Regulation of cavernous nerve injury-induced apoptosis by sonic hedgehog

Thirty to eighty-seven percent of patients treated by radical prostatectomy experience erectile dysfunction (ED). The reduced efficacy of treatments in this population makes novel therapeutic approaches to treat ED essential. We propose that abundant apoptosis observed in penile smooth muscle when the cavernous nerve (CN) is cut (mimicking the neural injury which can result from prostatectomy) is a major contributing factor to ED development. We hypothesize that decreased Sonic hedgehog (SHH) signaling is a cause of ED in neurological models of impotence by increasing apoptosis in penile smooth muscle. We examined this hypothesis in a bilateral CN injury model of ED. We found that the active form of SHH protein was significantly decreased 1.2-fold following CN injury, that SHH inhibition causes a 12-fold increase in smooth muscle apoptosis in the penis, and that SHH treatment at the time of CN injury was able to decrease CN injury-induced apoptosis (1-3-fold) in a dose-dependent manner. These results show that SHH stabilizes the alterations of the corpora cavernosal smooth muscle following nerve injury.     

Neural influences on sonic hedgehog and apoptosis in the rat penis

The role of sonic hedgehog (SHH) in maintaining corpora cavernosal morphology in the adult penis has been established; however, the mechanism of how SHH itself is regulated remains unclear. Since decreased SHH protein is a cause of smooth muscle apoptosis and erectile dysfunction (ED) in the penis, and SHH treatment can suppress cavernous nerve (CN) injury-induced apoptosis, the question of how SHH signaling is regulated is significant. It is likely that neural input is involved in this process since two models of neuropathy-induced ED exhibit decreased SHH protein and increased apoptosis in the penis. We propose the hypothesis that SHH abundance in the corpora cavernosa is regulated by SHH signaling in the pelvic ganglia, neural activity, or neural transport of a trophic factor from the pelvic ganglia to the corpora. We have examined each of these potential mechanisms. SHH inhibition in the penis shows a 12-fold increase in smooth muscle apoptosis. SHH inhibition in the pelvic ganglia causes significantly increased apoptosis (1.3-fold) and decreased SHH protein (1.1-fold) in the corpora cavernosa. SHH protein is not transported by the CN. Colchicine treatment of the CN resulted in significantly increased smooth muscle apoptosis (1.2-fold) and decreased SHH protein (1.3-fold) in the penis. Lidocaine treatment of the CN caused a similar increase in apoptosis (1.6-fold) and decrease in SHH protein (1.3-fold) in the penis. These results show that neural activity and a trophic factor from the pelvic ganglia/CN are necessary to regulate SHH protein and smooth muscle abundance in the penis.     

Altered Sonic hedgehog signaling is associated with morphological abnormalities in the penis of the BB/WOR diabetic rat

Erectile dysfunction (ED) is a common and debilitating pathological development that affects up to 75% of diabetic males. Neural stimulation is a crucial aspect of the normal erection process. Nerve injury causes ED and disrupts signaling of the Sonic hedgehog (Shh) cascade in the smooth muscle of the corpora cavernosa. Shh and targets of its signaling establish normal corpora cavernosal morphology during postnatal differentiation of the penis and regulate homeostasis in the adult. Interruption of the Shh cascade in the smooth muscle of the corpora cavernosa results in extensive changes in corpora cavernosal morphology that lead to ED. Our hypothesis is that the neuropathy observed in diabetics causes morphological changes in the corpora cavernosa of the penis that result in ED. Disruption of the Shh cascade may be involved in this process. We tested this hypothesis by examining morphological changes in the penis, altered gene and protein expression, apoptosis, and bromodeoxyuridine incorporation in the BB/WOR rat model of diabetes. Extensive smooth muscle and endothelial degradation was observed in the corpora cavernosa of diabetic penes. This degradation accompanied profound ED, significantly decreased Shh protein in the smooth muscle of the corpora cavernosa, and increased penile Shh RNA expression in the intact penis (nerves, corpora, and urethra). Localization and expression of Shh targets were also disrupted in the corpora cavernosa. Increasing our understanding of the molecular mechanisms that regulate Shh signaling may provide valuable insight into improving treatment options for diabetic impotence.     

Pharmacologic treatment of erectile dysfunction

Penile erection occurs in response to cavernous smooth muscle relaxation, increased blood flow to the penis, and restriction of venous outflow. These events are regulated by a spinal reflex relying on visual, imaginative, and olfactory stimuli generated within the central nervous system (CNS) and on tactile stimuli to the penis. Drugs can have a facilitatory or inhibitory effect either on the nerves regulating this reflex or on the cavernous smooth muscle. A balance between contractile and relaxant factors governs flaccidity/rigidity within the penis. Drugs that raise cytosolic calcium either prevent or abort erection. Conversely, drugs that lower cytosolic calcium relax smooth muscle and can initiate penile erection. Efficacy in treating erectile dysfunction (ED) with phosphodiesterase inhibitors, especially type 5; alpha-adrenergic-receptor antagonists; and dopamine agonists exploit these mechanisms within the penis or CNS. Recent advances in our understanding of the pharmacology of penile erection are being translated into effective therapies for ED.     

Mesenchymal stem cells alone or ex vivo gene modified with endothelial nitric oxide synthase reverse age-associated erectile dysfunction

Mesenchymal stem cells (MSCs) can be used in adult stem cell-based gene therapy for vascular diseases. To test the hypothesis that MSCs alone or endothelial nitric oxide synthase (eNOS)-modified MSCs can be used for treatment of erectile dysfunction (ED), syngeneic rat MSCs (rMSCs) were isolated, ex vivo expanded, transduced with adenovirus containing eNOS, and injected into the penis of aged rats. Histological analysis demonstrated that rMSCs survived for at least 21 days in corporal tissue after intracavernous injection, and an inflammatory response was not induced. Intracavernous administration of eNOS-modified rMSCs improved the erectile response in aged rats at 7 and 21 days after injection. The increase in erectile function was associated with increased eNOS protein, NOS activity, and cGMP levels. rMSCs alone increased erectile function of aged rats at day 21, but not at day 7, with the transplanted cells exhibiting positive immunostaining for several endothelial and smooth muscle cell markers. This change in rMSC phenotype was accompanied by upregulation of penile eNOS protein expression/activity and elevated cGMP levels. These findings demonstrate that an adenovirus can be used to transduce ex vivo expanded rMSCs to express eNOS and that eNOS-modified rMSCs improve erectile function in the aged rat. Intracavernous injection of unmodified wildtype rMSCs improved erectile function 21 days after injection through mechanisms involving improved endothelium-derived NO/cGMP signaling and rMSC differentiation into penile cells expressing endothelial and smooth muscle markers. These data highlight the potential clinical use of adult stem cell-based therapy for the treatment of ED.     

Downregulation of cGMP-dependent protein kinase-1 activity in the corpus cavernosum smooth muscle of diabetic rabbits

Increased guanosine 3',5'-cyclic monophosphate (cGMP), induced by nitric oxide release, is crucial for corpus cavernosum smooth muscle (CCSM) relaxation within the penis. This CCSM relaxation (necessary for penile erection) is impaired in men with erectile dysfunction (ED), especially those men with diabetes. One of the effector proteins for cGMP is cGMP-dependent protein kinase-1 (PKG-1). PKG-1 knockout mice exhibit detrusor overactivity (Am J Physiol Regul Integr Comp Physiol 279: R1112-R1120, 2000) and, more relevant to this study, ED (Proc Natl Acad Sci USA 97: 2349-2354, 2000), suggesting an in vivo role for PKG-1 in urogenital smooth muscle relaxation. In the current study, using normal rabbit CCSM, Western blot analysis revealed high expression of PKG-1 at levels almost equivalent to aorta (previously shown to have high PKG-1 expression) and that the two known alternatively spliced isoforms of PKG-1 (alpha and beta) are expressed in nearly equal amounts in the CCSM. However, in response to alloxan-induced diabetes, there was a decrease in expression of both PKG-1 isoforms at the mRNA and protein levels as determined by real-time RT-PCR and Western blotting, respectively, but with the PKG-1alpha isoform expression decreased to a greater extent. Moreover, diabetes was associated with significantly decreased PKG-1 activity of CCSM in vitro, correlating with decreased CCSM relaxation. Immunofluorescence microscopy revealed a diabetes-associated decrease in PKG-1 in the CCSM cells. In conclusion, our results demonstrate for the first time a significant downregulation of PKG-1 expression associated with decreased PKG-1 activity in the CCSM in response to diabetes. Furthermore, these results suggest a mechanistic basis for the decreased efficacy of phosphodiesterase V inhibitors in treating diabetic patients with ED.     

The vasculature of the rat penis: a scanning electron microscopic and histologic study.

As a basis for understanding the mechanism of erection in an animal model frequently used in research in reproductive biology, the angioarchitecture of the penis of the rat has been described using scanning electron microscopy. Study of the penile vasculature of the rat indicates that the corpora cavernosa penis and the corpus spongiosum are independent erectile tissues, each with its own arterial and venous vessels. The large vascular spaces and abundant smooth muscle of the penile crura are compatible with its role in regulating blood flow to more distal penile tissues. Helicine arteries of the crura, but not the parent deep penile artery or arteries elsewhere, have muscular cushions in their walls. The venous drainage of the penile crura is via subtunical veins which are thought to be compressed during erection to elevate pressure within the penis. Large, paired cavernous veins drain the shaft of the penis. A unique method for inhibiting blood flow from the penis is indicated by the division of the cavernous veins into smaller channels prior to joining the subtunical venous plexus. Erectile tissue in the bifid origins of the corpus spongiosum has abundant cavernous muscle, while in the remainder of the corpus spongiosum little smooth muscle lines the cavernous spaces. The cavernous spaces on either side of the urethra coalesce to form vessels, each of which communicates with cavernous spaces in the glans. In addition, a bypass of the glans is effected by communication of these vessels directly with the deep dorsal vein. The apparent absence of muscular pads in vessels of the spongiosum, the relative paucity of cavernous smooth muscle, and the ample venous drainage provided by the deep dorsal vein may account for the lack of a venous occlusive mechanism similar to that of the corpora cavernosa penis.     

Antioxidant icariside II combined with insulin restores erectile function in streptozotocin‐induced type 1 diabetic rats

Erectile dysfunction (ED) worsens in patients with diabetes mellitus (DM) despite good control of blood glucose level with insulin. Recent studies imply that diabetic vascular stresses (e.g. oxidative stress) persist in spite of glucose normalization, which is defined as metabolic memory. Studies suggest that the interaction between advanced glycation end products (AGEs) and their receptor (RAGE) mediates the development of metabolic memory. To investigate the effects of the antioxidant icariside II plus insulin on erectile function in streptozotocin (STZ)‐ induced type 1 diabetic rats. Fifty 8‐week‐old Sprague‐Dawley rats were randomly distributed into five groups: normal control, diabetic, insulin‐treated diabetic, icariside II‐treated diabetic, and insulin plus icariside II‐treated diabetic. Diabetes was induced by a single intraperitoneal injection of STZ. Eight weeks after induction of diabetes, icariside II was administered by gastric lavage once a day (5 mg/kg) for 6 weeks; and 2–6 units of intermediate‐acting insulin were given to maintain normal glycemia for 6 weeks. The main outcome measures were the ratio of intracavernous pressure (ICP) to mean arterial pressure (MAP); histology of penile endothelial cells and smooth muscle cells; neural nitric oxide synthase, AGEs and RAGE expression; malondialdehyde concentration; superoxide dismutase activity; and apoptosis index. Diabetic rats demonstrated a significantly lower ICP/MAP ratio, reduced penile endothelial cells, reduced smooth muscle cells, increased AGEs and RAGE, and increased apoptosis. Insulin and icariside II monotherapy partially restored erectile function and histological changes. However, the combination therapy group showed significantly better erectile parameters, cytological components and biochemistry, similar to those in the normal control group. These results suggest that, although insulin can effectively control glycemic levels, it does not completely alter the pathological changes in erectile tissues. Better efficacy could be expected with tight glycemic control plus the antioxidant icariside II. The proposed combination therapy might have the potential to eliminate metabolic memory by down‐regulating the AGEs‐RAGE‐oxidative stress axis.     

内质网应激在大鼠低氧高二氧化碳性肺动脉高压中的作用

目的：观察低氧高二氧化碳性肺动脉高压大鼠的肺血管重塑并探讨内质网应激（ERS）在肺动脉高压中的作用。方法：将40只SD大鼠随机分为四组：常氧对照组（N）、低氧高二氧化碳组（HH）、ERS通路抑制剂4-苯基丁酸（4-phenylbutyric acid）组（4-PBA）、ERS通路激动剂衣霉素（tunicamycin）组（TM）,n=10。测量各组大鼠的肺动脉平均压（mPAP）、颈动脉平均压以及右心室肥大指数,免疫荧光α-SMA标记法鉴定各组肺中小动脉平滑肌细胞,电镜观察肺组织及肺中小动脉形态学变化,原位末端标记法（TUNEL）检测各组肺动脉平滑肌细胞的凋亡指数,采用RT-PCR和Western blot分别检测各组大鼠葡萄糖调节蛋白78（GRP78）、C/EBP同源蛋白（CHOP）、c-Jun氨基末端激酶（JNK）、天冬氨酸特异性半胱氨酸蛋白酶-12（caspase-12）mRNA及蛋白质表达。结果：①与N组相比,HH组、4-PBA组、TM组mPAP、右心室游离壁重量/左心室加心室间隔重量[RV/（LV+S）]、肺动脉管壁面积/管总面积（WA/TA）比值增加（P<0.0 1）,肺动脉管腔面积/管总面积（LA/TA）比值减小（P<0.01）,细胞凋亡指数降低（P <0.05或P<0.01）。ERS相关蛋白质及mRNA的表达量升高,各差异均有统计学意义。②与HH组相比,4-PB A组mPAP和[RV/（LV+S）]、WA/TA值减小（P<0.01）,LA/TA值和细胞凋亡指数上升（P<0.05或P<0.01）,ERS相关蛋白质和mRNA的表达量均下调（P<0.05或P<0.01）;③与HH组相比,TM组mPAP、[RV/（LV+S）]、WA/TA值升高（P<0.05或P<0.01）;肺动脉中膜层增厚,LA/TA值和细胞凋亡指数降低（P<0.01）。ERS相关蛋白质及mRNA的表达量均升高,除GRP78蛋白质表达量无明显变化外,其余各差异均有统计学意义。结论：低氧高二氧化碳诱导的肺动脉高压大鼠肺血管重塑可能与肺动脉平滑肌细胞增殖过度及凋亡过少有关;ERS相关因子（JNK、caspase-12和CHOP）参与低氧高二氧化碳性肺动脉高压的调控。     

Chronic Cigarette Smoking Impairs Erectile Function through Increased Oxidative Stress and Apoptosis,Decreased nNOS,Endothelial and Smooth Muscle Contents in a Rat Model

Cigarette use is an independent risk factor for the development of erectile dysfunction (ED). While the association between chronic smoking and ED is well established, the fundamental mechanism(s) of cigarette-related ED are incompletely understood, partly due to no reliable animal model of smoking-induced ED. The present study was designed to validate an in vivo rat model of chronic cigarette-induced ED. Forty 12-week old male Sprague-Dawley rats were divided into 4 groups. Ten rats served as control group and were exposed only to room air. The remaining 30 rats were passively exposed to cigarette smoke (CS) for 4 weeks (n = 10), 12 weeks (n = 10), and 24 weeks (n = 10). At the 24-week time point all rats were assessed with intracavernous pressure (ICP) during cavernous nerve electrostimulation. Blood and urine were collected to measure serum testosterone and oxidative stress, respectively. Corporal tissue was assessed by Western blot for neuronal nitric oxide synthase (nNOS). Penile tissues were subjected to immunohistochemistry for endothelial, smooth muscle, and apoptotic content. Mean arterial pressure (MAP) was significantly higher in 24-week cigarette exposed animals compared to the control animals. Mean ICP/MAP ratio and cavernosal smooth muscle/endothelial contents were significantly lower in the 12- and 24-week rats compared to control animals. Oxidative stress was significantly higher in the 24-week cigarette exposed group compared to control animals. Mean nNOS expression was significantly lower, and apoptotic index significantly higher, in CS-exposed animals compared to control animals. These findings indicate that the rat model exposure to CS increases apoptosis and oxidative stress and decreases nNOS, endothelial and smooth muscle contents, and ICP in a dose dependent fashion. The rat model is a useful tool for further study of the molecular and cellular mechanisms of CS-related ED.     

Effects of dual endothelin receptor antagonist on antiapoptotic marker Bcl-2 expression in streptozotocin-induced diabetic rats

Erectile dysfunction (ED) affects approximately 50% of male patients with diabetes mellitus (DM) and is possibly due to the vascular and neuropathic complications of DM. Recently, apoptosis has been regarded as a downstream event in ED. More recently, the importance of alterations in apoptosis-related molecules in the mechanism of DM-induced ED has begun to be appreciated. Endothelin-1 (ET-1) plays a role via ET(A) and ET(B) receptors in the regulation of cavernosal smooth-muscle tone in penile tissues. We found that the ET-1 level in the penis of rats with DM was higher than that in the penis of control animals. The present study investigated a rat model in which DM was induced by a 3-week regimen of streptozotocin (STZ) to assess the expression of several apoptosis-related molecules in penile tissue and, concomitantly, the effects of ET antagonism on these changes. Male Sprague-Dawley rats (weight [+/-SD], 450 +/- 26 g) received a citrate saline vehicle or STZ (65 mg/kg ip). DM was confirmed by the presence of hyperglycemia. Diabetic animals were further separated into two treatment groups 1 week after onset of disease: one group received ET(A/B) dual receptor antagonist (SB209670) by means of osmotic minipump at a dosage of 1 mg/day, and the other group received saline. Rats in both groups were treated for 2 weeks and then sacrificed. Plasma glucose levels (+/-SD) in rats with DM were significantly higher than those in rats without DM (506 +/- 70 vs. 111 +/- 11 mg/dl). In the penile tissue of rats with DM, a 35% decrease in the expression of Bcl-2 protein (an important antiapoptotic marker detectable by immunoblotting) was seen, and ET(A/B) dual antagonist was observed to significantly counteract this decrease. Real-time polymerase chain reaction revealed that the expression of Bcl-2 mRNA was consistent with Bcl-2 protein expression. Levels of Bax and caspase-3, two important proapoptotic markers, were not significantly altered in the present study. Thus, we conclude that, in the penis of rats with early stage DM, the protection against apoptosis has decreased but can be improved by ET antagonism.     

Phenotypic Modulation of Corpus Cavernosum Smooth Muscle Cells in a Rat Model of Cavernous Neurectomy

Background

Patients undergoing radical prostatectomy (RP) are at high risk for erectile dysfunction (ED) due to potential cavernous nerve (CN) damage during surgery. Penile hypoxia after RP is thought to significantly contribute to ED pathogenesis.

Aim

We previously showed that corpora cavernosum smooth muscle cells (CCSMCs) undergo phenotypic modulation under hypoxic conditions in vitro. Here, we studied such changes in an in vivo post-RP ED model by investigating CCSMCs in bilateral cavernous neurectomy (BCN) rats.

Methods

Sprague-Dawley rats underwent sham (n = 12) or BCN (n = 12) surgery. After 12 weeks, they were injected with apomorphine to determine erectile function. The penile tissues were harvested and assessed for fibrosis using Masson trichrome staining and for molecular markers of phenotypic modulation using immunohistochemistry and western blotting. CCSMC morphological structure was evaluated by hematoxylin-eosin (H&E) staining and transmission electron microscopy (TEM).

Results

Erectile function was significantly lower in BCN rats than in sham rats. BCN increased hypoxia-inducible factor-1α and collagen protein expression in corpora cavernous tissue. H&E staining and TEM showed that CCSMCs in BCN rats underwent hypertrophy and showed rough endoplasmic reticulum formation. The expression of CCSMC phenotypic markers, such as smooth muscle α-actin, smooth muscle myosin heavy chain, and desmin, was markedly lower, whereas vimentin protein expression was significantly higher in BCN rats than in control rats.

Conclusions

CCSMCs undergo phenotype modulation in rats with cavernous neurectomy. The results have unveiled physiological transformations that occur at the cellular and molecular levels and have helped characterize CN injury–induced ED.     

阴茎勃起及勃起功能障碍的研究进展

勃起功能障碍的基础研究须近十年来取得了较大进展,一氧化氮－cGMP(NO-cGMP)通路的发现使得阴茎平滑肌松驰的机制进一步阐明。一氧化氮合酶（NOS）、磷酸二酯酶（PDEs）的研究为勃起功能障碍的临床治疗提供了坚实的基础,进而促使了万艾可的问世。目前,勃起功能障碍的基因治疗停留在实验室阶段,但随着分子生物学的深入研究,转基因疗法可能成为临床上治疗勃起功能障碍的有效方法之一。     

Distribution and origin of neuropeptide Y-immunoreactive fibers in the penis of the rat

Summary The present study investigated the distribution of neuropeptide Y-immunoreactive fibers to the penis of the rat. In the corpora cavernosa penis, a dense plexus of fibers was asociated with arteries, intrinsic cavernosal muscle, and veins including the deep dorsal vein. In the corpus spongiosum, immunoreactive fibers were present around vascular smooth muscle and at the periphery of the acini of the paraurethral glands. Immunohistochemistry of penile neurons identified by retrograde tracer injection into the penis indicates that about 5% of the penile neurons in the pelvic plexus contained the neuropeptide while larger percentages of penile neurons in the sympathetic chains were immunoreactive for neuropeptide Y. Chemical and surgical sympathectomy greatly reduced the neuropeptide Y- and catecholamine-containing fibers in the erectile tissue but had no clear effect on the neuropeptide Y fibers around the paraurethral glands; a tissue that is not innervated by adrenergic fibers. It is concluded that (1) the widespread distribution of neuropeptide Y indicates that it may function in the control of penile blood flow, (2) with the possible exception of the paraurethral glands, the sympathetic chain is the most likely source of neuropeptide Y fibers in both erectile bodies of the penis, and (3) this peptide may play a role in the secretory functions of the paraurethral glands.     

Localisation and measurement of VIP in the genitourinary system of man and animals

VIP is present in the genitourinary system of man and animals. In man the highest concentrations are found in the penis, the uterus and vagina and in the urinary bladder. VIP nerves heavily innervate the erectile tissue of the male external genitalia, the uterine smooth muscle and blood vessels, the seromucous glands of the cervix, and the lamina propria and vaginal epithelium. In the urinary bladder, VIP nerves are located beneath the transitional epithelium, in the lamina propria and in the smooth muscle. Other areas well innervated by VIP nerves include the prostate, seminal vesicles and vasa deferentia. Chemical (phenol- and 6-OHDA) or surgical (hypogastric or pelvic nerve section) extrinsic denervation fail to deplete the genitourinary system of its VIP content, supporting the view that VIP-containing nerves originate from local ganglion cells. Indeed, neuronal cell bodies containing VIP are seen in the paracervical ganglia of the female genitalia, the para- or intramural bladder ganglia and scattered through the base of the cavernosum body, the neck of the bladder and the prostate. The finding of elevated levels of VIP in the local circulation after induced penile erection in man and mammals and the ability of VIP to relax the detrusor muscle of the bladder suggests that the peptide may be involved in penile erection and bladder relaxation, as does the marked VIP depletion in the penis or bladder in patients suffering from diabetic impotence or bladder instability.     

Does diabetes mellitus affect the progress of tolerance to isosorbide dinitrate (ISDN) in corporal tissue?

For erection to take place, the penile arteries and sinusoids have to dilate, thereby increasing the blood flow into the penis. There is increasing evidence that release of l-arginine derived nitric oxide (NO) from nonadrenergic-noncholinergic (NANC) nerves and from the sinusoidal endothelium is a major event in penile smooth muscle relaxation and promotes the endogenous formation of cyclic guanosine monophosphate (cGMP). Nitrovasodilators can be attributed to the activation of soluble guanylate cyclase, resulting in an increase in intracellular level of cyclic guanosine monophosphate, but prolonged exposure to high levels of nitroglycerine and other organic nitroesters induces tolerance against the cardiovascular effect. In this study, the aim was to determine the effect of diabetes on the corporal smooth muscle relaxant effect of ISDN and the effect of diabetes on the process of tolerance to the drug. For this purpose, alloxan-induced diabetic rabbits were used to form diabetes group. The responses of the corpus cavernous strips obtained from control and alloxan-induced diabetic rabbit were studied in organ chamber. In conclusion, prolonged in vitro exposure of corpus cavernosum strips obtained from control and diabetic groups to high concentrations of ISDN caused significant desensitization to the relaxant effect the drug. So, prolonged exposure of corporal tissue to the agents like nitroglycerine, used for treatment of impotence, may render ineffective the therapy in diabetic erectile impotence. However, intolerance to nitric oxide provides a rationale for the concept of using nitro oxide agents (like SNP) in the treatment of diabetic erectile dysfunction.     

Effects of Intracavernous Injection of Adipose-Derived Stem Cells on Cavernous Nerve Regeneration in a Rat Model

The aim of this study was to investigate effects of intracavernous injection of adipose-derived stem cells (ADSCs) on cavernous nerve (CN) regeneration and functional status in a nerve-crush rat model. Thirty Sprague–Dawley male rats were randomly divided into three equal groups: one group underwent sham operation, while two groups underwent bilateral CN crush. Crush-injury group was treated at the time of injury with intracavernous injection of ADSCs, or injured control group with no further intervention. Erectile function was assessed by CN electrostimulation after 3 months. Penile tissue and crushed nerves were collected for histology. Three months after surgery, in the group that underwent bilateral nerve crushing with no further intervention, the functional evaluation showed a lower mean maximal intracavernous pressure (ICP) and maximal ICP per mean arterial pressure (MAP) with CN stimulation than those in the sham group. In the group with an immediate intracavernous injection of ADSCs, the mean maximal ICP and maximal ICP/MAP were significantly higher than those in the injured control group. Histologically, the group with the intracavernous injection of ADSCs had more myelinated axons of CNs and more NADPH-diaphorase-positive nerve fibers than the injured control group but fewer than the sham group. Intracavernous injection of ADSCs treatment had beneficial effects on the smooth muscle/collagen ratio in the corpus cavernosum. These results show that the intracavernous injection of ADSCs to the site of CN-crush injury facilitates nerve regeneration and recovery of erectile function. Our research indicates that penile injection of ADSCs can improve recovery of erectile function in a rat model of neurogenic ED.     

Early treatment with fumagillin, an inhibitor of methionine aminopeptidase-2, prevents Pulmonary Hypertension in monocrotaline-injured rats

Pulmonary Hypertension (PH) is a pathophysiologic condition characterized by hypoxemia and right ventricular strain. Proliferation of fibroblasts, smooth muscle cells, and endothelial cells is central to the pathology of PH in animal models and in humans. Methionine aminopeptidase-2 (MetAP2) regulates proliferation in a variety of cell types including endothelial cells, smooth muscle cells, and fibroblasts. MetAP2 is inhibited irreversibly by the angiogenesis inhibitor fumagillin. We have previously found that inhibition of MetAP2 with fumagillin in bleomycin-injured mice decreased pulmonary fibrosis by selectively decreasing the proliferation of lung myofibroblasts. In this study, we investigated the role of fumagillin as a potential therapy in experimental PH. In vivo, treatment of rats with fumagillin early after monocrotaline injury prevented PH and right ventricular remodeling by decreasing the thickness of the medial layer of the pulmonary arteries. Treatment with fumagillin beginning two weeks after monocrotaline injury did not prevent PH but was associated with decreased right ventricular mass and decreased cardiomyocyte hypertrophy, suggesting a direct effect of fumagillin on right ventricular remodeling. Incubation of rat pulmonary artery smooth muscle cells (RPASMC) with fumagillin and MetAP2-targeting siRNA inhibited proliferation of RPASMC in vitro. Platelet-derived growth factor, a growth factor that is important in the pathogenesis of PH and stimulates proliferation of fibroblasts and smooth muscle cells, strongly increased expression of MetP2. By immunohistochemistry, we found that MetAP2 was expressed in the lesions of human pulmonary arterial hypertension. We propose that fumagillin may be an effective adjunctive therapy for treating PH in patients.     

Epac1 is upregulated during neointima formation and promotes vascular smooth muscle cell migration

Vascular remodeling after mechanoinjury largely depends on the migration of smooth muscle cells, an initial key step to wound healing. However, the role of the second messenger system, in particular, the cAMP signal, in regulating such remodeling remains controversial. Exchange protein activated by cAMP (Epac) has been identified as a new target molecule of the cAMP signal, which is independent from PKA. We thus examined whether Epac plays a distinct role from PKA in vascular remodeling. To examine the role of Epac and PKA in migration, we used primary culture smooth muscle cells from both the fetal and adult rat aorta. A cAMP analog selective to PKA, 8-(4-parachlorophenylthio)-cAMP (pCPT-cAMP), decreased cell migration, whereas an Epac-selective analog, 8-pCPT-2'-O-Me-cAMP, enhanced migration. Adenovirus-mediated gene transfer of PKA decreased cell migration, whereas that of Epac1 significantly enhanced cell migration. Striking morphological differences were observed between pCPT-cAMP- and 8-pCPT-2'-O-Me-cAMP-treated aortic smooth muscle cells. Furthermore, overexpression of Epac1 enhanced the development of neointimal formation in fetal rat aortic tissues in organ culture. When the mouse femoral artery was injured mechanically in vivo, we found that the expression of Epac1 was upregulated in vascular smooth muscle cells, whereas that of PKA was downregulated with the progress of neointimal thickening. Our findings suggest that Epac1, in opposition to PKA, increases vascular smooth muscle cell migration. Epac may thus play an important role in advancing vascular remodeling and restenosis upon vascular injury.