Archivio Italiano di Urologia e Andrologia 2013; 85, 128

INTRODUCTION
The Peyronie's disease (PD) is an idiopathic disorder of
connective tissue of the penis, that involves the tunica
albuginea of the corpora cavernosa and the adjacent are-
olar space. A fibrous plaque replaces the elastic fibers
that normally constitute the tunica albuginea, limiting its
extensibility, and causing pain during erection, erectile
dysfunction and difficulty of penetration, as a result of
the deviation of the penis.
Although it was once considered a relatively infrequent
disease, some studies show that the prevalence of IPP is

ORIGINAL PAPER

Animal experimental model of Peyronie’s disease: 
A pilot study 

Maria Angela Cerruto 1, Carolina D’Elia 1, 2, Alberto Molinari 1, 
Francesca Maria Cavicchioli 1, Antonio D’Amico 1, Walter Artibani 1

1 Urology Clinic, A.O.U.I. Verona, Verona, Italy;
2 Urology Clinic, Santa Chiara Hospital, Trento, Italy.

Objective: The Peyronie's disease (PD) is an idiopathic disorder of connective tissue of
the penis, that involves the tunica albuginea of the corpora cavernosa and the adjacent
areolar space. It is a growing clinical evidence to support the therapeutic potential of
mesenchymal stem cells and histological findings has assumed a possible application of
lipofilling technique in patients with PD. The objective of this experimental study is the

creation of a murine experimental model of PD, evaluating with MRI the penis of the rats (feasi-
bility study), in order to plane the application of lipofilling technique in an animal model.
Methods: Four male Wistar rats were anesthetized, fixed in prone position and subjected to MRI.
The animals underwent, subsequently, an injection of thrombin in the tunica albuginea and MRI
images were acquired at 7 and 21 days after injection with incision of the dartos.
Results: The MRI acquisitions, both in coronal and axial projection, showed an adequate visibili-
ty of the anatomical structures. At 7 days after thrombin injection with the dartos incision it was
evident an oedematous portion, visible as a hyperintense area, located at the injection area. At 21
days after injection, oedema was partially resolved: the injection part of the hyperintense area
remains unchanged, while the remaining area appears to be part of a re-absorption and re-organ-
ization process.
Conclusions: Since none of the various treatment modalities currently available for the manage-
ment of PD is able to bring healing, the researchers’ attention is increasingly directed towards
innovative treatment programs, such as the use of stem cells of mesenchymal origin. At the pres-
ent time, the research in PD is hampered by the lack of universally accepted animal model and
this is likely attributed to the limited insight into PD mechanisms and the difficulties faced by cur-
rent animal models to truly represent the complexity.

KEY WORDS: Peyronie’s disease; Murine animal model.

Submitted 14 January 2013; Accepted 31 January 2013 No conflict of interest declared

Summary

comparable to that of diabetes or of urolithiasis (1, 2). A
recent epidemiological study reported a prevalence of
3.2% (3), highlighting the potential psychosocial impact
of this disease.
The Peyronie's disease affects predominantly adulthood
(approximately 60% of cases between 40 and 60 years)
and is still present at young age (10-20% at the age of 40)
and advanced age (15-20% over 60 years of age) (4).
Further studies have tried to identify risk factors associ-
ated with PD: smoking, hypertension, hyperlipidemia

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29Archivio Italiano di Urologia e Andrologia 2013; 85, 1

Animal experimental model of Peyronie’s disease: A pilot study 

ischaemia, hyperpermeability, fibrosis and increased
ischemia itself. This vicious circle can be broken using
the proangiogenetic capacity of mesenchymal stem cells. 
It is a growing clinical evidence to support the therapeu-
tic potential of mesenchymal stem cells for the revascu-
larization of ischemic tissues and the recovery of their
function.
In a recent italian study Rigotti et al. (20) have tested the
effectiveness of purified lipoaspirates on 20 patients
operated on for breast cancer with severe side effects and
irreversible functional damage from postoperative radio-
therapy. 
60 to 80 cc of adipose tissue were collected from the
medial region of the knee, from the abdominal region or
trochanteric: subsequently, the lipoaspirates was cen-
trifuged at 2700 rpm for 15 minutes, in order to remove
most of the deposits of triglycerides and to cause injury
of membrane in adipocytes, in order to allow the rapid
clearance once injected. In this way, the study authors
have created a sort of concentration of adult stem cells,
which, maintained their three-dimensional microenvi-
ronment, appear to be more effective in the neoangio-
genetic process (21).
In the same paper the ultrastructure of the tissue dam-
aged by radiotherapy has been studied before injection
of lipoaspirates and after 1, 2, 6 and 12 months. 
Prior to injection, the tissue showed fibrotic patterns
similar to those of scleroderma: duplication in the base-
ment membrane of the capillaries, spaces between the
endothelial cells, adipocytes containing lysosomes or
clusters of mitochondria, accumulation of collagen.
The after-treatment ultrastructural analysis showed signs
of neoangiogenetic, neoadipogenetic and antifibrotic
activity, due to the activity of adult stem cells contained
in lipoaspirates (22-24).
At a clinical level, this activity corresponds to a dramat-
ic improvement of symptoms, disappearance of necrosis
and wound healing.
Based on these innovative clinical and histological find-
ings has assumed a possible application of lipofilling
technique in patients with PD. 
It was therefore drawn up an experimental protocol con-
sists of two basic steps:
1. creating an animal model of PD on the basis of exper-

iments already conducted (16, 25) verifying with MRI
(Magnetic Resonance Imaging) stages of development
of the disease;

2. the application of lipofilling technique in an animal
model.

The objective of this experimental study is the creation
of a murine experimental model of IPP (16), evaluated
with MRI (feasibility study).

MATERIALS AND METHODS
Four male Wistar rats weighting between 420 and 450 g.
were fed with standard rodent diet (Rieper SpA, Italy) and
maintained under conditions of controlled temperature
and light, respecting the principles of the "NIH Guide for
the Care and Use of Laboratory Animals" and the directives
of the European Community.

have all been taken into account, but basically consid-
ered to be associated with erectile dysfunction more than
strictly to PD. 
Microscopically, the tunica albuginea is largely com-
posed by fibrillar collagen type I, woven with elastin
fibers, in order to form organized structures (5). Even
the plaques of PD are mostly constituted from collagen
type I and III (6), but the elastin component, which gives
to the tunica albuginea elongation properties up to
150%, is altered: the decrease elastin, as well the alter-
ations in the arrangement of the tunic layers, are charac-
teristics of PD, that can lead to deviations in penile erec-
tion and erectile dysfunction (7).
More recent papers have identified a strong association of
PD with both Dupuytren's disease and the HLA B-27 (8).
Other studies have taken into account immunological fac-
tors, showing that patients with PD in 75.8% of cases pre-
sented an abnormal immunological test, 48.5% had   alte -
rations in cell-mediated immunity and the 37.9% pre-
sented increase in markers for autoimmune diseases (9)
but it seems that this predisposition, rather than systemic,
is localized to the tunica albuginea. Another study has
documented evidence of high anti-elastin antibodies in
the serum of patients with IPP, hypothesizing, the same
way, an autoimmune etiology (10).
The role of free radicals in the pathogenesis of plaque
seems to be important; Bivalacqua et al. (11) have shown,
on the basis of other studies (12-14), that the production
of Nitric Oxyd (NO) can inhibit the deposition of colla-
gen in the murine model of PD.
The most widely accepted theory considers the penile
trauma as main causative factor in the pathogenesis of
PD: an acute traumatic event or a repetitive microtrauma
(e.g. during sexual activity) can result in delamination of
the tunica albuginea and microvascular damage (8, 15).
Fibrin was found to be potentially involved in the devel-
opment of the fibrotic plaque in a murine model of PD
(16). In the same study, and in another more recent
(17), was also reported the presence of increased levels
of PAI-1, an important inhibitor of fibrinolysis and col-
lagenolysis: its increased activation appears to play a
key role in the development of penile plaque.
The PD has been associated with many other conditions
in urologic procedures such as radical retropubic prosta-
tectomy, cystoscopy and introduction of indwelling ure-
thral catheters, genital or peritoneal trauma, urethritis,
lipomas (18). 
Even atherosclerosis has been mentioned as a target area
on the study of PD, due to the similarities about the
pathophysiology (19), regarding the common mecha-
nism of cell hyperproliferation, that leads to the forma-
tion of a plaque.
Since none of the various treatment modalities currently
available for the management of PD is able to bring hea -
ling to the patients who are affected, the researchers'
attention is increasingly directed towards innovative
treatment programs based on a better understanding of
the pathophysiological disease.
Moreover, the overexpression of cytokines and autoim-
mune and genetic factors have been cited as cofactors (8). 
The resulting inflammatory process would be responsi-
ble for the onset of a vicious circle of vascular injury,

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30

Before being subjected to MRI, the rats were anesthetized
in an induction chamber by inhalation of a mixture
70/30% of nitrous oxide and oxygen containing 5%
isoflurane. During the execution of the examination, anes-
thesia was maintained via buccal mask with the same
mixture of nitrous oxide and oxygen with isoflurane 3%.
Image acquisition was performed using a scanner
Biospec (Bruker, Karlsruhe, Germany) equipped with a
horizontal magnet operating at 4.7 Tesla with opening of
33 cm (Oxford Ltd, Oxford, UK) (Figure 1). The rats were
fixed in a prone position on a birdcage type coil diame-
ter of 7.2 cm (Figure 2). 
In order to verify the correct positioning of the animal
with respect to the coil, T1-weighted coronal projection
acquisitions were performed, while, in order to obtain
pre-and post-injection anatomical details, acquisitions
T2-weighted images in axial view were performed.
The coronal images were acquired with a relaxation time
(TR) of 486 ms, echo time (TE) of 11 ms, with the use
of 2 medium for 20 slices, each of thickness 1 mm. The
field of view (FOV) was 6 x 6 cm and the matrix choice
of 256 x 256, corresponding to a spatial resolution of
0234 mm2/pixel.
Axial images were acquired with a relaxation time (TR)
of 5000 ms, echo time (TE) of 67 ms, with the use of 2
medium for 25 slices, each of thickness 1 mm. The field
of view (FOV) was 6 cm x 3 matrix and the choice of
512 x 256, corresponding to a spatial resolution of
0.117 mm2/pixel.
The animals underwent, subsequently, an injection of 60
mL of thrombin in the tunica albuginea, using a 1 mL
syringe with 30 G needle. In the first measure, the throm-
bin was injected without opening the dartos (Figure 3):
thereafter, considering the substantial ineffectiveness of
the first surgery, an incision was performed along the dar-
tos of about 2 mm along the penile axis, in order to have
a more secure access to the subdartoic spaces, and the
thrombin was injected with the same method (Figure 4).
MRI images were acquired in the same manner as
described above at 7 and 21 days after injection with
incision of the dartos.

Figure 1.

Figure 2.

Figure 3.

Figure 4.

RESULTS
The acquisitions, both in projection coronal both in
axial, have demonstrated adequate visibility of anatomi-
cal structures.
Figure 5 shows a T1-weighted image, a coronal projec-
tion of the penis, while Figure 6, T2-weighted axial view,
shows the results produced by the thrombin injection,
offering a good view of the albuginea and its relationship
with the superficial and deeper structures.
At 7 days after thrombin injection with incision of the
dartos it was evident a edematous portion (Figure 7)
located at the injection area. Oedema, visible as a hyper-
intense area, appeared to affect the section previously
seen as a hypointense rim surrounding the corpora cav-
ernosa of the penis, leaving them intact and with a signal
intensity comparable to that seen in the image acquired
before the injection.
At 21 days after injection, edema was partially resolved
(Figure 8): in the injection part of the hyperintense area
remains unchanged, while the remaining area appears to
be part of a reabsorption and reorganization process.

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Animal experimental model of Peyronie’s disease: A pilot study 

DISCUSSION
The low success rate obtained by standard therapies has
fueled our interest in the development of new treatment
options, such as the use of stem cells of mesenchymal ori-
gin. In literature has recently emerged a growing increase
of the clinical evidence to support the therapeutic poten-
tial of mesenchymal stem cells for the revascularization of
ischemic tissues and the recovery of their function (20). 
Significant clinical results were obtained from autolo-
gous endothelial and hematopoietic stem cells from the
bone marrow in ischemic lesions at the level of the limbs,
myocardium and of the retina in vitro and in animal
models, thereby elucidating the mechanisms of stem cell
– mediated neo-angiogenesis.
The use of stem cells could be useful for the management

Figure 5. Figure 6.

Figure 7. Figure 8.

of fibrotic connective tissue diseases such as PD, breaking
a vicious circle of self-perpetuating inflammatory condition
with post-traumatic vascular injury, subsequent ischemia,
hyperpermeability, increased fibrosis and ischemia itself. 
Since there is no data in the literature concerning the
role of stem cells in management of PD, we planned to
develop a murine model of stable IPP, monitored with
imaging (MRI), and then try to corroborate the hypoth-
esis that mesenchymal stem cells can have a positive
influence on the natural history of this disease. 
In order to create a murin model of PD we injected 60 μl
of thrombin at the level of the tunica albuginea obtain-
ing after 3 weeks after injection the persistence of the
inflammatory reaction at the site of inoculation, identifi-

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32

able to control MRI, without secure evidence of fibrosis.
Moreover, a review of the literature on human MRI of the
penis was carried out, in order to assess the feasibility of
the application of this method in animals: although ultra-
sound is the primary mode of imaging penis, the MRI may
use a higher spatial resolution and improved contrast soft
tissue. MRI is to be considered as an opportunity for
progress in penile imaging and is able to solve clinical
questions for which the ultrasound is not conclusive.
In addition to PD, other diseases can be investigated by
this method, such as primitive or secondary penile neo-
plasms, periurethral abscess, venous thrombosis, frac-
tures; can also be rated the positioning of penile pros-
theses after surgery. 
The review of the literature has concerned in particular
the possibility to discriminate with the MRI the tunica
albuginea compared to the remaining structures in
humans, so as to apply sequences of acquisition and
anatomical similarities also in rats.
The T2-weighted images provide more detailed anatom-
ical details: in particular, the tunica albuginea is visible as
a hypointense area that surrounds the corpora cavernosa
and which also includes Buck's fascia. 
In a work of Pretorius et al. (26) also the PD was studied
by magnetic resonance imaging: there was evidence of a
thickening of the tunica albuginea, visible in both T2-
weighted both in those T1- weighted images.
Davila et al. (16) have recently proposed an animal
model of IPP injecting at the level of the tunica albuginea
of rat 30 μ l of fibrin and thrombin, inducing edema after
one week after inoculation and fibrosis after three weeks. 
The use of these components would trigger the final
steps of the coagulation cascade and reproduces experi-
mentally what happens after a trauma at the level of the
tunica albuginea. The profibrotic effect is most likely due
to the fibrin and thrombin deposition in a fibroblasts rich
and hypovascularized tissue. 
The study of Davila et al. also showed that, compared to
previous methods that used injection of TGF-!1 (27),
the fibrin induced plaque develops more quickly (three
weeks instead of six weeks). The TGF-!1, in fact, has an
anti-inflammatory action in reducing macrophage activa-
tion and inhibition of cytotoxic T cells (28, 29): this may
explain the study of El-Sakka et al. (27) the non-forma-
tion of plaque in three weeks and the discovery of the
same six weeks, the latter a result of the action of TGF-
!1 profibrotic same.
In our study, after this period there has been found the
persistence of the inflammatory condition (edema),
probably due to the lower speed of action of thrombin
compared to fibrin alone. It is important to observe, that
the literature data are only based on histological founds,
not also on imaging, as in our study. 
While the exact mechanism of Peyronie's disease (PD)
remains an enigma, the pathophysiology of PD is con-
sidered to be multifactorial, with interactions of genetic
predisposition, trauma, tissue inflammation and aberrant
wound healing. 
The most frequently reported models of PD can be clas-
sified as TGF !1, fibrin and surgical trauma-induced
models. In vitro studies using Peyronie's fibroblast cul-
ture media have also provided further insights into cel-

lular mechanism of PD. At the present time, the research
in PD is hampered by the lack of universally accepted
animal model and this is likely attributed to the limited
insight into PD mechanisms and the difficulties faced by
current animal models to truly represent the complexity
and complete spectrum of human disease.

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Correspondence

Maria Angela Cerruto, MD, FEBU
mariaangela.cerruto@univr.it

Carolina D’Elia, MD, FEBU (Corresponding Author)
karolinedelia@gmail.com

Alberto Molinari, MD
yefet83@yahoo.it

Francesca Maria Cavicchioli, MD
francesca.cavicchioli@gmail.com

Antonio D’Amico, MD
antonio.damico@ospedaleuniverona.it

Prof. Walter Artibani, MD
walter.artibani@univr.it
Urology Clinic, A.O.U.I. Verona 
P.zza L.A. Scuro 10 - 37134 Verona, Italy

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