation for use in soft tissue
reconstruction as an alternative to current techniques and
fillers. However, with current technology the ability to scale
such hydrogels to provide adequate volume, shape, and
structure for reconstruction of the entire breast mound at a
viable price is questionable. It is for this reason that our group
advocates the use of degradable aliphatic polyesters.
The utility of these materials in TE has been aided by the
rise of additive manufacturing (three-dimensional [3D]
printing), which allows the production of scaffolds with
customizable micro and macro-architecture in a number of
different biocompatible materials.
Any solid biomaterial to be used in breast regeneration
must be biocompatible, sterilizable, and deformable so as to
be comfortable but also robust enough to maintain its shape
37
and support growth. In addition, it should allow further
diagnostic imaging for early stage breast cancer detection. To
be acceptable to the patient the material should have a similar
consistency to the normal breast. The use of a bio-material
that is already FDA approved for use in implan-tation would
also fast-track the translation toward commercialization and
clinical use. The table below shows the mechanical properties
of the major components of the breast tissues compared to
some of the clinically used bio-degradable and bioresorbable
polymers already approved in many medical devices and
implants (Table 1).
51
Configured into a highly porous architecture scaffolds can
be fabricated to have mechanical properties similar to the
suspensory ligaments that are the main components pro-
viding structural support to the breast, with the potential to
feel natural on palpation.
52,53
Importantly, the quest is not to
replicate/copy the physical structure of the tissue to be re-generated but to
guide the regeneration process by designing a scaffold architecture, which
inherits an anchoring presence to guide the tissue formation. Aliphatic
polyesters such as PCL degrade via the Krebs cycle and are metabolized to
lactate and other acidic monomers, which are easily pro-cessed by natural
metabolic pathways in the body.54 Fast degrading polymers such as PGA and
PLGA may create a local acidic environment, which can have adverse tissue
effects.54 In contrast, PCL degrades slowly over the course of a number of
years, and provides adequate structural support for not only regeneration but
also remodeling into functional tissue, while maintaining physiological pH in
the surrounding tissue.
55
BREAST TISSUE ENGINEERING
Table 1. The Mechanical Properties of Major Components of the Breast and Common Biopolymers
Component of breast tissue Elastic moduli (GPa)
-6
Adipose tissue 0.5–25 · 10
-6
Glandular tissue 2–66 · 10
Suspensory ligaments 0.04–0.4 (40% decrease with age)
Adapted from Gefen and Dilmoney52 and Chhaya et al.53
There is significant heterogeneity in breast shape and
composition between women, and personal differences in
the desired outcome for both augmentation and recon-
53,56,57
struction. To reflect these differences scaffolds can
be tailor-made via 3D imaging software and additive
53,56,57
manufacturing. Silicone implants offer limited choi-
ces in consistency and are mostly targeted at recreating the
58
look and feel of a youthful breast. An approach that pro-
vides the possibility of a more naturally shaped and feeling
breast for their age may encourage more widespread uptake in
older women many of whom currently reject recon-
59
struction. A few years ago the idea of mass production via
additive manufacturing might have been questionable but the
rapid development of the technology in this field is promising.
Just last year the team at carbon-3D announced their new
approach to stereolithography (SLA) called con-tinuous liquid
60
interface production to much acclaim. This technology
promises much faster production than traditional SLA and in
their patent application they even suggest that the technology
can be used for printing biodegradable resins previously not
61
widely available in this line of printing.
Cells
Since the pioneers of the field Langer, Vacanti, and Atala
62
first considered the approach in 1994 there have been only a
small number of teams investigating a regenerative medicine-
based breast TE concept. In contrast many more teams have
conducted main stream research in adipose TE. Not surprising
from a clinical point of view little progress
63
has been made with as of yet only one human trial ; and only
64–66 67,68
one Australian and a German/Australian team
have succeeded in regenerating clinically relevant volumes of
tissue. The key problem in TE&RM is the regeneration of a
functional vascular network into the tissue-engineered
construct (TEC). This is especially important in adipose TE,
with fat cells being highly metabolically active and under-
69
going necrosis when not adequately supplied. In a com-
prehensive review in 2004, the lack of the TE research
community to reach clinical translation was acknowledged
and the so far published studies were categorized into two
approaches (1) those isolating and culturing preadipocytes for
implantation and (2) those using the tissue growth fac-tors to
20
recruit resident preadipocytes. In the last 10 years these
methods have remained similar and many groups have
combined them, commonly utilizing growth factors like FGF-
2, IGF-1, and VEGF, matrigel (a combination of ECM/ growth
factors), ADSCs, preadipocytes, HUVECs (endothe-
lial stem cells), and the use of bioreactors to promote an-
65,70–76
giogenesis and adipogenesis. ADSCs are commonly
isolated from the SVF via adherence and cultured before
implantation. While many of these studies have demonstrated
285
Biomaterial Elastic moduli (GPa)
Poly-D,L-lactide 1.5–1.9
Poly-L-lactide 1.4–2
Polycaprolactone 0.31
the possibility of producing a vascularized adipose construct
they have been limited to constructs of small volume—pre-
dominantly <2 cc that is clearly not clinically relevant to
breast constructs, which would require at least >80 cc to
77
match the smallest commercially available implants. One of
the major limitations in scaling up experiments is cost of these
growth factors and particularly of culturing the stem cells that
53
require certified laboratories. This poses serious concerns
for the commercial viability of such approaches on a larger
scale. Such techniques would also require extensive
investigation before FDA approval for human use to prove
that the use of such growth factors and stem cells did not
encourage malignant growth. In a promising strategy Witt-
man et al. demonstrated successful adipose tissue growth in a
mouse model using the entire SVF in fibrin gels, without
40
preculture of cells. This technique is readily translatable and
has the potential to allow a one-step procedure in the oper-
ating theater with the implantation of isolated SVF combined
with TissuCol (FDA-approved fibrin gel). The authors sug-
gest that the precursor cells might allow simultaneous an-
giogenesis and adipogenesis.
A microsurgical driven concept to vascularization is the
use of an arterio-venous (AV) loop. This has been used by
the Morrison group to produce a clinically relevant volume
of adipose tissue, being one of only two teams to achieve
this in an animal model and the first group to publish a
63,64
human trial in March this year. In results published in
Plastic and Reconstructive Surgery (2011) they demon-strated
the production of 80 mL of soft tissue by encasing an AV loop
(alone, with a muscle flap, with a fat autograft, or with a fat
flap) and a sponge scaffold in a perforated hard plastic
chamber, and implanting it subcutaneously in a pig model.
The pedicled fat flap (starting volume 5 mL) was the only
group to produce significant adipose volumes with eight
samples filling the 80 mL hard chambers with an av-erage
volume of *25 mL fat tissue (1/3) and the other 2/3 of the
volume consisting of fibrovascular tissue. A single sample was
left in situ for 22 weeks, and after removing the hard casing at
12 weeks there seemed to have been fibro-vascular regression
leaving a volume of *60 mL consisting almost entirely of
adipose tissue. The authors noted that the lack of adipose
tissue growth in the fat autograft group (li-pofilling) compared
to the fat flap, was likely due to the autograft being placed on
a two-dimensional vascular ped-icle rather than in a
vascularized 3D space. In a reply in the same journal Yuan
suggests that the success of the approach with fat flap pedicles
was due to the modulation of physical forces promoting
78
adipogenesis. In other work Yuan has shown that
adipogenesis is inhibited by mechanical forces in contrast to
musculoskeletal tissues that are known to pro-liferate in
response to physical stress—this is consistent with the
79
observation of volume loss in lipofilling. In a similar
286
FIG. 2. A three-dimensional scaffold of clinically relevant volume with lipoaspirate was implanted in a large animal (mini
pig) model. (a) PCL scaffold with specifically designed voids filled with lipaspirate preimplantation. (b) After 8 months
implantation a histological image stained with Goldner’s trichrome demonstrating PCL scaffold struts ( ), ingrowth of
+
highly structured fibrous tissue ( ), and adipose tissue survival (*). PCL, polycaprolactone.
large animal model our group has produced porous PCL
scaffolds, which allowed for support of adipose tissue growth
and highly structured fibrous tissue ingrowth (Fig. 2). The
formation of such a large specific 3D shape is simply not
possible with lipofilling alone. While the work of the
Morrison group is significant as the first group to engi-neer
clinically relevant volumes of adipose tissue its wide-spread
clinical application is questionable. In January this year they
published a proof-of-concept pilot study in human volunteers,
using the same approach of vascularized fat pedicles inside
hard plastic casing (this time sans sponge) of various volumes
63
in five patients with unilateral mastecto-mies. In one of the
patients the fat flap grew from 30 mL to fill the 210 mL
encasing, which remained stable for 12 months inside the
encasing and retained some volume for 6 months following
removal. In surgery the tissue was re-ported to resemble partly
fat and partly fibrous tissue. In the other subjects, three
showed only fibrous and no adipose growth and one patient
pulled out of the trial due to pain from the implant at 7 weeks.
The reasons for adipose tissue growth in only one of the
patients are unclear but the authors suggest the contrast in
results compared to their animal trial may be because pigs
continue to grow throughout life whereas humans do not. The
formation of thick fibrous capsules in 3/4 of the human
patients is significant where the primary aim was to grow
adipose tissue; this is a recurring issue in large animal studies
and something future research in the field needs to address.
The use of hard plastic casing in this approach necessitates
reoperation for removal, and may also be uncomfortable for
the patient, as demonstrated by one of the patients
withdrawing. It would seem more practical to have an
implantable degradable scaffold alone with sufficient strength
to resist shear forces.
Combining lipofilling and scaffold-based TE
The central issues facing lipofilling are volume loss due to
internal space limitations and compromised vascular supply.
VISSCHER ET AL.
#
For TE, it is the impracticality of up-scaling current ap-
proaches. A common TE approach is to isolate stem cells and
culture them before reimplantation—the idea being that they
20
are able to proliferate unlike mature adipocytes. However,
the relative success of lipofilling suggests that this may be an
unnecessary step. The lipoaspirate following simple on-site
concentration protocols may be rich enough in these stem cells
to promote adipose regeneration—and this is a more cost-
effective approach. Although the stem cells are more resistant
to reduced vascular supply, mature adipocytes too may be able
to survive when sufficient vascularization is maintained. Ad-
ditionally, the risk of injecting a cocktail of stem cell-enriched
cells into a breast environment might trigger or activate the
80
dormant cancer cells in a breast cancer patient.
Khouri and Vecchio suggests that with advances in lipoas-
pirate harvesting protocols the current limiting factor in graft
survival is the recipient site rather than the graft material, with
31
cell survival dependent on distribution. This is consistent
with the observation that smaller grafts have higher survival
rates due to a higher surface to volume ratio to the vascular
27
bed. He advocates the use of his technology Brava (Brava,
LLC, Miami, FL) a vacuum-based external soft-tissue ex-
pansion system to aid fat grafting, whereby a pump attached to
polyurethane domes applies negative pressure to the breast to
encourage micro-angiogenesis and volume expansion with
31
stretching of the skin envelope. Breast reconstruction using
injected lipoaspirate is technically possible even without
Brava technology through multiple lipotransfer sessions.
However, the result is sometimes difficult to predict and re-
sorption rates of the fat is different in specific anatomic com-
81
partments of the breast.
A more practical concept to increase available internal
space and vascular supply is the implantation of a patient-
specific scaffold.68 This allows the body to act as a biore-actor,
allowing tissue ingrowth and establishment of a vas-cular
supply before injection of the lipoaspirate (Fig. 3). 82 This
ensures that angiogenesis precedes adipogenesis, which is a
critical step in adipose tissue regeneration.20 The scaffold
BREAST TISSUE ENGINEERING
also provides mechanical support encouraging adipose pro-
liferation. This approach combines TE and lipofilling
aspects with each accounting for the others disadvantages.
It is important to distinguish between augmentation/skin
sparing mastectomy and full mastectomy with loss of the
skin envelope. In the latter procedure there is very little
volume remaining under the skin, and expandable implants
are required to promote stretching of the skin. This puts
great mechanical pressure on the area such that lipofilling
83
alone is not viable. It is possible the inclusion of a
scaffold could provide the required mechanical support for
tissue growth after expansion. Different protocols will need
to be established for the use of TE scaffolds depending on
the surgical procedure.
Our group has demonstrated successful growth of large
volume adipose tissue in a pilot study of four pigs with a
FIG. 4. Protocol steps and equipment for our large animal breast TE study. (a) Liposuction technique for lipoaspirate
harvesting. (b) Next generation PCL breast scaffold. (c) Preparation of a subcutaneous pocket for the scaffold with an
implanted scaffold in the background.
287
FIG. 3. A breast before
mastectomy (a), postmastec-
tomy with preimplantation of
a TE scaffold (b), allowing
vascular ingrowth before li-
pofilling (c), with later deg-
radation of the scaffold (d).
TE, tissue engineering.
67
unique concept of delayed fat injection. Porous degradable
PCL scaffolds of clinically relevant volume (75 cc) were
produced via melt extrusion by Osteopore according to
international standards, and implanted subcutaneously in pigs
for 24 weeks. Autologous fat graft was harvested and 4 mL
injected into the implanted scaffold either immediately or after
a 2 weeks delay in a separate group. This delay was thought to
allow prevascularization and this group produced the greatest
change with a sixfold increase in adipose tissue to give almost
30 mL. The tissue in the scaffold consisted of around 47%
67
adipose tissue, similar to the natural breast tissue control.
These results show proof of principle for our approach to
breast TE. Our group has now started a high powered large
animal study using next generation scaffolds and more
advanced fat grafting techniques to pave the way for human
trials (Fig. 4).
288
Safety
Patient safety in any development of a new treatment
concept in medicine must be the primary consideration.
Important considerations for local breast cancer recurrence
are factors that might impede detection or promote malig-
nant regrowth.
Cancer detection
Potential lipofilling complications may be reduced with
correct harvesting and injection technique and the introduc-
tion of specialized devices, although there is still a risk of fat
84
necrosis, oil cyst formation, and calcification. Previously,
there were concerns that these secondary changes would be
indistinguishable from malignant recurrence on mammo-
85
graphy and MRI. In 2009 the ASPS Fat Graft Taskforce
found no evidence that these changes interfere with cancer
imaging, however, they suggested that more studies were
25
needed to confirm these findings. Since then comprehen-
84
sive reviews by Mizuno and Hyakusoku and Kasem et al.
28
concluded that modern imaging techniques and experienced
radiologists can easily distinguish between these lesions. It has
since been shown that any breast operation may result in the
formation of calcification, but lipofilling actually pro-duces
less than many other common procedures.
85
The addition of a TEC could also impede the detection of
new malignant growth. These scaffolds likely share some risks
with silicone implants—which compress tissue, cause scar
formation, and increase the radio-density thus interfering with
identifying lesions. Silicone implants have been shown to
obscure mammographic imaging of the breast, however, this
has not translated into any change in morbidity or mor-tality
with patients presenting at the similar stages of breast
86
cancer. This is believed to be due to implants—especially
those subpectorally located, causing atrophy of the sub-
glandular breast parenchyma facilitating palpation of new
86,87
lesions. It is important to recognize key differences be-
tween a silicone implant and a scaffold—which will have
significant implications on risk. A porous scaffold would be
less radiopaque than a silicone implant, and produce less scar
tissue allowing better visualision of surrounding tissue, after
degradation the scaffold would cease to interfere with im-
aging entirely. However, it could also obscure new growth
within its boundaries. A scaffold would also be less uniform in
texture possibly reducing the sensitivity of a manual breast
examination. There are no studies specifically examining this
risk, and this is an issue that needs consideration in the future.
Cancer recurrence
There are serious theoretical concerns that lipofilling
may encourage cancerous recurrence or new growth,
because its regenerative effects are based on the same
hormones, growth factors, and stem cells that have been
shown to promote cancer progression and neoplastic
88
angiogenesis in laboratory research.
Donnenberg et al. reviewed animal and cell culture models
80
to question the safety of regenerative therapy after cancer.
Their conclusions delineate a difference between dormant
cancer cells and high-grade tumors, suggesting that only the
growth of these high-grade cells is encouraged by regenerative
80
efforts. They conclude that attempted tissue-
VISSCHER ET AL.
engineered reconstruction should be delayed until cancer
remission is firmly established—which would seem intuitive
at the outset. The possible correlation between their cancer
cell types and breast cancer clinical staging or histological
grading is uncertain and so its clinical relevance is unclear.
In a retrospective cohort study of 646 lipofilling proce-
dures Petit et al. found a small increase in locoregional
recurrence rates with breast-conserving surgery but not for
mastectomy compared to rates from the European Institute
89
of Oncology. They advise that definitive conclusions
cannot be made from separate retrospective studies and
advocate the need for larger prospective trials or a registry.
Another smaller observational study followed the onco-
logic outcomes of 60 patients over 5 years and stratified
88
them according to TNM cancer stage. Their long-term
results suggest that lipofilling is safe for treating women
after mastectomy for stage 1 breast cancer and most likely
stage 2. It may even be possible for stage 3 patients after
radiotherapy if the pathologic grading is prognostically fa-
vorable; their results indicate that triple negative breast
cancers should be avoided. This suggests possible correla-
tion with the classification of high-grade tumor cells by
Donnenberg et al.
In a comprehensive systematic review in 2013 Krastev et
al. analyzed 20 clinical trials of autologous lipoaspirate
grafting in the breast and found overall no significant dif-
ferences in locoregional recurrence in either mastectomy or
90
breast-conserving treatment. The authors stress that these
results are not conclusive and that larger prospective trials
with longer follow-up are required. The likely reason there
are so few studies available is because the ASPS dismissed
the technique until a few years ago.
The theoretical risks of lipofilling encouraging
malignant growth are concerning. There is currently no
high level evidence suggesting significant increases in
breast cancer recurrence post lipofilling, but there is
limited research in the area. The evidence supporting its
28
safety is sparse but slowly accumulating. There is a clear
need for further research with high quality trials to
establish the safety of this promising technique. The
clinical consensus suggest that lipofilling is safe, in
patients with determined to have a low risk of recurrence.
Surgical Considerations
The risks of encouraging cancer recurrence and hindering
detection would change depending on placement of the con-
struct. Traditionally implants for breast augmentation were placed
subglandularly—in theory the best position for natural shape and
form of the breast, more comfortable for the patient and sparing
91
incision of the pectoralis. This placement position has fallen out
of favor with silicone implants due to the finding of increased
capsular contracture rates. Current recommenda-tions are for both
augmentation and reconstructive patients to have implants
inserted subpectorally, with the alternative option of subfascial
92
placement for augmentation. This ap-proach with a TE
construct that aims to reduce the incidence of capsular contracture
through degradation of the implant would once again offer the
possibility of subglandular place-ment. The safest option for a TE
construct would be to place it subpectorally—separating it with a
layer of muscle from the glandular tissue from where breast
cancer is thought to
BREAST TISSUE ENGINEERING
originate. This should also provide better blood supply as
muscle has greater vascularization than adipose tissue. How-
ever, this may be more uncomfortable for the patient, and will
place increased mechanical stress on the construct, which
would likely not be conducive to long-term adipose tissue re-
tention. The choice may also take into account whether the
indication is reconstruction or augmentation, which tradition-
ally have different aims. Reconstruction usually aims to create
bilateral medium-sized breast to restore anatomical shape but
with the future risk of cancer recurrence being the primary
93
concern. This is in contrast to augmentation, which is per-
formed for aesthetic reasons, designed to the patients’ prefer-
ences. The placement choice remains a dilemma that needs to
be agreed in consultation between bioengineers, surgeons, and
the patient.
Future Directions
Breast conserving surgery applications
An approach combining patient-specific scaffolds and
lipofilling also offers a new alternative in reconstruction after
breast-conserving surgery that silicone implants could never
provide. Breast conserving surgery consists of a combination
of lumpectomy, and radiation therapy—and is often the
87
preferred treatment choice in small early stage breast cancer.
Currently, conservation is performed in over 60% of breast
94
cancer surgeries. Lumpectomy volumes of greater than 15%
often result in unsatisfactory aesthetic outcomes and may
require oncoplastic procedures with volume displacement
(larger breasts to fill defect) or re-placement (smaller breasts,
94
with fat flaps). Irradiation may compromise cosmesis for
silicon-based implant reconstruc-tion and autologous flaps
26,95
may be preferred in some cir-cumstances. The concept of
autologous fat grafting to lumpectomy defects has been
recognized as an exciting approach, as alternative techniques
are inappropriate and because what is surgically removed is
26
mostly adipose tis-sue. The combination of a TE approach
and lipofilling is especially appealing—as construct of any
size can be de-signed and the scaffold can provide support to
the adipose cells maintaining the desired shape.
Additional breast reconstruction applications
A TE approach offers advantages over traditional tech-
niques for breast reconstruction in a number of different
applications in addition to breast cancer surgery. A number of
congenital conditions, trauma, surgical complications, and
severe capsular contracture can lead to significant asymmetric
deformation. Poland syndrome is a unilateral congenital
deformity affecting 1/10,000 women where the sternal portion
of the pectoralis major and the anterior mammary fold are
96
absent commonly presenting with fibrotic breast tissue.
Tuberous breast deformity is another con-genital anomaly
where the breast fails to develop properly and results in
97
hypolastic breast mounds that may be asym-metrical. The
correction of these deformities presents a challenge to the
plastic surgeon who must either use a complicated series of
tissue flaps or custom-made silicone implants. The ability to
custom design and print TE scaf-folds for an individual patient
has the potential to simplify operations and improve outcomes
for these patients.
289
Nipple areola complex reconstruction
The nipple areola complex (NAC) is central to the aes-
thetics of the breast. It is ideally conserved in a nipple-sparing
mastectomy, but depending on the location and stage of breast
cancer this is not always possible. The NAC can also be
deformed as a result of infection, trauma, or burns. In
conventional surgery the color and texture of the areola can be
mimicked via tattooing or skin grafting. The projection of the
nipple is usually achieved via one of a series of flaps that are
98
mostly derivatives of the same design. However, a common
complication is projection loss and around a third of patients
99
rate their NAC reconstruction as fair to poor. Techniques
aimed to maintain projection in-clude use of a cartilage graft,
alloderm, or a silicone spacer.
This represents a challenge that could be addressed by
TE. While TE commonly aims to replace the lost tissue
with the same tissue type, in this instance an aesthetic
recreation of a certain shape is required with tattooing able
to provide color. TE approaches to NAC reconstruction are
few and far between. In 1998 Cao et al. used pluronic F-
127 to deliver chondrocytes into a pig in a subcutaneous
pouch surrounded by a circumferential suture to produce
100
projection. More recently Pashos et al. presented an
abstract detailing pre-liminary in vitro results for
decellularizing the nipples of Rhesus monkeys for use as a
101
TE scaffold on which to seed the patient’s own cells.
Pashos et al. has founded a startup BioAesthetics to bring
the product to market, although regulatory and ethical
hurdles remain. Another startup TeVido biodevices also
promises patients tissue-engineered personalized nipples
but have yet to ship product. Our group is investigating the
potential for biodegradable polymer-based scaffolds for
nipple reconstruction, which can be designed to specific
size requirements and could be seeded with a number of
different cell types including fat graft or chondrocytes.
Therapeutics
The use of lipoaspirate shows promise not only for breast
reconstruction but also could concomitantly act as a therapy
for treating the side effects of radiation therapy. Post-
mastectomy radiation is commonly recommended in patients
with locally advanced breast cancer, large tumors, or with
19
significant lyphovascular/lymph node involvement. How-
ever, it is associated with increased complication rates and
19
reconstructive failure in all current methods. The radiation
targets the rapidly dividing cancer cells disrupting replication
and cellular architecture, but results in unavoidable damage to
the surrounding normal local tissue causing significant der-
matitis, subcutaneous fibrosis, skin hyperpigmentation, and
26,86
impaired healing. This can lead to a self-maintaining
pathological condition that can last for years; its
pathogenesis is thought to a vicious cycle of vessel injury,
29
hyperperme-ability, altered blood flow, and ischemia.
In a study Rigotti et al. showed that the transplantation
of lipoaspirate into chronic radiotherapy induced tissue
injury can lead to both clinical and microscopic
improvement, with revascularization and restoration of
29
function. This was thought to be due to the presence of
the ADSC in the lipoaspirate, which are known to induce
neovascularization. They are theorized to improve the
29
capillary: adipocyte ratio breaking the cycle of damage.
290
FIG. 5. Treatment concept integrating multidisciplinary expertise in tissue engineering and lipofilling.
Interestingly, others have shown that ADSCs retain their
regenerative potential after chemotherapeutic treatments,
102
which are also clinically relevant in breast cancer.
Multiphasic biomaterial and scaffold-design
There are numerous potential advantages that could be
incorporated into this approach in the future. One group
has proposed the use of tannic acid cross-linked to collagen
as a scaffold material that is thought to have antitumor
103
properties and could prevent cancer recurrence. Another
group claims to be developing technology with both
therapeutic and diagnostic capacities, with the delivery of
chemothera-peutic drugs and/or tumor-detecting chemicals
on breast prosthesis. This has the potential not only to
37
reduce cancer recurrence but also aid early detection.
In our group, a holistic approach to breast TE has been conceived
based on our multidisciplinary expertise profile (Fig. 5). This
treatment concept involves scanning/imaging of patients,
multifunctional scaffold design, fabrication, and im-plantation
together with lipofilling. Imaging of the breasts (presurgery) or
contralateral breast after a single mastectomy translated into a
computer aided design format allows for the design of a breast
scaffold to match the patients natural breast shape, with the added
potential for the patient to change size, projection, and shape.
Multifunctional scaffolds being investigated support the growth of
adipose tissue with local drug delivery for chemotherapy, infection
prophylaxis, or adipocytic differentiation. The aim is to achieve
highly patient-specific treatment with significantly improved patient
outcome.
VISSCHER ET AL.
Conclusion
The concept of breast reconstruction and augmentation
utilizing the combination of TE principles and lipofilling
offers a promising alternative to silicone implants and to
current techniques using free flaps. Clinically, lipofilling is
established as an effective and safe technique yet with sig-
nificant limitations in large tissue volume generation that
could theoretically be addressed by the preimplantation of
a scaffold to provide mechanical support and a vascular
sup-ply. The materials are currently available to fabricate a
biodegradable implant that should address the issue of local
tissue reaction that plagues current implants. The current
evidence supports the safety of this approach, although this
is an area that requires further and ongoing investigation
due to serious theoretical risks. This approach also has the
ca-pacity to be therapeutic in postradiation injury and the
possibility to be used after breast conserving surgery. Fi-
nally, there is ongoing research that could yield useful
therapeutic and diagnostic additions to this approach. This
evidence points to the need for large-scale animal trials of
lipofilling with clinically relevant implant scaffold volumes
to pave the way for human trials and clinical translation.
Acknowledgments
Some of the work and figures shown in the review was
supported by the National Breast Cancer Foundation
(NBCF IN-15-047 to D.W.H.) Australian Research Council
(ARC Training Centre in Additive Biomanufacturing -
IC160100 026 to D.W.H.).