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Stem Cell Therapy for Erectile Dysfunction: A Critical Review

Authors:
  • City University of Macao

Abstract and Figures

Erectile dysfunction (ED) is a prevailing health problem that seriously impacts quality of life. Current treatment options are less effective for patients having cavernous nerve (CN) injury or diabetes mellitus-related ED. These 2 types of ED are thus the main focus of past and current stem cell (SC) therapy studies. In a total of 16 studies so far, rats were exclusively used as disease models and SCs were mostly derived from bone marrow, adipose tissue, or skeletal muscle. For tracking, SCs were labeled with LacZ, green fluorescent protein, 4',6-diamidino-2-phenylindole, DiI, bromodeoxyuridine, or 5-ethynyl-2-deoxyuridine, some of which might have led to data misinterpretation. SC transplantation was done exclusively by intracavernous (IC) injection, which has been recently shown to have systemic effects. Functional assessment was done exclusively by measuring increases of IC pressure during electrostimulation of CN. Histological assessment usually focused on endothelial, smooth muscle, and CN contents in the penis. In general, favorable outcomes have been obtained in all trials so far, although whether SCs had differentiated into specific cell lineages remains controversial. Recent studies have shown that intracavernously injected SCs rapidly escaped the penis and homed into bone marrow. This could perhaps explain why intracavernously injected SCs had systemic antidiabetic effects and prolonged anti-ED effects. These hypotheses and the differentiation-versus-paracrine debate require further investigation.
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CONCISE REVIEW
Stem Cell Therapy for Erectile Dysfunction:
A Critical Review
Ching-Shwun Lin,
1
Zhong-Cheng Xin,
2
Zhong Wang,
3
Chunhua Deng,
4
Yun-Ching Huang,
5
Guiting Lin,
1
and Tom F. Lue
1
Erectile dysfunction (ED) is a prevailing health problem that seriously impacts quality of life. Current treatment
options are less effective for patients having cavernous nerve (CN) injury or diabetes mellitus-related ED. These
2 types of ED are thus the main focus of past and current stem cell (SC) therapy studies. In a total of 16 studies so
far, rats were exclusively used as disease models and SCs were mostly derived from bone marrow, adipose
tissue, or skeletal muscle. For tracking, SCs were labeled with LacZ, green fluorescent protein, 4’,6-diamidino-2-
phenylindole, DiI, bromodeoxyuridine, or 5-ethynyl-2-deoxyuridine, some of which might have led to data
misinterpretation. SC transplantation was done exclusively by intracavernous (IC) injection, which has been
recently shown to have systemic effects. Functional assessment was done exclusively by measuring increases of
IC pressure during electrostimulation of CN. Histological assessment usually focused on endothelial, smooth
muscle, and CN contents in the penis. In general, favorable outcomes have been obtained in all trials so far,
although whether SCs had differentiated into specific cell lineages remains controversial. Recent studies have
shown that intracavernously injected SCs rapidly escaped the penis and homed into bone marrow. This could
perhaps explain why intracavernously injected SCs had systemic antidiabetic effects and prolonged anti-ED
effects. These hypotheses and the differentiation-versus-paracrine debate require further investigation.
Introduction
Erectile dysfunction (ED) is a prevailing health problem
that seriously impacts the quality of life of men and their
spouses or partners [1]. In the United States alone, an esti-
mated 30 million men suffer from different degrees of ED [2].
The majority of ED patients can now be treated satisfactorily
with phosphodiesterase type-5 (PDE5) inhibitors, such as
sildenafil (Viagra; Pfizer Inc., New York, NY), vardenafil
(Levitra; Bayer AG, Leverkusen, Germany), and tadalafil
(Cialis; Lily-ICOS, Indianapolis, IN) [3]. However, PDE5 in-
hibitors are strictly contraindicated with concomitant nitrates
because of the danger of their synergistic hypotensive effects
[3]. PDE5 inhibitors are known to cause a variety of adverse
side effects that may reduce their suitability for some patients
[3]. More importantly, PDE5 inhibitors are only partially ef-
fective in treating certain types of ED including those associ-
ated with diabetes mellitus (DM) and surgery-induced CN
injuries (mainly due to radical prostatectomy) [3]. As such,
alternative treatments, particularly those that can treat the
underlying disease process of ED, are highly desirable. In this
regard, one of the strategies currently being evaluated is stem
cell (SC) therapy.
Rationale for Using SC Therapy
Despite being conventionally classified in the urinary
system, the penis is in fact a vascular organ. The penile
corpora cavernosa are composed of sinusoids that are lined
with a single layer of endothelial cells (ECs) and are sur-
rounded by multiple layers of circular and longitudinal
cavernous smooth muscle cells (CSMCs) (Fig. 1). In the
flaccid penis, CSMCs are in a contracted state and maintain a
small amount of blood flow in and out of the sinusoids.
When a man is sexually aroused, nitric oxide (NO) is re-
leased from terminal fibers of cavernous nerves (CNs) and
enters the neighboring CSMCs, resulting in CSMC relaxa-
tion. Blood rushes in as a consequence and engorges the
sinusoids, leading to the initial phase of penile erection.
Maintenance of erection, that is, continued CSMC relaxation,
is believed to derive from additional NO release from the
sinusoidal ECs. The further sinusoidal engorgement causes
1
Knuppe Molecular Urology Laboratory, Department of Urology, School of Medicine, University of California, San Francisco, California.
2
Andrology Center, Peking University First Hospital, Beijing, China.
3
Department of Urology, Ninth People’s Hospital Affiliated to Medical College of Shanghai Jiao-Tong University, Shanghai, China.
4
Department of Urology, First Affiliated Hospital of Sun Yet-Sen University, Guangzhou, China.
5
Division of Urology, Department of Surgery, Chang Gung Memorial Hospital, Chia-Yi, Taiwan, China.
STEM CELLS AND DEVELOPMENT
Volume 21, Number 3, 2012
Mary Ann Liebert, Inc.
DOI: 10.1089/scd.2011.0303
343
the compression of venules located between the trabeculae
and the tunica albuginea, resulting in nearly total occlusion
of venous outflow. This combination of sinusoidal engorge-
ment and venous occlusion leads to the full erection of the
penis [4]. Thus, structurally, the key components of erection
are the ECs, CSMCs, and CN [more specifically, neuronal
nitric oxide synthase (nNOS)-positive nerves], and func-
tionally, accurate interactions among these 3 constituents are
critical. In different types of ED, these structures and/or in-
teractions are often altered, as described briefly in the fol-
lowing text.
The purpose of a radical prostatectomy is to remove the
cancerous prostate; however, the procedure can damage the
CN, which run alongside the prostate [5]. The short-term
consequence of CN injury is neurogenic ED, which is re-
versible, whereas the long-term consequence is atrophy of
CSMCs, which can lead to irreversible ED [6]. In men with
DM, the high blood glucose also causes reduction of CN, EC,
and CSMC contents [7]. In men with hyperlipidemia, im-
paired EC function is well documented, but whether it is
accompanied with structural changes is not as certain [7]. In
our recent studies we have quantified structural changes in
the penises of rats with CN injury, type 2 DM (T2DM), and
hyperlipidemia [8–10]; these changes are summarized in Fig.
2. Note that hyperlipidemia was induced by high-fat feeding,
and T2DM by high-fat feeding plus streptozotocin injection.
High-fat feeding causes increased CSM content in the rat
[10], and this may have compensated the CSM loss that is
usually associated with DM, resulting in the preservation of
CSM in the T2DM rats [8].
SCs are believed to be able to differentiate into various cell
types including ECs, smooth muscle cells (SMCs), Schwann
cells, and neurons [11]. Therefore, SC therapy for ED was
originally based on the hypothesis that transplantation of
SCs into penis through intracavernous (IC) injection might
replenish the depleted EC and/or CSMC pools. However, in
published studies whether cellular differentiation occurred
in the treated animals is controversial and will be discussed
in more details in a later section under Histological assessment.
On the other hand, there were also considerations that IC
transplanted SCs might encourage the regeneration of the
host’s own ECs and CSMCs or might restore proper inter-
actions between ECs and SMCs. In other words, paracrine
actions as opposed to cellular differentiation are responsible
for SCs’ therapeutic efficacy, and based on several of our
studies in both ED and non-ED fields, this seems to be the
main mechanism [12].
With regard to restoring damaged CN, it should be
pointed out that the nerve cell bodies are located in the major
pelvic ganglia (MPG) some distance away from the penis. As
such, in the first SC-for-ED study, SCs were injected into the
MPG of one group of CN injury rats, and the results showed
improved erectile function [13]. Importantly, in the same
study, SCs were also injected into the penis of another group
of CN injury rats, and similar results were obtained. This
finding is cited as basis for using IC injection in the next SCs
for CN injury study [14], and since then this injection method
has been apparently accepted as legitimate, although no one
knew how it worked until now (see Stem cell transplantation
section).
Current Status
There are currently 16 publications in the field of SC
therapy for ED (Table 1). Five additional publications tested
SC transplantation without assessing erectile function and
are thus not SC therapy per se.
Clinical Trial
A clinical trial of SC therapy for ED has been carried out in
Korea [25]. In this study, 7 T2DM men ranging from 57 to 87
years of age were each treated with IC injection of 15 million
allogeneic umbilical cord blood SCs. Morning erection was
regained in 3 patients within 1 month and in 6 patients
within 3 months. However, despite having increased penile
rigidity, none was able to achieve vaginal penetration unless
aided by taking sildenafil before coitus. During 11-month
follow-up, only one treated subject maintained erection suf-
ficient for coitus. Interestingly, SC therapy appears to have
antidiabetes effects as all treated subjects except the oldest
had reduced blood glucose and glycosylated hemoglobin
levels. These results provide further evidence not only for the
frequently observed antidiabetes effects of SCs but also for
IC injection being a systemic application as we recently re-
ported (see Stem cell transplantation section). Also noteworthy
is that, despite being allogeneic in the absence of immuno-
suppressant, SC transplantation did not cause any adverse
effects, thus providing further evidence for the immuno-
suppressive effects of SCs.
Preclinical Trials
A typical preclinical trial of SC therapy for ED is sche-
matically depicted in Fig. 3. It involves the isolation,
FIG. 1. Penile histology of a
normal rat. The cross-section
of rat penis was stained with
Alexa-488-conjugated phal-
loidin, which detects smooth
muscle (green stain), with
Alexa-594-conjugated anti-
RECA antibody, which de-
tects endothelium (red stain),
and with DAPI, which detects
cell nuclei (blue stain). DAPI,
4’,6-diamidino-2-phenylindole;
CC, corpus cavernosum; Pha,
phalloidin; RECA, rat en-
dothelial cell antigen.
344 LIN ET AL.
cultivation, sorting, and modification of SCs, followed by
labeling them with a cell-tracking agent. The labeled SCs
were then injected into the corpus cavernosum (IC injection)
of an ED animal model. Weeks or months later, the animals
are tested for erectile function, usually by measurement of
increases in intracavernous pressure (ICP) during electro-
stimulation of CN. The animals are then sacrificed for his-
tological assessment of corpus cavernosum and tracking of
injected SCs.
Animal models
Rat is the most commonly used animal in ED research and
was used in all preclinical SC-for-ED studies thus far. Un-
derscoring clinical needs, CN injury and DM are the most
commonly tested disease models. CN injury, tested in 7
studies, was induced by either crush or resection of CN bi-
laterally. T2DM patients outnumber T1DM patients 9 to 1,
yet most experimental studies chose T1DM over T2DM. This
FIG. 2. Histological changes
in the penises of ED rats. Re-
presentative images from
normal rats and indicated ED
models illustrate ED-associated
changes in smooth muscle (left
column), endothelium (middle
column), and nNOS-positive
nerves (right column). Quantifi-
cation of these changes in 9 rats
in each group of rats was done
as described in our previous
studies, and the results are
shown at the bottom.For
smooth muscle and endothe-
lium, the unit on the Y-axis is
number of pixels/high-power
field (HPF). For nNOS, the unit
is number/HPF, where ‘‘num-
ber’’ refers to the red dots seen in
the images. Note that all tissue
sections were costained with
DAPI for the visualization of
cell nuclei (blue). ED, erectile
dysfunction; nNOS, neuronal
nitric oxide synthase.
STEM CELL THERAPY FOR ERECTILE DYSFUNCTION 345
Table 1. Erectile Dysfunction-Related Stem Cell Transplantation Studies
Publication year/first
author
Animal
model/patients Cell type
Transplantation
method/cell
number
Cell tracking
method
Assessment
time point
ICP
assessment
Penile histological
assessment
2003/Deng [15] 25-month-old rats Rat eNOS-BMSC IC/500,000 None 1 week No IHC for eNOS
2004/Bochinski [13] CN injury rats Rat BDNF-ESC IC or Intra-MPG/
500,000
GFP label 3 months Yes NADPH, IHC for TH,
neurofilament
2006/Kim [14] CN resection rats Rat SkMDC IC/1,000,000 LacZ label 2 and 4 weeks Yes IHC for PGP9.5
2007/Bivalacqua [16] 25-month-old rats Rat eNOS-BMSC IC/500,000 LacZ label 1 and 2 weeks Yes IF for SMA, vWF,
eNOS, PECAM,
SM-MHC, CD45
2007/Song [17] 10-week-old rats Human v-myc-BMSC IC/1,000,000 IF for human
nucleus
2 weeks No IF for desmin, calponin,
SMA, vWF, CD31
2008/Song [18] 10-week-old rats Human v-myc-NCSC IC/1,000,000 IF for human
nucleus
2 weeks No IF for desmin, calponin,
SMA, vWF, CD31
2008/Nolazco [19] 20-month-old rats Mouse SkMSC IC/500,000–
1,000,000
DAPI label 2 and 4 weeks Yes IHC for CD34, Sca1
2009/Fall [20] CN resection rats Rat BMMN cells IC/10
7
PKH-26 label 3 and 5 weeks Yes IHC for SMA, CD31,
vimentin
2009/Song [21] 6-month-old rats Rat fetal brain stem cells IC/1,000,000 GFP label 2, 4, and 6 weeks No IF for SMA, calponin,
VEGF
2010/Garcia [22] Zucker T2DM rats Autologous ADSC IC/1,000,000 BrdU label 3 weeks Yes IHC for RECA, nNOS
2010/Huang [23] Hyper-lipidemic rats Autologous ADSC IC/2,000,000 EdU label 3, 14, 28 days for
histology.
28 days for ICP
Yes IF for RECA, nNOS,
SMA
2010/Abdel Aziz [24] 2 to 2.5-year-old rats Rat BMSC IC/1,000,000 GFP label 3 and 4 weeks; 3
and 4 months
Yes HE stain
2010/Bahk [25] T2DM patients Umbilical cord blood
stem cells
IC/15,000,000 None Up to 11 months No None
2010/Albersen [26] CN injury rats Autologous ADSC and
lysate
IC/1,000,000 EdU label 4 weeks Yes IF for nNOS, SMA,
b-III-tubulin
2010/Kendirci [27] CN injury rats P75-selected rat BMSC IC/500,000 GFP transgene 4 weeks Yes IF for GFP
2011/Qiu [28] STZ T1DM rats Rat BMSC IC/400,000 DiI label 4 weeks Yes IF for calponin, SMA,
vWF, CD31
2011/Qiu [29] STZ T1DM rats Rat VEGF-BMSC IC/500,000 DiI label 4 weeks Yes IF for SMA, CD31
2011/Gou [30] STZ T1DM rats VEGF-transfected EPC IC/2,000,000 DAPI 3 weeks Yes IHC for CD34
2011/Lin [31] CN resection rats ADSC-seeded adipose
matrix
Nerve grafts EdU label 3 months Yes None
2011/Lin [32] CN resection rats Rat ADSC IC/1,000,000 EdU label 2 and 7 days No None
2011/Fandel [33] CN resection rats Autologous ADSC IC/2,000,000 EdU label 1, 3, 7, and 28
days
Yes IF for nNOS, S100,
SDF-1. Trichrome
ICP, intracavernous pressure; eNOS, endothelial nitric oxide synthase; CN, cavernous nerve; BMSC, bone marrow stem cells; IC, intracavernous; IHC, immunohistochemistry; BDNF, brain-derived
neurotropic factor; ESC, embryonic stem cell; MPG, major pelvic ganglia; TH, tyrosine hydroxylase; SkMDC, skeletal muscle-derived cells; PGP, protein gene product; IF, immunofluorescence; vWF, von
Willebrand factor; PECAM, platelet endothelial cell adhesion molecule; SM-MHC, smooth muscle myosin heavy chain; DAPI, 4’,6-diamidino-2-phenylindole; BMMN, bone marrow mononuclear; T2DM,
type 2 diabetes mellitus; ADSC, adipose-derived stem cell; EdU, 5-ethynyl-2-deoxyuridine; BrdU, 5-bromo-2-deoxyuridine; VEGF, vascular endothelial growth factor; RECA, rat endothelial cell antigen;
nNOS, neuronal nitric oxide synthase; HE, hematoxylin and eosin; SMA, smooth muscle actin; STZ, streptozotocin; DiI, one of several dialkylcarbocyanines; EPC, endothelial progenitor cells.
346
is due to the fact that T1DM can be easily induced by in-
traperitoneal injection of streptozotocin, whereas T2DM is
more difficult to induce or requires the purchase of costly
genetically modified animals. So, in SC-for ED field, T1DM
model was used in 3 studies and T2DM in 1. Aging-related
ED is largely manageable with PDE5 inhibitors, so SC
studies in this category (a total of 3) were most likely moti-
vated by not requiring any special treatment on the animal.
Finally, hyperlipidemia-associated ED, which requires feed-
ing animals with costly high-fat diet, was used in one study.
Stem cells
Types of SCs that have been used in experimental ED
treatment (number of studies in brackets) are bone marrow
(6), adipose (5), skeletal muscle (2), embryonic (1), endothe-
lial progenitor (1), and umbilical cord blood (1, a clinical
trial). Some studies have provided reason for choosing 1
particular SC type over the other; however, personal pref-
erences and/or preexisting circumstances (eg, prior experi-
ence with a particular type of SC) probably played bigger
roles than sound scientific rationales did. In any case, the
pros and cons of various types of SCs can be found abun-
dantly elsewhere and will not be discussed here.
Nearly all transplantations were done either autologously
or allogeneically, the single exception being from mouse to
rat. The transplanted cell number averages around 1 million
per recipient rat. Most studies (total of 10) employed un-
modified SCs, whereas others used SCs that were transfected
or fractionated or in combination with other agents. Trans-
fection or fractionation inevitably introduces risk factors (eg,
virus) into the system and/or substantially reduces the
number of treatment cells. Thus, knowing that the majority
of studies employed unmodified SCs with satisfactory
outcomes, the need to use modified or sorted SCs requires
additional evidence.
Cell labeling
A wide variety of methods have been used to monitor the
distribution and survival of transplanted SCs. In SC-ED field,
2 studies by the same group of researchers transplanted
human SCs to rats; therefore, cell tracking was done by the
identification of human-specific nuclear protein. However,
FIG. 3. A schematic repre-
sentation of the experimental
procedures of a typical pre-
clinical stem cell therapy for
ED. The donor rat and recip-
ient rat can be the same
(autologous) or different (al-
logeneic). The isolation and
cultivation of SCs vary from
one type to another. Mod-
ification and sorting of SCs are
desired by some researchers,
although a higher benefit ver-
sus risk ratio has not been
demonstrated. Labeling of
SCs, which is unnecessary if
from a GFP donor rat, usually
incorporates a chemical agent
that can be later detected by
color or fluorescence. Trans-
plantation of the labeled SCs
has so far been conducted
universally by IC injection, as
indicated with the cross-
sectional view of the penis of
an animal whose erectile func-
tion has been compromised by
various means, for example,
CN injury and streptozotocin
injection. Weeks or months
after SC transplantation, the
animals are tested for erectile
function, usually by measure-
ment of increases in intra-
cavernous pressure (ICP)
during electrostimulation of
CN. The animals are then sacrificed for histological assessment of corpus cavernosum and identification of the injected SCs. SCs,
stem cells; GFP, green fluorescence protein; IC, intracavernous; CN, cavernous nerve.
STEM CELL THERAPY FOR ERECTILE DYSFUNCTION 347
these studies did not assess erectile function and thus will
not be discussed further. All other SC-ED studies used cells
labeled with LacZ, 4’,6-diamidino-2-phenylindole (DAPI),
green fluorescent protein (GFP), DiI (also known as PKH-26),
bromodeoxyuridine (BrdU), or 5-ethynyl-2-deoxyuridine
(EdU). Because the accuracy of data interpretation depends
on the reliability of these labeling methods, potential prob-
lems are summarized below.
LacZ is a bacterial gene that encodes b-galactosidase (b-
gal); however, many mammalian cells and tissues contain
endogenous b-gal, making the detection of LacZ-transfected
cells after their transplantation technically challenging [34].
GFP is a protein from jellyfish. However, because of auto-
fluorescence in mammalian tissues, GFP detection can seem
like ‘‘seeing the wood through the trees’ [35]. DAPI binds to
DNA noncovalently; therefore, it can leak from labeled cells
after transplantation and be adsorbed by host cells, resulting
in false-positive detection [36]. DiI binds to cell membrane
noncovalently and can leak from transplanted cells to host
cells. In addition, because of DiI’s cytotoxicity, transplanted
cell preparations may contain debris of dead cells. Adsorp-
tion of this DiI-labeled debris by host cells can lead to false-
positive identification [37–41]. BrdU is incorporated into
newly synthesized DNA and such labeled cells are detected
with anti-BrdU antibody. However, the immnodetection of
BrdU requires harsh treatment of tissue samples, resulting in
distorted histological images [42]. In addition, the denaturing
treatment can cause loss of antigenicity of cellular proteins,
making it impossible to detect cellular differentiation
through immunohistochemical colocalization of cell type-
specific protein. Even if the protein of interest survives the
denaturing treatment, it is still difficult to identify the BrdU
label with confidence, because its brown color cannot be
easily distinguished from the purplish nuclear stain [43].
EdU is a newer thymidine analog and is detected by a simple
chemical reaction that requires no special tissue treatment
[42]. However, similar to BrdU, if a labeled cell is replicative
after transplantation, its EdU label gets diluted with each
round of cell division. So, long-term detection of trans-
planted cells is possible only if the cells are relatively qui-
escent. In our experience with EdU-labeled adipose-derived
SCs (ADSCs), their detection in transplanted tissue is possi-
ble for at least 5 months after transplantation.
Stem cell transplantation
Before the introduction of PDE5 inhibitors, which are ta-
ken orally, IC injection of erectogenic agents was the most
effective treatment for ED [4]. Indeed, even nowadays pa-
tients who do not respond to or cannot take PDE5 inhibitors
are still prescribed with IC injection of vasodilators. As this
route of drug administration targets the organ of failure di-
rectly, it is commonly believed that IC injection is a locally
applied intervention. However, we have observed that IC
injected growth factors were able to restore erectile function
through repair of damaged CN, whose cell bodies reside in
the MPG [44–46]. Further, in our first SC-for-ED study, we
observed that intracavernously injected SCs could treat CN
injury-related ED [13], and all subsequent studies using dif-
ferent types of SCs confirmed the validity of this injection
method for treating CN injury-related ED. Together, these
data strongly suggest that IC injection is systemic in nature.
In all of our published SC-for-ED studies we reported
difficulties in finding the transplanted SCs in penile tissues
even though the animals clearly demonstrated functional
and structural improvements [13,22,23,26]. In studies pub-
lished by others, intracavernously injected SCs were simi-
larly difficult to find. In one of our studies we examined the
presence of SCs in the penis at 2, 14, and 28 days after IC
injection; the results clearly showed a time-dependent de-
cline of the number of SCs [23]. In our recent studies we
conducted more definitive quantitative analyses and the re-
sults showed that the majority of intracavernously injected
SCs exited the penis within 1 day [32,33]. Further, in 1 of
these 2 studies we showed that intracavernously injected SCs
preferentially traveled to the bone marrow [32]; in the other
we found that intracavernously injected SCs also traveled to
the MPG of CN injury rats, and this appears to be mediated
by upregulated SDF-1 in the MPG [33]. Together, these data
suggest that (i) IC injection is essentially like intravenous (IV)
injection—because of the fact that the cavernous sinusoids
are essentially bundled venules (Fig. 1), (ii) the therapeutic
efficacy of SCs for CN injury is due to SC trafficking to the
MPG, (iii) systemically (IC or IV) applied SCs home-in to
bone marrow, in support of the concept that mesenchymal
SCs originate from bone marrow, and (iv) home-in of SCs to
bone marrow may permit establishment of SC reservoirs for
sustained regenerative and/or repair activities.
Functional assessment
In animal experimentation, the most commonly used
method for functional assessment of erection is measurement
of intracavernous pressure (ICP) during electrostimulation of
CN. This procedure requires laparotomy followed by sacri-
ficing the animals; therefore, it is done near the end (most
commonly at 1 month post-treatment) of a preclinical trial.
As mentioned in the Rationale for Using SC Therapy section
earlier, sexual stimulation triggers CN to release NO, which
then causes CSMC relaxation and sinusoidal engorgement.
In animal experiments, stimulation of CN with electric cur-
rent mimics sexual stimulation and causes an increase of ICP
that can approach systemic blood pressure, depending on
the amperage of the applied electric current. Typically, at
settings of 1.5 mA, 20 Hz, and pulse width 0.2 ms, the elec-
trostimulation causes an increase of ICP (in cmH
2
O) from a
baseline of 20 to around 100 in normal rats. In ED rats, the
rise is usually to around 30, and a successful SC treatment
usually restores the value to about 70.
Histological assessment
At the end of functional assessment, penile tissues are
commonly prepared for examination by immunohisto-
chemistry or immunofluorescence. The purposes of these
examinations are to (i) locate transplanted SCs, (ii) correlate
structural with functional changes, and (iii) identify possible
SC differentiation. Localization of transplanted cells was
discussed earlier under Cell labeling section. Assessment of
structural changes invariably focuses on the 3 key compo-
nents that regulate penile erection, ECs, CSMCs, and CN.
ECs are commonly identified with antibodies against rat
endothelial cell antigen, CD31, endothelial nitric oxide syn-
thase, and/or von Willebrand factor (vWF). CSMCs are most
348 LIN ET AL.
commonly detected with anti-smooth muscle actin (anti-
SMA) antibody. The most functionally relevant marker for
CN is nNOS, as it identifies NO-releasing nerve fibers.
The concept of SC therapy was originally based on the
premise that SCs have the ability to differentiate into var-
ious cell lineages. Thus, most SC therapy studies have
strived to identify such events by checking whether the
labeled SCs express cell type-specific proteins such as CD31
for ECs and SMA for CSMCs. So, it is obvious that the
accurate identification of cell differentiation depends on the
reliability of the SC trait/label, the differentiated cell mar-
ker, and the histological image. As discussed earlier in the
Cell labeling section, with the exception of EdU, cell labels
that have been employed in SC-ED studies cannot be de-
tected with confidence. Moreover, histological images pre-
sented in most SC-ED studies are of low resolution and
thus difficult to judge whether the so-called protein ex-
pression is indeed cellularly localized. In our experience,
seemingly colocalized stains at low magnification often
turned out not to be cellularly associated when viewed at
higher mag nifications (eg, 1,00 0 ·). Thus, it is crucial that
claims of cell differentiation be backed by clearly discern-
able histological images.
Future Directions
As CN injury- and DM-related ED patients are less re-
sponsive to PDE5 inhibitors, these 2 types of ED will con-
tinue to be the main targets for future research. Current CN
injury rat models have been well characterized and are
clinically relevant; they can thus continue to be used for fu-
ture SCs for ED research. On the other hand, current DM
models either are too costly or do not adequately represent
clinical situations. To mitigate these problems, we recently
established an inexpensive T2DM rat model that exhibits ED
symptoms and penile structural changes that resemble those
of T2DM patients [8]. This model can thus generate more
clinically relevant data in future SC-ED studies.
With regard to choosing a particular SC type, it is im-
portant to consider what is most practically applicable in
clinical situations. At present, ADSCs is the only cell type
that can be isolated and autologously transplanted on a
same-day basis. Further, several devices for automated iso-
lation of ADSCs are now commercially available. Thus, in
terms of cost, risks, ethics, expediency, and effectiveness,
ADSCs should compete very favorably. The most significant
risk—promotion of tumor growth—is shared by different
types of SCs and requires further research.
As we have now demonstrated that IC injection is similar
to IV injection [32], it is advisable that future SC-ED studies
examine the systemic distribution of the transplanted SCs.
This of course requires that the cells be labeled with a reliable
tracking dye. In our experience with many commonly em-
ployed dyes such as BrdU, DiI, DAPI, GFP, and EdU, we
have found that labeling with EdU is the easiest and most
reliable. With regard to SC distribution, we advise the ex-
amination of bone marrow as SCs seem to have a tendency to
travel there. What do these SCs do in bone marrow is the
next question that needs to be addressed. Specifically, do
they play any role in terms of short-term and long-term
therapeutic efficacy? Finally, the issue of cellular differenti-
ation versus paracrine action needs to be further investi-
gated. To do so, first, it is important to know that, although
cellular differentiation is a presumed SC property, it does not
have to happen in order for SC to exert therapeutic effects.
With this concept in mind, then, a reliable tracking dye is
used to label the SCs, followed by generating high-quality
high-resolution histological images, and it should be possible
to make accurate and unbiased interpretations.
Acknowledgments
This work was supported by grants from the Arthur
Rock Foundation and the National Institutes of Health
(DK045370).
Author Disclosure Statement
No competing financial interests exist.
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Address correspondence to:
Dr. Ching-Shwun Lin
Knuppe Molecular Urology Laboratory
Department of Urology
School of Medicine
University of California
San Francisco, CA 94143-0738
E-mail: clin@urology.ucsf.edu
Received for publication June 13, 2011
Accepted after revision July 26, 2011
Prepublished on Liebert Instant Online July 27, 2011
STEM CELL THERAPY FOR ERECTILE DYSFUNCTION 351
... ED is a major health problem affecting the life quality of more than 150 million men worldwide, and this number is predicted to reach approximately 322 million by 2025 [1,2]. Although substantial advances have been achieved in the pathophysiological mechanisms of ED, ultimately, it seems few effective therapies for various clinical cases, especially for cavernous nerves (CNs) injury related ED [3][4][5]. To address this issue, various cutting-edge therapeutic strategies have been investigated and one of which is currently being extensively evaluated is SC therapy [3]. ...
... Although substantial advances have been achieved in the pathophysiological mechanisms of ED, ultimately, it seems few effective therapies for various clinical cases, especially for cavernous nerves (CNs) injury related ED [3][4][5]. To address this issue, various cutting-edge therapeutic strategies have been investigated and one of which is currently being extensively evaluated is SC therapy [3]. Attracted by various signals released by injured tissues, such as growth factors and cytokines, SCs can migrate to the injured tissue [6]. ...
... MPG and corpus cavernosum were locally injected with PBS, BPNS, SDF1-α, and BP@SDF1-α, respectively (details were shown above). The tissues were harvested at different time points (1,3,7, and 14 days post treatment) for fluorescent staining. ...
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Stem cell (SC) therapy has been shown high prospects in erectile dysfunction (ED) treatment. Without ethical issues and risks of immune rejection and tumorigenesis of exogenous SC therapy, endogenous stem/progenitor cells (S/PCs) have a better potential for ED management, and their homing and redistribution are controlled by SDF1-α/CXCR4 axis. Considering black phosphorus nanosheet (BPNS) has emerged as an efficient and safe drug vehicle due to its large surface area, biodegradability, and the ability to retain and slowly release its loaded drugs, BPNS is utilized to load SDF1-α, a chemokine for S/PCs, to construct the BP@SDF1-α complex to efficiently recruit stem cells (SCs) by injury-site injection and thus ameliorate ED within the bilateral cavernous nerve injury (BCNI) rat models. We find that BP@SDF1-α can efficiently recruit exogenous SCs and endogenous S/PCs to corpus cavernosum and main pelvic ganglion (MPG) by local administration. Of note, ascribing to endogenous S/PCs recruitment, it also successfully alleviates ED in BCNI rat models by enhancing the protein expression levels of α-SMA, CD31, and nNOs, and eliciting less collagen deposition in the penis after its combined injection at corpus cavernosum and MPG. Thus, this study provides a new insight into the treatment of ED with endogenous S/PCs. Biodegradable Nano Black Phosphorus based SDF1-α delivery system ameliorates Erectile Dysfunction in a cavernous nerve Injury Rat Model by recruiting endogenous Stem/Progenitor cells
... Erection in the long term remains unknown. Complications such as diabetes mellitus or chronic rejection affecting the vascular tree may cause secondary impotence, which may need to be addressed by medical [46], stem-cell therapy [47][48][49][50][51][52][53][54][55][56][57][58][59][60][61][62], or surgical treatment. However, surgical interventions like the insertion of a surgical prosthesis may carry additional risks of infection and possible rejection. ...
... Both small and large animal models are being utilized for studying erectile dysfunction related to novel drug discovery, including diabetic, castration, smoking, and hypercholesterolemia models [63]. Stem cell therapy for erectile dysfunction has shown promise in animal models [50,52,53,57,61,[64][65][66][67][68][69][70][71][72][73][74][75][76][77] and clinical trials [60,[78][79][80][81][82][83][84][85]; however, its routine clinical translation requires further study. Additionally, small and large animal models of cavernous nerve reconstruction to restore erectile function have been investigated [86][87][88][89][90][91][92][93][94]. ...
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Animal research is an essential contributor to the medical achievements of the last century. The first step of studying a disease in animals is the development of a model which is relevant to the clinical situation in humans. Thus, a good animal model is the sine qua non of the experimental research. This review aims to assess the contemporary literature on animal models for penile transplantation, examining their applicability and significance in the context of clinical scenarios. We also revisit, evaluate, and emphasize the interesting and important findings of certain animal models to bring the reader up to date from the perspective of allotransplantation. Their current and future clinical applicability and feasibility have been discussed, shedding light on worldwide experience in Vascularized Composite Allotransplantation (VCA).
... The remaining references listed in Figure 6 are detailed in the References List. [37][38][39][40][41][42][43][44][45][46][47][48] ...
... They have published a large number of studies, many of which were highimpact documents, and have made significant contributions to the development of the field. 38,42,43 Publications on the application of SCT in ED tended to be published in specialized journals of urology and andrology, such as The Journal of Sexual Medicine, Andrology, and Asian Journal of Andrology. Moreover, an increasing number of high-quality documents have been published in journals in other fields and comprehensive journals in recent years, 49,57,58 indicating that the scientific research community is widely recognizing research in this field. ...
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Purpose As a common male disease, erectile dysfunction (ED) seriously affects the physical and mental health of patients. In recent years, studies have continued to point out the great potential of stem cell therapy (SCT) in the treatment of ED. The purpose of this study is to comprehensively analyze the research of SCT for ED and understand the development trends and research frontiers in this field. Methods Publications regarding SCT and ED were retrieved and collected from the Web of Science Core Collection. CiteSpace and VOSviewer software were then utilized for bibliometric and visualization analysis. Results A total of 524 publications were eventually included in this study. The annual number of publications in this field was increasing year by year. China and the USA were the two most productive countries. Lin GT, Lue TF and Lin CS, and the University of California San Francisco where they worked were the most productive research group and institution, respectively. The journal with the largest number of publications was The Journal of Sexual Medicine, and the following were mostly professional journals of urology and andrology. Diabetes mellitus-induced ED and cavernous nerve injury-related ED were the two most commonly constructed models of ED in studies. Concerning the types of stem cells, mesenchymal stem cells derived from adipose and bone marrow were most frequently used. Moreover, future research would mainly focus on exosomes, tissue engineering technology, extracorporeal shockwave therapy, and clinical translation. Conclusion The research of SCT for ED will receive increasing global attention in the future. Our study provided bibliometric and visualization analysis of published literature, helping researchers understand the global landscape and frontiers in this field. More preclinical and clinical studies should be conducted to more deeply explore the underlying mechanisms of treatment and promote clinical translation.
... However, it is noteworthy that the retention of injected cells within the penis is remarkably low, often falling below 1%, and these cells typically dissipate within a short span of a few days. Despite the limited duration of cell presence, their mechanism elicits a notable response regarding tissue recovery [54][55][56][57][58]. ...
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Male infertility arises from a complex interplay of factors affecting reproductive organs and various physiological pathways. Among these, erectile dysfunction (ED), a widespread global issue, plays a key role. While existing ED treatments address some aspects, achieving complete reversibility and avoiding side effects remains a challenge. In this context, stem cell therapy emerges as a promising, potentially transformative approach. Preliminary evidence from preclinical animal models and clinical trials highlights stem cell therapy’s remarkable efficacy and effectiveness for ED. This novel strategy offers several advantages, including enhanced effectiveness and a reported absence of adverse side effects. This review delves into the causes of male infertility, with a particular focus on ED and its pathophysiology. We explore the current treatment landscape, highlighting therapy’s existing strategies’ limitations and stem cell therapy’s unique potential. By examining relevant preclinical and clinical studies, we provide a comprehensive picture of this innovative approach and its promising future in restoring men’s fertility and quality of life.
... SCs differentiate into CN cells, SMCs, or ECs [75]. Second, SCT could heal penile tissue through a paracrine effect which induces anti-inflammatory and anti-apoptotic effect [76]. In most of the animal studies, intracavernosal pressure (ICP), and the ratio of ICP and mean arterial pressure were analyzed to assess EF. ...
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Objective This systematic review aimed to analyze animal and human trial data to better understand the efficacy of stem cell therapy (SCT) for erectile dysfunction (ED) and the obstacles that may hinder its application in this field. Methods We searched electronic databases, including PubMed and Scopus, for published studies with the Medical Subject Heading terms of “erectile dysfunction” (AND) “stem cell therapy” (OR) “erectile dysfunction” (AND) “clinical trial of stem cell therapy” (OR) “stem cell therapy” (AND) “sexual dysfunction”. The search was limited to English-language journals and full papers only. The initial search resulted in 450 articles, of which 90 relevant to our aims were included in the analysis. Results ED is a multifactorial disease. Current treatment options rely on pharmacotherapy as well as surgical options. Patients may have side effects or unsatisfactory results following the use of these treatment options. SCT may restore pathophysiological changes leading to ED rather than treating the symptoms. It has been evaluated in animal models and shown promising results in humans. Results confirm that SCT does improve erectile function in animals with different types of SC use. In humans, evidence showed promising results, but the trials were heterogeneous and limited mainly by a lack of randomization and the small sample size. Many challenges could limit future research in this field, including ethical dilemmas, regulation, patient recruitment, the cost of therapy, and the lack of a standardized SCT regimen. Repairing and possibly replacing diseased cells, tissue, or organs and eventually retrieving normal function should always be the goals of any therapy, and this can only be guaranteed by SCT. Conclusion SCT is a potential and successful treatment for ED, particularly in patients who are resistant to the classic therapy. SCT may promote nerve regeneration and vascular cell regeneration, not only symptomatic treatment.
... Preclinical research in animal models has corroborated improvements in ED following stem cell administration and has also confirmed in vitro stem cell differentiation in these cell lines [18,19]. ...
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Adipose tissue-derived stem cells (ADSCs) have been shown to improve erectile function in animal models of erectile dysfunction. However, few studies have been carried out using a reliable in vivo imaging method to trace transplanted cells in real time, which is necessary for systematic investigation of cell therapy. The study aims to explore the feasibility of non-invasively monitoring intracavernous injection of ADSCs in rat and miniature pig corpus cavernosum using in vivo magnetic resonance (MR) imaging. Thirty-six male Sprague Dawley rats (10 weeks old) and six healthy, sexually mature male miniature pigs (20 kg weight) were obtained. ADSCs were isolated from paratesticular fat of donor rats and cultured. Then ADSCs were labeled with superparamagnetic iron oxide nanoparticles (SPIONs), a type of MR imaging contrast agent, before transplantation into rats and pigs. After intracavernous injection, all rats and pigs underwent and were analyzed by MR imaging at the day of ADSC transplantation and follow-up at 1, 2 and 4 weeks after transplantation. In addition, penile histological examination was performed on all rats and pigs before (n = 6) and at 1 day (n = 6), 1 week (n = 6), 2 weeks (n = 6) or 4 weeks (n = 12) after ADSC transplantation. SPION-labeled ADSCs demonstrated a strong decreased signal intensity compared with distilled water, unlabeled ADSCs or agarose gel. SPION-labeled ADSCs showed a hypointense signal at all concentrations, and the greatest hypointense signal was observed at the concentration of 1 × 10⁶. MR images of the corpus cavernosum showed a hypointense signal located at the injection site. T2*-weighted signal intensity increased over the course of 1 week after ADSCs transplantation, and demonstrated a similar MR signal with that before ADSCs transplantation. After SPION-labeled ADSC injection, T2*-weighted MR imaging clearly demonstrated a marked hypointense signal in pig corpus cavernosum. The T2*-weighted signal faded over time, similar to the MR imaging results in rats. Obvious acute inflammatory exudation was induced by intracavernous injection, and the T2*-weighted signal intensity of these exudation was higher than that of the injection site. The presence of iron was detected by Prussian blue staining, which demonstrated ADSC retention in rat corpus cavernosum. Lack of cellular infiltrations were demonstrated by H&E staining before and 4 weeks after transplantation, which indicated no negative immune response by rats. Prussian blue staining was positive for iron oxide nanoparticles at 2 weeks after transplantation. SPION-labeled ADSCs showed a clear hypointense signal on T2-weight MRI in vitro and in vivo. The MR signal intensity in the corpus cavernosum of the rats and miniature pigs faded and disappeared over time after ADSC transplantation. These findings suggested that MR imaging could trace transplanted ADSCs in the short term in the corpus cavernosum of animals.
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Introduction: While phosphodiesterase type 5 inhibitors (PDE5is) and others are used to treat Erectile dysfunction (ED), many patients are either unresponsive or resistant to it. Stem cell therapy (SCT) is a promising alternative approach. Numerous preclinical trials have demonstrated improved erectile function in animal models using SCT, although the number of clinical trials investigating SCT for men with ED is limited. Nonetheless, findings from human clinical trials suggest that SCT may be a useful treatment option. Areas covered: Biomedical literature, including PubMed, ClinicalTrials.gov, and European Union Clinical Trails Registry, were analyzed to summarize and synthesize information on stem cell therapy for ED in this narrative review. The achievements in preclinical and clinical evaluations are presented and critically analyzed. Expert opinion: SCT has demonstrated some benefits in improving erectile function, while further studies are urgently needed. Such studies would provide valuable insights into the optimal use of stem cell therapy and its potential as a therapeutic option for ED. Taking advantage of different mechanisms of action involved in various regenerative therapies, combination therapies such as SCT and low-energy shock waves or platelet-rich plasma may provide a more effective therapy and warrant further research.
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ERECTILE DYSFUNCTION: CURRENT APPROACH AND EMERGING STRATEGIES
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The intracavernous (i.c.) injection of stem cells (SCs) has been shown to improve erectile function in various erectile dysfunction (ED) animal models. However, the tissue distribution of the injected cells remains unknown. In this study we tracked i.c.-injected adipose-derived stem cells (ADSCs) in various tissues. Rat paratesticular fat was processed for ADSC isolation and culture. The animals were then subject to cavernous nerve (CN) crush injury or sham operation, followed by i.c. injection of 1 million autologous or allogeneic ADSCs that were labeled with 5-ethynyl-2-deoxyuridine (EdU). Another group of rats received i.c. injection of EdU-labeled allogeneic penile smooth muscle cells (PSMCs). At 2 and 7 days post injection, penises and femoral bone marrow were processed for histological analyses. Whole femoral bone marrows were also analyzed for EdU-positive cells by flow cytometry. The results show that ADSCs exited the penis within days of i.c. injection and migrated preferentially to bone marrow. Allogenicity did not affect the bone marrow appearance of ADSCs at either 2 or 7 days, whereas CN injury reduced the number of ADSCs in bone marrow significantly at 7 but not 2 days. The significance of these results in relation to SC therapy for ED is discussed.
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Lower urinary tract diseases are emotionally and financially burdensome to the individual and society. Current treatments are ineffective or symptomatic. Conversely, stem cells (SCs) are regenerative and may offer long-term solutions. Among the different types of SCs, bone marrow SCs (BMSCs) and skeletal muscle-derived SCs (SkMSCs) have received the most attention in pre-clinical and clinical trial studies concerning the lower urinary tract. In particular, clinical trials with SkMSCs for stress urinary incontinence have demonstrated impressive efficacy. However, both SkMSCs and BMSCs are difficult to obtain in quantity and therefore neither is optimal for the eventual implementation of SC therapy. On the other hand, adipose tissue-derived SCs (ADSCs) can be easily and abundantly obtained from "discarded" adipose tissue. Moreover, in several head-on comparison studies, ADSCs have demonstrated equal or superior therapeutic potential compared to BMSCs. Therefore, across several different medical disciplines, including urology, ADSC research is gaining wide attention. For the regeneration of bladder tissues, possible differentiation of ADSCs into bladder smooth muscle and epithelial cells has been demonstrated. For the treatment of bladder diseases, specifically hyperlipidemia and associated overactive bladder, ADSCs have also demonstrated efficacy. For the treatment of urethral sphincter dysfunction associated with birth trauma and hormonal deficiency, ADSC therapy was also beneficial. Finally, ADSCs were able to restore erectile function in various types of erectile dysfunction (ED), including those associated with diabetes, hyperlipidemia, and nerve injuries. Thus, ADSCs have demonstrated remarkable therapeutic potentials for the lower urinary tract.
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Objective To explore the effects of transplantation of bone marrow-derived mesenchymal stem cells (BM-MSCs) on improving erectile function of streptozocin (STZ)-induced diabetic rats. Methods Male Sprague Dawley rats were injected either with STZ to induce diabetes or with citrate buffer as controls. Rat BM-MSCs were harvested and labeled with Chloromethyl Benzamido derivatives of 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanine perchlorate (CM-DiI), and then transplanted into corporal cavernosum of STZ-induced diabetic rats. Four weeks after transplantation, all rats were analyzed for erectile function and penile histology. Results After BM-MSCs transplantation, the ICP/MAP ratio was increased significantly compared with diabetic controls. Content of smooth muscle and endothelium in corporal cavernosa of BM-MSCs transplanted rats was significantly increased compared to diabetic controls. Immunofluorescence analysis demonstrated that CM-DiI-labeled BM-MSCs could stay in corporal cavernosa for at least 4 weeks and some of them expressed von Willebrand Factor, CD31, calponin, or α-smooth muscle actin, cells markers for endothelial cells or smooth muscle cells, respectively. Conclusions Intracavernous transplantation of BM-MSCs had beneficial effects on erectile function of diabetic rats and increased the content of endothelium and smooth muscle in corporal cavernosum.
Article
OBJECTIVE To test the hypothesis that an intracavernosal injection with brain-derived neurotrophin factor (BDNF) and vascular endothelial growth factor (VEGF) can facilitate nerve regeneration and recovery of erectile function after cavernosal nerve injury.MATERIALS AND METHODS The study included 25 Sprague-Dawley rats; four had a sham operation, seven bilateral nerve crushing with no further intervention, and 14 bilateral nerve crushing with either an immediate (seven) or delayed for 1 month (seven) intracavernosal injection with BDNF+VEGF. Erectile function was assessed by cavernosal nerve electrostimulation at 3 months, and neural regeneration by NADPH-diaphorase staining and tyrosine hydroxylase (TH) staining of penile tissue and major pelvic ganglia (MPG).RESULTSAfter nerve crushing, the functional evaluation at 3 months showed a lower mean (sd) intracavernosal pressure (ICP) with cavernosal nerve stimulation, at 33.9 (15.3) cmH2O, than in the sham group, at 107.8 (18.1) cmH2O. With an immediate injection with BDNF+VEGF the ICP was significantly higher than in the controls, at 67.8 (38.5) cmH2O. Even delayed injection with BDNF+VEGF improved the ICP, to 78.0 (21.8) cmH2O. Histological analysis of specimens stained for NADPH and TH showed a significant change in the morphology of terminal branches of the cavernosal and dorsal nerves, and the staining quality of the neurones in the MPG. The number of positively stained nerve fibres tended to revert to normal after treatment with BDNF+VEGF.CONCLUSION An intracavernosal injection with BDNF+VEGF appears to both prevent degeneration and facilitate regeneration of neurones containing neuronal nitric oxide synthase in the MPG, dorsal nerve and intracavernosal tissue. Therefore it might have therapeutic potential for enhancing the recovery of erectile function after radical pelvic surgery.
Article
What's known on the subject? and what does the study add? Increased cavernous smooth muscle content has been repeatedly observed in rat models of hyperlipidaemia - associated erectile dysfunction. This study shows that the increased smooth muscle content is due to hyperplasia. • To investigate the structural changes, including possible smooth muscle hyperplasia, in the penis of a hyperlipidaemia-associated erectile dysfunction (ED) animal model. • Hyperlipidaemia was induced in rats through a high-fat diet. • Penile tissues of normal and hyperlipidaemic rats were stained with Alexa-488-conjugated phalloidin and/or with antibodies against rat endothelial cell antigen, neuronal nitric oxide synthase (nNOS), and collagen type IV (Col-IV) before image and statistical analyses were carried out. • The main outcome measures were the smooth muscle, endothelial, Col-IV and nNOS content of the corpus cavernosum. • Phalloidin intensely stained all smooth muscle in the penis, revealing the circular and longitudinal components of cavernous smooth muscle (CSM). • The CSM content was significantly higher in the hyperlipidaemic than in the normal rats (P < 0.05). • Cell numbers in both circular and longitudinal CSM were significantly higher in the hyperlipidaemic than in the normal rats (P < 0.05). • Cavernous endothelial content was significantly lower in hyperlipidaemic than in normal rats (P < 0.05). • nNOS-positive nerves within the dorsal nerves, around the dorsal arteries, and in the corpora cavernosa were all significantly lower in the hyperlipidaemic than in the normal rats (P < 0.05). • Hyperlipidaemia is associated with reduced nNOS-positive nerves, reduced endothelium, and increased CSM in the penis. • The increased CSM is attributable to hyperplasia. • These structural changes may explain why hyperlipidaemic men are more likely to develop ED.
Article
To investigate whether fluorochrome-conjugated phalloidin can delineate cavernous smooth muscle (CSM) cells and whether it can be combined with immunofluorescence (IF) staining to quantify erectile dysfunction (ED)-associated changes. ED was induced by cavernous nerve crush in rats. Penile tissues of control and ED rats were stained with Alexa-488-conjugated phalloidin and/or with antibodies against rat endothelial cell antigen (RECA), CD31, neuronal nitric oxide synthase (nNOS), and collagen-IV (Col-IV). Phalloidin was able to delineate CSM as composed of a circular and a longitudinal compartment. When combined with IF stain for CD31 or RECA, it helped the identification of the helicine arteries as covered by endothelial cells on both sides of the smooth muscle layer. When combined with IF stain for nNOS, it helped the identification that nNOS-positive nerves were primarily localized within the dorsal nerves and in the adventitia of dorsal arteries. When combined with IF stain for Col-IV, it helped identify that Col-IV was localized around smooth muscles and beneath the endothelium. Phalloidin also facilitated the quantitative analysis of ED-related changes in the penis. In rats with cavernous nerve injury, RECA or Col-IV expression did not change significantly, but CSM and nNOS nerve contents decreased significantly. Phalloidin stain improved penile histology, enabling the visualization of the circular and longitudinal compartments in the CSM. It also worked synergistically with IF stain, permitting the visualization of the dual endothelial covering in helicine arteries, and facilitating the quantification of ED-related histologic changes.
Article
Intracavernous (IC) injection of stem cells has been shown to ameliorate cavernous-nerve (CN) injury-induced erectile dysfunction (ED). However, the mechanisms of action of adipose-derived stem cells (ADSC) remain unclear. To investigate the mechanism of action and fate of IC injected ADSC in a rat model of CN crush injury. Sprague-Dawley rats (n=110) were randomly divided into five groups. Thirty-five rats underwent sham surgery and IC injection of ADSC (n=25) or vehicle (n=10). Another 75 rats underwent bilateral CN crush injury and were treated with vehicle or ADSC injected either IC or in the dorsal penile perineural space. At 1, 3, 7 (n=5), and 28 d (n=10) postsurgery, penile tissues and major pelvic ganglia (MPG) were harvested for histology. ADSC were labeled with 5-ethynyl-2-deoxyuridine (EdU) before treatment. Rats in the 28-d groups were examined for erectile function prior to tissue harvest. IC pressure recording on CN electrostimulation, immunohistochemistry of the penis and the MPG, and number of EdU-positive (EdU+) cells in the injection site and the MPG. IC, but not perineural, injection of ADSC resulted in significantly improved erectile function. Significantly more EdU+ ADSC appeared in the MPG of animals with CN injury and IC injection of ADSC compared with those injected perineurally and those in the sham group. One day after crush injury, stromal cell-derived factor-1 (SDF-1) was upregulated in the MPG, providing an incentive for ADSC recruitment toward the MPG. Neuroregeneration was observed in the group that underwent IC injection of ADSC, and IC ADSC treatment had beneficial effects on the smooth muscle/collagen ratio in the corpus cavernosum. CN injury upregulates SDF-1 expression in the MPG and thereby attracts intracavernously injected ADSC. At the MPG, ADSC exert neuroregenerative effects on the cell bodies of injured nerves, resulting in enhanced erectile response.
Article
To conduct a pilot study to investigate functional, metabolic, and penile morphologic changes in a novel model of lean DM2. Erectile dysfunction (ED) is a frequent sequela in patients with type 2 diabetes mellitus (DM2). Eight rats received a high-fat diet and 2 weeks later, 2 intraperitoneal injections of streptozotocin (STZ, 30 mg/kg). Five age-matched rats served as controls. Insulin challenge tests were performed at 6 and 12 weeks after induction of DM2. At 12 weeks, erectile function was tested by measurement of intracavernous pressure (ICP) increase upon cavernous nerve stimulation. Penile tissue and serum samples were harvested for histology and biochemistry, respectively. A lean DM2 model was established as demonstrated by decreased insulin resistance, elevated nonfasting plasma glucose levels, hyperlipidemia, and decreased insulin concentration in the absence of obesity. ICP/mean arterial pressure was significantly decreased in DM2 animals (0.29) compared with controls (0.81). Expression of neuronal nitric oxide synthase and rat endothelial cell antigen-1, and the smooth muscle/collagen ratio were significantly decreased in the penis of DM2 animals. We propose an inexpensive nongenetic animal model of lean DM2-associated ED. Microanatomical changes in the erectile tissue that reflect an advanced stage of the disease were observed.