EXPERIMENTAL

Temporary Angiogenic Transformation of the

Skin Graft Vasculature after Reperfusion
Nicole Lindenblatt, M.D.
Background: In the era of tissue engineering, the physiologic process of skin
Uwe Platz graft revascularization remains unclear, preventing the successful development
Martina Althaus of skin substitutes. Therefore, the authors developed a new in vivo model with
Niels Hegland which to visualize the process of engraftment and its microvascular architecture.
Christian A. Schmidt, M.D., The aim of this study was to specifically investigate the vascular transformations
Ph.D. within the skin graft to gain applicable knowledge on how vascular processes
Claudio Contaldo, M.D. during engraftment occur.
Brigitte Vollmar, M.D. Methods: Microsurgical preparation of the modified dorsal skinfold chamber
Pietro Giovanoli, M.D. including autologous skin grafting was performed in male C57BL/6J mice (n ⫽
Maurizio Calcagni, M.D. 10). In addition, immunohistochemistry of angiogenic factors, endothelial cells,
Zürich, Switzerland; and Rostock, and pericytes, and corrosion casting were performed to further characterize the
Germany specific mechanisms.
Results: The graft exhibited capillary widening starting at day 3, resulting in the
temporary formation of spherical protrusions at the graft capillary divisions
starting in the center of the graft 24 to 48 hours after revascularization. Confocal
microscopy showed the simultaneous expression of CD31 and desmin. Corro-
sion casting and evaluation by light microscopy and scanning electron micros-
copy showed the three-dimensional formation of capillaries in the wound bed
that connected to the preexisting capillary loops of the skin graft.
Conclusions: The authors were able to show for the first time a temporary
angiogenic response within the capillaries of the skin graft. This most likely
represents a reaction to reperfusion allowing the supply of proangiogenic factors
to the hypoxic skin graft. The detection of an angiogenic response within the
graft capillaries is for the first time made possible in the newly developed model
and is therefore completely novel. (Plast. Reconstr. Surg. 126: 61, 2010.)

S
kin grafting still represents the preferred cause long-term survival of cells within these con-
technique for covering skin defects.1 Despite structs demands the rapid establishment of ade-
the widespread use of this technique, we still quate blood supply.4 As a consequence, strategies
do not fully understand the precise cellular and are now directed toward engineering of pre-
vascular mechanisms involved with graft survival formed microvascular networks within tissue-en-
and incorporation. The development of tissue- gineered constructs.5 These developments sup-
engineered skin substitutes has reanimated the port efforts to understand the physiologic process
discussion about graft revascularization, because of graft revascularization and to establish new
these constructs still lack reliable incorporation models with which to study the revascularization
into the human body.2,3 This is in most cases be- of skin grafts or skin substitutes in vivo. Most re-

From the Division of Plastic and Reconstructive Surgery and

the Division of Cardiovascular Surgery, Department of Sur-
gery, University Hospital Zurich, and the Institute for Ex-
perimental Surgery, University of Rostock. Disclosure: This study was supported financially
Received for publication October 20, 2009; accepted Decem- by the Hartmann-Muller-Foundation, Zurich, Swit-
ber 10, 2009. zerland, and the FORUN Program of the University
Presented in part at the European Congress of Scientists and of Rostock, Rostock, Germany. The authors declare
Plastic Surgeons, in Bern, Switzerland, September 12 that they have no competing financial interests,
through 13, 2008, and at the European Plastic Surgery commercial associations, or financial disclosures
Research Council, in Hamburg, Germany, August of 2009. that might pose or create a conflict of interest with
Copyright ©2010 by the American Society of Plastic Surgeons information presented in the article.
DOI: 10.1097/PRS.0b013e3181da87f6

www.PRSJournal.com 61
 Plastic and Reconstructive Surgery • July 2010

cently, we developed a new model allowing for the Intravital Microscopy

simultaneous in vivo visualization of the microcir- Repetitive intravital microscopic analyses of
culation of wound bed and skin graft. Using this the skin graft were carried out daily over a period
approach, we were able to show that angiogenesis of 10 days (n ⫽ 4). Microscopic images were taken
within the wound bed resulted in reperfusion of at four different areas within the center and the
the graft capillaries at day 3, with almost complete periphery of the graft, respectively (Fig. 2). After
restoration of the skin microcirculation at day 5.6 injection of 0.2 ml of fluorescein isothiocyanate–
Based on these first results, we assumed an labeled dextran (2%; molecular weight, 150,000;
ingrowth of newly formed blood vessels from the Sigma-Aldrich, Munich, Germany), the microcir-
wound bed into the graft and subsequent connec- culation of the skin graft was visualized by intravital
tion to the existing vasculature rather than a direct fluorescence microscopy (Leica DM/LM; Leica Mi-
early linkage of existing bed and graft vessels, as crosystems, Wetzlar, Germany). Microscopic images
several authors have proposed before.7–9 However, were captured by a charge-coupled device television
the fate of the graft vasculature has still to be camera (Kappa Messtechnik, Gleichen, Germany)
clarified. It is on the one hand thinkable that vessels and recorded on video (50 Hz; AG-7350-SVHS; Pa-
from the wound bed connect directly to the existing nasonic, Tokyo, Japan) for subsequent off-line anal-
graft vessels without further development of the vas- ysis. Using 10⫻ (N-Plan 10⫻/0.25 LD; Leica), 20⫻
cular system. In this case, the exact mechanisms (HCX Apo 20⫻/0.50 W; Leica), 40⫻ (HCX Apo
would be of major interest, representing a relevant 40⫻/0.8 W; Leica), and 63⫻ (HCX APO L63⫻/
phenomenon of vessel interconnection.10 In con- 0.90 W; Leica) objectives, blood flow was monitored
trast, previous studies suggested that graft vessels in capillaries of the superficial and deep dermal
may degenerate after reperfusion, leaving the base- plexus of the skin graft.
ment membrane as a potential conduit for further
vessel ingrowth.11–13 Therefore, it was the aim of this Microcirculatory Analysis
study to gain further insight into the morphology
and development of the vascular structures within Microvascular perfusion was quantified off-
the skin graft after reperfusion and to three-dimen- line by analysis of the videotaped images using a
sionally visualize its capillary network. computer-assisted image analysis system (CapIm-
age; Zeintl Software, Heidelberg, Germany). This
included the determination of functional capillary
MATERIALS AND METHODS density (in centimeters per square centimeter),
vessel diameter (in microns), and red blood cell
Mouse Modified Dorsal Skinfold Chamber velocity (in microns per second). Furthermore,
On approval by the local government, all exper- morphology of vascular changes within the skin
iments were carried out in accordance with the Ger- graft were assessed and quantified as number per
man and Swiss legislation on protection of animals. area (square millimeters) of observation.
Male C57BL/6J mice with a body weight of 23 to 27 g
(n ⫽ 10) were anesthetized by intraperitoneal in- Immunohistochemistry
jection of ketamine (90 mg/kg body weight) and Five days after transplantation, cross-section tis-
xylazine (25 mg/kg body weight). sue blocks of the modified dorsal skinfold chamber
To study the revascularization process, we were harvested. Four- (n ⫽ 3) and 25-␮m (n ⫽ 3)
used the modified dorsal skinfold chamber, as sections were cut. The 4-␮m sections were treated
recently published by our study group.6 On day 3 by microwave for antigen unmasking and by per-
after the dorsal skinfold chamber preparation, oxidase and protein block. Goat anti–Ang-1 (1:
skin and most parts of the hypodermal fat layer 200), mouse anti–Flk-1 (1:200), rabbit anti–Flt-1
were carefully removed in a circular area 7 mm in (1:200), and rabbit anti-bFGF (1:200) (all from
diameter from the back of the chamber, leaving Santa Cruz Biotechnology, Santa Cruz, Calif.)
the panniculus carnosus as the wound bed. A full- were used as primary antibodies followed by the
thickness skin graft of identical size was harvested respective secondary antibodies and counter-
from the groin of the animal and placed into the stained with DAPI (Sigma-Aldrich).The 25-␮m
defect in the back of the chamber (Fig. 1, above). sections were fixed in acetone, dried, and washed
It was thus possible to view the microcirculation of in phosphate-buffered saline for 5 minutes, fol-
the wound bed from the front of the chamber and, lowed by overnight blocking with 10% donkey
at the same time, of the graft in the back of the serum in phosphate-buffered saline/Tween 0.1%.
chamber (Fig. 1, below). Rabbit anti-CD31 (1:50; Abcam, Cambridge,

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Volume 126, Number 1 • Angiogenic Transformation in Skin Grafts

Fig. 1. Images of the modified dorsal skinfold chamber. From the front of the chamber (above, left), the muscular wound bed
(panniculus carnosus) and the larger subcutaneous vessels are visible and accessible to intravital microscopy. Simultaneously,
the microcirculation of the skin graft can be visualized in vivo from the back (above, right). The microcirculation of the wound
bed showed a strong angiogenic response after 48 hours (below, left), leading to reperfusion of the graft capillaries after 72 hours
(below, right). Original magnification, ⫻200.

United Kingdom) and goat anti-desmin (1:50;

Santa Cruz Biotechnologies) were used as primary
antibodies, followed by incubation with the sec-
ondary antibodies and DAPI nuclear staining. The
4-␮m cryosections were evaluated by fluorescence
microscopy (Zeiss Axioscop 40; Carl Zeiss,
Oberkochen, Germany). In addition, confocal mi-
croscopy of the 25-␮m cryosections was performed
(SP5 Confocal Microscope; Leica Microsystems).

Corrosion Casting and Evaluation by Light and

Scanning Electron Microscopy
Standard methods were used for vascular cor-
rosion casting. Mice were anesthetized with pen-
tobarbital sodium (100 mg/kg body weight ad-
ministered intraperitoneally). The chest was
opened by means of a midline sternotomy and the
Fig. 2. Intravital microscopy was performed in eight different left ventricular chamber was entered using a 23-
areas within the center and periphery of the skin graft. gauge intravenous “butterfly” cannula. The right

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 Plastic and Reconstructive Surgery • July 2010

atrium was incised to allow for exsanguination of Statistical Analysis

the animal, and the circulatory system was flushed Differences between values were assessed us-
with 20 ml of artificial cerebrospinal fluid. The ing one-way analysis of variance or Kruskal-Wallis
casting medium PU4ii was used as described tests followed by the appropriate post hoc com-
previously.14 Then, the tissue specimen was ex- parison tests (Holm-Sidak and Dunn test, respec-
planted after complete resin curing after 4 to 6 tively). All data were expressed as means ⫾ SEM
days at room temperature. Soft tissue of the skin- and overall statistical significance was set at p ⬍
fold chamber was macerated in 7.5% potassium 0.05. Statistics and graphics were performed using
hydroxide at 50°C overnight. the software packages SigmaStat and SigmaPlot
For light microscopy, untreated PU4ii casts (Jandel Corp., San Rafael, Calif.).
were evaluated (Leica MZ 16A; Leica Microsys-
tems). Lyophilized samples consisting of the cast- RESULTS
ing material were mounted on metal stubs using
double-sided gluing tape and colloidal silver. Sam- Intravital Microscopy of the Skin Graft
ples were sputter coated with gold at 1.5 kV for 3 Functional capillary density, capillary diame-
minutes and examined using a scanning electron ter, red blood cell velocity, and the changes in
microscope at 15⫻ to 1000⫻ magnification (SU- capillary morphology were assessed. Baseline mi-
PRA 50 VP Scanning electron microscope; Zeiss). crocirculatory parameters of the groin skin did not

Fig. 3. Skin graft microcirculation in the graft center and periphery over a period of 10 days. Functional capillary density (FCD) (above,
left), capillary diameter (CD) (below, left), capillary blood flow velocity (RBV) (above, right), and density of the newly discovered “spherical
structures” (below, right) are shown. Mean ⫾ SEM; *p ⬍ 0.05 versus day 0; #p ⬍ 0.05 versus periphery.

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Volume 126, Number 1 • Angiogenic Transformation in Skin Grafts

reveal significant differences between center and

periphery for any of the parameters (Fig. 3). After
transplantation, the graft did not show any signs of
perfusion for 48 hours. After 3 days, slow blood
flow appeared within the graft vasculature, dem-
onstrating a pattern that was comparable to that of
the original microvasculature of the groin skin.
Most likely, this indicated the beginning of the
revascularization of the preexisting graft capillar-
ies, resulting in a functional capillary density of
38.1 ⫾ 7.7 cm/cm2 in the center of the graft and
22.2 ⫾ 4.6 cm/cm2 in the periphery (p ⬍ 0.05)
(Fig. 3, above, left). Functional capillary density
within the graft increased steadily, leading to al-
most identical values for the periphery and the
center following day 6. In line with this, red blood
cell velocity increased steadily after reperfusion
(Fig. 3, below, left). Capillary diameter of the graft
vessels increased significantly within the entire
graft after reperfusion, returning to normal values
at day 10 (Fig. 3, above, right). Most interestingly,
we observed the formation of spherical protru-
sions of the graft capillaries starting 24 to 48 hours
after reperfusion occurring predominantly at the
capillary divisions (Fig. 3, below, right).

Formation of Spherical Protrusions

The formation of spherical protrusions was
transient and disappeared after day 8, leading to
normal and slender capillary morphology at day
10. The process started in the center of the graft
at day 3 and progressed into the periphery. The
newly formed protrusions within the graft vascu-
lature were part of the capillaries and were scat-
tered at a maximum density of 6.2 ⫾ 2.1 per square
millimeter in the graft periphery at day 6 (Figs. 3,
below, right, and 4). They were up to seven times
wider than normal dermal capillaries and con-
tained clumped blood cells resulting from re-
duced microvascular flow.

Immunohistochemistry
Analysis of the expression of the angiogenesis
factors Ang-1, Flk-1, Flt-1, and bFGF within the Fig. 4. The spherical structures originated from the graft capillaries
vascular walls of the wound bed and the graft starting at day 3 after reperfusion and continued to grow larger in
exhibited an up-regulation of all molecules at day diameter (above). The structures appeared to contain cellular mate-
5, underscoring the angiogenic process that was rial, were up to seven times wider than normal capillaries, and pre-
seen using intravital microscopy (Fig. 5). sented slow but maintained blood flow (center and below).
Staining of the endothelium and pericytes
with CD31 and desmin in thick cryosections and
subsequent evaluation by confocal microscopy re- Light and Scanning Electron Microscopy
vealed that both molecules were present within Light microscopy showed large wound bed ves-
the vascular wall of the spherical structures within sels that could be identified because of their dark
the graft at day 5 (Fig. 6). blue color, whereas the smaller capillaries ap-

65
 Plastic and Reconstructive Surgery • July 2010

Fig. 5. Histologic evaluation of the expression of angiogenic factors within the wound bed and the skin graft revealed the
up-regulation of Ang-1 (above, left), the VEGF receptors Flt-1 (above, right ) and Flk-1 (below, left), and bFGF (below, right). DAPI
nuclear staining; original magnification, ⫻200. Goat anti–Ang-1, mouse anti–Flk-1, rabbit anti–Flt-1, and rabbit anti-bFGF were
used as primary antibodies followed by secondary staining with Texas red (above, left) and fluorescein isothiocyanate (above,
right, below, left, and below, right).

peared light blue and white. Larger magnifica- der the scanning electron microscope also stressed
tions showed the microvasculature of the skin the angiogenic transformation of the wound bed
graft itself, which could be identified because of its vessels, making reperfusion of the existing graft vas-
loop-like structures (Fig. 7, above). Vessels from culature as a consequence of the connection with
the wound bed and the typical capillaries of the newly formed bed vessels highly likely. At a greater
superficial and deep dermal plexus were inter- magnification, the connection from vessels originat-
connected. Interestingly, we also were able to ing from the wound bed to the original graft vascu-
identify the spherical structures at the capillary lature was seen (Fig. 7, below).
divisions of the graft vasculature with this method.
Evaluation by scanning electron microscopy DISCUSSION
made the two different natures of the vascular The vascularization of tissues is a well-known
beds even more clear: on the one hand, the wound aspect in plastic surgery.15–17 Tissue engineering as a
bed vessels with angiogenic transformation; on the therapeutic approach to regenerate lost or diseased
other hand, the graft vascular plexus in a more su- tissues by means of delivery of cells, constructs, and
perficial plane (Fig. 7, center). The graft intrinsic biomolecules is assumed to demand a functional
vascular dermal plexus appeared to be preserved as vascular network for adequate integration.3,18,19 This
well in this technique. The high magnification un- is the reason why the physiologic process of revas-

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Volume 126, Number 1 • Angiogenic Transformation in Skin Grafts

process has never been described. A simple “con-

tact” of two existing vessels in the wound bed and
skin graft would have to imply that these two ves-
sels lie in direct proximity to each other after
transplantation. Otherwise, this inosculation pro-
cess would have to involve some type of vessel
growth. According to earlier studies demonstrat-
ing dermal vessel growth at a rate of 5 ␮m/hour,
a 300-␮m skin graft may be pierced by newly
formed microvessels within 72 hours.21 In contrast
to this would stand the complete de novo vascu-
larization of the graft by vessels originating from
the wound bed, a process that most likely would
involve several weeks.22
In a previous study, it was suggested that the
autochthonous vessels of the graft provide merely
a conduit or sheath for the angiogenic vessels from
Fig. 6. Fluorescent immunohistochemistry and evaluation by the wound bed.23 In addition, Capla et al. found
confocal microscopy showed that the vascular wall of the spher- in an immunohistologic study that host vessels
ical structures expressed CD31 (green) and the pericyte marker invade the graft from the periphery along preex-
desmin (red), resulting in yellow overlay. Rabbit anti-CD31 and isting channels, replacing the graft vasculature.12
goat anti-desmin were used as primary antibodies followed by We propose the following interpretation for the
incubation with the secondary antibodies anti–rabbit-Cy2 and temporary angiogenic response. After reperfusion,
anti– goat-Cy3 and DAPI nuclear staining. Original magnifica- the graft is flushed with blood and, subsequently,
tion, ⫻630. angiogenic factors and nutrients reach the hypoxic
tissue. Because the angiogenic response does not
start until 24 to 48 hours after reperfusion, it can be
cularization of skin grafts and the possible inoscu- deduced that during the 3-day period of ischemia no
lation of a newly formed to an existing vascular net- angiogenesis takes place within the graft. This is
work is of great interest. most likely attributable to lack of energy as a con-
In the present study, we describe for first time sequence of hypoxia and also contradicts the theory
the transient formation of spherical protrusions in that the graft itself is able to perform angiogenesis
the graft capillaries between days 3 and 8 after before reperfusion.24,25 After reperfusion, the in-
grafting, which closely resembled angiogenic creasing oxygen supply from the wound bed
buds. Immunohistologic analyses revealed the ex- causes the still-functioning graft vascular network
pression of proangiogenic molecules within the to react. Angiogenesis starts in the center of the
wound bed and the graft vasculature. In addition, graft already at day 3, implying that the hypoxic
confocal microscopy showed that the wall of the stimulus may be most prominent in this area. It
structures expressed CD31 and desmin, suggesting then progresses to the periphery, where the max-
the presence of both endothelial cells and imum density of buds was found on day 6. It is
pericytes.20 Therefore, the formation of these spher- reasonable to argue that the hypoxic stimulus and
ical protrusions most likely represents an angiogenic subsequently the continuous production of proan-
response of the autochthonous graft capillaries. Pre- giogenic molecules are terminated after successful
sumably, this was preceded by a docking of sprouting revascularization, leading to a pruning of unnec-
vessels from the wound bed and subsequently a con- essary vascular buds until day 8. Another theory
nection to the preexisting graft vessels. These find- may be that the graft vasculature itself degenerates
ings contradict the generally accepted delay of sev- and is subsequently replaced by ingrowing host
eral weeks of neovascularization7 and also the vasculature. These results also support the theory
traditional assumption of an early direct inoscula- that revascularization of tissues with a preexisting
tion between adjacent capillaries within the wound microvascular network such as the skin is predom-
bed and skin graft.8,9 inantly a process driven by the respective recipient
Previous studies assumed a simple initial in- bed rather than the graft itself.
osculation process between existing vessels from In addition, by applying vascular corrosion
the wound bed with existing capillaries in the skin casting, a dense network of angiogenic vessels
graft after 48 hours.13 However, the nature of this from the wound bed connecting to the existing

67
 Plastic and Reconstructive Surgery • July 2010

dermal plexus of the skin graft was detectable. How-

ever, one major limitation of the technique of cor-
rosion casting is the lack of visibility of the vascular
wall itself. The exact evaluation of the molecular
changes at the interconnection sites is of great rel-
evance and the subject of ongoing studies.
After angiogenic stimulation, endothelial
cells begin to degrade the vascular basement
membrane.26 Subsequently, endothelial “tip” cells
migrate into the perivascular space, leading to bud
and sprout formation.27 The coverage of the de-
veloping tubes of endothelial cells by pericytes
signals the maturation of the newly formed vas-
cular structures.28 Based on the data obtained by
confocal microscopy, the spherical structures in
the presented study contain both endothelial cells
and pericytes. Pericytes and vascular smooth mus-
cle cells cover the endothelial cells in the vascu-
lature and have been reported to take part in the
process of angiogenesis.29,30 Historically, the re-
cruitment of pericytes into the angiogenic sprout
was thought to take place after the capillary plexus
had formed.31 However, later studies in fact sug-
gest that pericytes lead capillary sprouts.32–35 In
addition, pericytes have been described to be a
major source of VEGF production, which in the
early phase of angiogenesis is caused by a in-
creased nitric oxide production.36 To our knowl-
edge, this temporary angiogenic response in skin
grafts has never been described before. This is
most likely because no appropriate intravital mi-
croscopic model existed until now.
Angiogenesis and the contributing factors
have been studied extensively in the past.37,38 How-
ever, in revascularization of tissues with preexist-
ing vascular networks, ingrowing vessels somehow
need to “lyse” the existing capillary to connect to
it. New cellular junctions have to be established, a
process that has largely been unknown until today
and is ideally represented in the mechanism of
skin graft revascularization. One possibility may be
that the outgrowing angiogenic vessels still express
proteases similar to those they expressed during
initiation of the angiogenetic process. Several pro-
teases, such as chymase, plasminogen activator,
Fig. 7. Corrosion casting and subsequent light microscopy re-
tryptase, and metalloproteinases, have been sus-
vealed a dense three-dimensional microvascular network of
pected to play a role in this process.37 However,
newly formed vessels within the wound bed after skin graft-
their role in the connection of angiogenic vessels
ing. At a higher magnification, the formation of spherical pro- from the wound bed to the existing vascular net-
trusions of the graft capillaries after reperfusion was visible. work of the skin graft has never been investigated.
Vessels of the wound bed (asterisks) and the loop-like oriented
vessels of the graft (arrows) can be identified (above). Scan-
ning electron microscopy confirmed angiogenesis within the connections between newly developed tortuous vessels from
wound bed and reperfusion according to the original pattern the wound bed to the graft capillaries can be identified (below).
of graft vasculature (red) (center). At high magnifications, Original magnification, ⫻200 to ⫻1000.

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Volume 126, Number 1 • Angiogenic Transformation in Skin Grafts

The clarification of these mechanisms to apply ization of skin grafts in vivo: The role of angiogenesis. Plast
them onto the more complex process of the re- Reconstr Surg. 2008;122:1669–1680.
7. Goretsky MJ, Breeden M, Pisarski G, Harriger MD, Boyce ST,
vascularization of skin substitutes or preengi- Greenhalgh DG. Capillary morphogenesis during healing of
neered biomaterials should be addressed in fur- full-thickness skin grafts: An ultrastructural study. Wound
ther studies using modern in vivo models. Repair Regen. 1995;3:213–220.
8. Okada T. Revascularization of free full thickness skin grafts
in rabbits: A scanning electron microscope study of micro-
CONCLUSIONS vascular casts. Br J Plast Surg. 1986;39:183–189.
Applying a newly developed in vivo model to 9. Young DM, Greulich KM, Weier HG. Species-specific in situ
visualize skin graft revascularization, we were able hybridization with fluorochrome-labeled DNA probes to
to show for the first time a temporary angiogenic study vascularization of human skin grafts on athymic mice.
J Burn Care Rehabil. 1996;17:305–310.
response within the capillaries of the skin graft 10. Laschke MW, Vollmar B, Menger MD. Inosculation: Con-
between days 3 and 8 after transplantation starting necting the life-sustaining pipelines. Tissue Eng Part B Rev.
in the center of the graft. Immunohistochemistry 2009;15:455–465.
revealed the existence of endothelial cells and 11. Henry LD, Marshall DC, Friedman EA, Goldstein DP, Dam-
pericytes within the temporary angiogenic buds. min GJ. A histologic study of the human skin autograft. Am
J Pathol. 1961;39:317–332.
This most likely represents a reaction to reperfu-
12. Capla JM, Ceradini DJ, Tepper OM, et al. Skin graft vascu-
sion, implying the supply of angiogenic factors to larization involves precisely regulated regression and re-
the hypoxic skin graft. In addition, corrosion cast- placement of endothelial cells through both angiogenesis
ing supported the theory of angiogenesis within and vasculogenesis. Plast Reconstr Surg. 2006;117:836–844.
the wound bed and subsequent connection to the 13. O’Ceallaigh S, Herrick SE, Bluff JE, McGrouther DA, Fer-
existing vascular network within the skin graft. guson MW. Quantification of total and perfused blood ves-
sels in murine skin autografts using a fluorescent double-
Further studies to investigate the exact mecha- labeling technique. Plast Reconstr Surg. 2006;117:140–151.
nism of vessel connection and the development of 14. Krucker T, Lang A, Meyer EP. New polyurethane-based mate-
the graft vasculature are in progress. rial for vascular corrosion casting with improved physical and
imaging characteristics. Microsc Res Tech. 2006;69:138–147.
Nicole Lindenblatt, M.D. 15. Garrè C. Über die histologischen Vorgänge bei der Anhei-
Division of Plastic and Reconstructive Surgery lung der Thierschen Transplantationen. Beitr Klin Chir. 1888;
University Hospital Zurich 4:395.
Ramistraße 100 16. Hübscher C. Beiträge Hautverpflanzung nach Thiersch. Zen-
8091 Zurich, Switzerland tralbl All Pathol. 1890;505:1.
[email protected] 17. Goldmann EE. Die künstliche Überhäutung offener Krebse
durch Hauttransplantationen nach Thiersch. Beitr Klein Chir.
ACKNOWLEDGMENTS 1890;4:395.
18. Shepherd BR, Enis DR, Wang F, Suarez Y, Pober JS, Schech-
The authors thank Kerstin Abshagen and Dorothea ner JS. Vascularization and engraftment of a human skin
Frenz, Institute for Experimental Surgery, University of substitute using circulating progenitor cell-derived endothe-
Rostock, Rostock, Germany; Eric P. Meyer, Institute of lial cells. FASEB J. 2006;20:1739–1741.
Zoology; Martha Bain, Department of Visceral and 19. Tremblay PL, Hudon V, Berthod F, Germain L, Auger FA.
Transplantation Surgery; and Klaus Marquardt, Cen- Inosculation of tissue engineered capillaries with the host’s
vasculature in a reconstructed skin transplanted on mice.
ter for Microscopy and Image Analysis (all from the
Am J Transplant. 2005;5:1002–1010.
University of Zürich, Zürich, Switzerland) for their ex- 20. Gerhardt H, Betsholtz C. Endothelial-pericyte interactions in
cellent technical assistance. angiogenesis. Cell Tissue Res. 2003;314:15–23.
21. Zarem HA. Development of the microcirculation in full
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