Cavernous Angiogenesis Research Articles

Vasohibin-1 rescues erectile function through up-regulation of angiogenic factors in the diabetic mice

Neovascularization of the erectile tissue emerges as a beneficial curative approach to treat erectile dysfunction (ED). Here we for the first time report the unexpected role of vasohibin-1 (VASH1), mainly known as an anti-angiogenic factor, in restoring erectile function in diabetic mice. A diabetic patient has lower cavernous VASH1 expression than in the potent man. VASH1 was mainly expressed in endothelial cells. There were significant decreases in cavernous endothelial cell and pericyte contents in VASH1 knockout mice compared with those in wild-type mice, which resulted in impairments in erectile function. Intracavernous injection of VASH1 protein successfully restored erectile function in the diabetic mice (~ 90% of control values). VASH1 protein reinstated endothelial cells, pericytes, and endothelial cell–cell junction proteins and induced phosphorylation of eNOS (Ser1177) in the diabetic mice. The induction of angiogenic factors, such as angiopoietin-1 and vascular endothelial growth factor, is responsible for cavernous angiogenesis and the restoration of erectile function mediated by VASH1. Altogether, these findings suggest that VASH1 is proangiogenic in diabetic penis and is a new potential target for diabetic ED.

Intracavernous delivery of Dickkopf3 gene or peptide rescues erectile function through enhanced cavernous angiogenesis in the diabetic mouse.

Severe peripheral angiopathy in patients with diabetes is a major contributing factor for low response rate to phosphodiesterase-5 inhibitors. To examine whether and how Dickkopf3 (DKK3), a secreted modulator of the Wnt pathway that known to be involved in endothelial cell repair and vascular progenitor cell migration, restores erectile function in diabetic mice. Eight-week-old C57BL/6 mice received intraperitoneal injections of streptozotocin (50mg/kg for 5days). Eight weeks after the diabetes was induced, the efficacy of DKK3 was determined by three independent experiments: experiment 1 (DKK3 peptide [5μg in 20μL PBS]); experiment 2 (DKK3 plasmid DNA with electroporation [10, 40, or 100μg in 20μL PBS, respectively]); and experiment 3 (DKK3 adenovirus [1×107 , 1×108 , 1×109 virus particles per 20μL, respectively]). Erectile function was measured by electrical stimulation of the cavernous nerve one week (for peptide) or two weeks (for genes) after treatment. The angiogenic activity of DKK3 was determined in diabetic penis in vivo and in primary cultured mouse cavernous endothelial cells (MCECs) in vitro. The cavernous expression of DKK3 protein was significantly lower in the diabetic mice than in controls. DKK3 peptide or adenovirus significantly improved erectile function in diabetic mice (70% of the control values). DKK3 adenovirus profoundly restored cavernous endothelial cell and pericyte contents and increased endothelial junction proteins in diabetic mice in vivo. DKK3 peptide induced upregulation of angiogenic factors (angiopoietin-1, vascular endothelial growth factor, and basic fibroblast growth factor) and accelerated tube formation in MCECs cultivated under the high-glucose condition in vitro. DKK3 restored cavernous vascular integrity and improved erectile function in diabetic mice. Therapeutic cavernous angiogenesis by the use of DKK3 will be a promising therapeutic strategy to treat diabetic erectile dysfunction.

Angiopoietin-Tie Signaling Pathway in Endothelial Cells: A Computational Model.

SummaryThe angiopoietin-Tie signaling pathway is an important vascular signaling pathway involved in angiogenesis, vascular stability, and quiescence. Dysregulation in the pathway is linked to the impairments in vascular function associated with many diseases, including cancer, ocular diseases, systemic inflammation, and cardiovascular diseases. The present study uses a computational signaling pathway model validated against experimental data to quantitatively study various mechanistic aspects of the angiopoietin-Tie signaling pathway, including receptor activation, trafficking, turnover, and molecular mechanisms of its regulation. The model provides mechanistic insights into the controversial role of Ang2 and its regulators vascular endothelial protein tyrosine phosphatase (VE-PTP) and Tie1 and predicts synergistic effects of inhibition of VE-PTP, Tie1, and Tie2 cleavage on enhancing the vascular protective actions of Tie2.

Intracavernosal Adeno-Associated Virus-Mediated S100A1 Gene Transfer Enhances Erectile Function in Diabetic Rats by Promoting Cavernous Angiogenesis via VEGF-A/VEGFR2 Signaling

IntroductionNovel therapeutic targets for diabetes-induced erectile dysfunction (DED) are urgently needed. Previous studies have proved that S100A1, a small Ca2+-binding protein, is a pluripotent regulator of cardiovascular pathophysiology. Its absence is associated with endothelial dysfunction, the central event linking cardiovascular changes in diabetes. However, the role of S100A1 in DED remains unknown. AimTo explore the effect and underlying mechanisms of S100A1 in restoring erectile function in type I diabetic rat model. MethodsDiabetes was induced by intraperitoneal injection of streptozotocin and then screened by apomorphine (APO) to confirm erectile dysfunction. Rats that met the criteria of penile erection were marked as APO-positive; otherwise, the result was APO-negative. In experiment 1, S100A1 gene expression alterations in the corpus cavernosum in moderate and established stages of DED were analyzed. In experiment 2, S100A1 and control GFP gene were delivered into the corpus cavernosum in APO-negative rats by adeno-associated virus (AAV) serotype 9. Erectile function was assessed at 4 weeks after gene therapy. Main Outcome MeasuresErectile response, histologic and molecular alterations. ResultsS100A1 protein was localized to the area surrounding the cavernosal sinusoids in the penis, and it was gradually downregulated synchronized with the progression of DED. Compared with an injection of AAV-GFP, a single injection of AAV-S100A1 significantly restored erectile function in diabetic rats. S100A1 overexpression significantly upregulated the expression of endogenous VEGF-A, promoted VEGFR2 internalization, and subsequently triggered the protein kinase B–endothelial nitric oxide synthase pathway in diabetic erectile tissues. Marked increases in nitric oxide and endothelial content were noted in AAV-S100A1-treated diabetic rats. Clinical ImplicationsLocal S100A1 overexpression may be an alternative therapy for DED and should be further investigated by future clinical studies. Strength & LimitationsThis is the first study demonstrating the angiogenic role of S100A1 in DED, but does not preclude the contribution of the effects of S100A1 in other tissues such as the neuronal tissue on the functional effects observed in erectile responses. ConclusionThe decreased expression of S100A1 during hyperglycemia might be important in the development of erectile dysfunction. S100A1 may play a potential role in restoring erectile function in rats with DED through modulating cavernous angiogenesis.Yu Z, Zhang Y, Tang Z, et al. Intracavernosal Adeno-Associated Virus-Mediated S100A1 Gene Transfer Enhances Erectile Function in Diabetic Rats by Promoting Cavernous Angiogenesis via VEGF-A/VEGFR2 Signaling. J Sex Med 2019;16:1344–1354.

AB035. Vasohibin-1 as a potential therapeutic target for erectile dysfunction

Induction of neovascularization is a promising strategy to treat erectile dysfunction (ED). Here we for the first time report the unexpected role of vasohibin-1, which is previously defined as an anti-angiogenic factor, in the restoration of erectile function by enhancing cavernous (penile) angiogenesis in an animal model of diabetic ED. The cavernous expression of vasohibin-1 was down-regulated in diabetic mice and in patients with diabetic ED; vasohibin-1 was mainly expressed in endothelial cells. Vasohibin-1 knockout mice revealed decrease in cavernous endothelial and pericyte content compared with wild type mice, which resulted in deterioration of erectile function. Local delivery of vasohibin-1 peptide into the corpus cavernosum of diabetic mice induced significant restoration of erectile function, which reached up to 85% of control values at the concentration of 4 µg/20 µL. Vasohibin-1 significantly increased cavernous endothelial cell content and induced eNOS phosphorylation (Ser1177). Vasohibin-1 decreased extravasation of oxidized-LDL by restoring pericytes and endothelial cell-cell junction proteins in the diabetic mice. By using proteome profiler array kit, we found that the induction of angiogenic factors, including angiopoietin-1, vascular endothelial growth factor, and basic fibroblasts growth factor, mainly in fibroblasts is a major molecular mechanism responsible for vasohibin-1-mediated cavernous angiogenesis and subsequent restoration of erectile function. Our findings suggest that vasohibin-1 is proangiogenic in diabetic penis and is a promising therapeutic target for ED from vascular causes.

AB084. intracavernous administration of embryonic stem cell-derived exosome restores erectile function through penile neurovascular regeneration in the streptozotocin-induced diabetic mouse

BackgroundExosome contains a variety of protein, mRNA, and miRNA and is known to play an important role in intercellular communication as a bio-nanoparticle with a diameter of 40 to 100 nm. Recent studies have demonstrated the therapeutic potential of exosome in a variety of animal models for cardiovascular diseases and neuropathies. The aim of this study was to investigate effectiveness of embryonic stem cell (ESC)-derived exosome in restoring erectile function in diabetic mice.MethodsDiabetes was induced by intraperitoneal injection of streptozotocin into 8-week-old C57BL/6 male mice. At 8 weeks after the induction of diabetes, the animals were distributed into 3 groups: control nondiabetic mice and diabetic mice receiving two successive intracavernous injections of phosphate buffered saline (PBS) (days −3 and 0; 20 µL) or ESC-derived exosome (days −3 and 0; 1 µg in 20 µL of PBS). Two weeks after treatment, we measured erectile function by electrical stimulation of the cavernous nerve. The penis was harvested and stained with antibodies to PECAM-1, smooth muscle α-actin, NG2, and βIII tubulin. We also determined angiogenic potential of ESC-derived exosome in an ex vivo aortic ring assay and in primary cultured mouse cavernous endothelial cell (MCEC) and pericyte (MCP) mono-culture or co-culture system in vitro.ResultsIntracavernous injections of ESC-derived exosome significantly improved erectile function in diabetic mice, which reached up to 90% of control values. ESC-derived exosome induced significant restoration of cavernous contents of endothelial cells, smooth muscle cells, pericytes, and neuronal cells in diabetic condition. Moreover, ESC-derived exosome promoted microvascular sprouting from aortic ring and accelerated tube formation in primary cultured MCEC and MCP mono-culture or co-culture system in vitro.ConclusionsESC-derived exosome successfully restored erectile function through enhanced cavernous angiogenesis and neural regeneration in diabetic mice. Further studies are needed to clarify mechanism by which ESC-derived exosome induces neurovascular repair.

275 Inhibition of proNGF pathway restores erectile function by enhancing cavernous angiogenesis and neural regeneration in streptozotocin-induced diabetic mouse

Patients with diabetic erectile dysfunction (ED) usually respond poorly to oral PDE5 inhibitors due to a lack of bioavailable nitric oxide from severe endothelial and neural dysfunction. ProNGF and its receptor p75NTR are known to be up-regulated in diabetic condition and to induce endothelial cell apoptosis and neuronal degeneration. The aim of this study was to investigate effectiveness of anti-proNGF neutralizing antibody (proNGF-Ab) in restoring erectile function in streptozotocin-induced diabetic mouse. Diabetes was induced by intraperitoneal injection of streptozotocin (50 mg/kg) into 8-week-old C57BL/6 male mice for 5 consecutive days. At 8 weeks after the induction of diabetes, the animals were distributed into 3 groups: controls, streptozotocin-induced diabetic mice receiving repeated intracavernous injections of PBS (days -3 and 0; 20 μL) or proNGF-Ab (days -3 and 0; 20 μg in 20 μL of saline). We measured erectile function by electrical stimulation of the cavernous nerve at 2 weeks after treatment. The penis was then harvested for histologic and biochemical studies.

Current Literature Review.

Current Literature Review.

MP89-01 VASOHIBIN-1 IS A NOVEL THERAPEUTIC TARGET FOR DIABETIC ERECTILE DYSFUNCTION IN MICE

You have accessJournal of UrologySexual Function/Dysfunction: Basic Research & Pathophysiology II1 Apr 2016MP89-01 VASOHIBIN-1 IS A NOVEL THERAPEUTIC TARGET FOR DIABETIC ERECTILE DYSFUNCTION IN MICE Woo Jean Kim, Kang-Moon Song, Jin-Mi Park, Anita Limanjaya, Kalyan Ghatak, Nguyen Nhat Minh, Soo-Hwan Park, Won-Hee Park, Jiyeon Ock, Guo Nan Yin, Ji-Kan Ryu, Jun-Kyu Suh, and In Rae Cho Woo Jean KimWoo Jean Kim More articles by this author , Kang-Moon SongKang-Moon Song More articles by this author , Jin-Mi ParkJin-Mi Park More articles by this author , Anita LimanjayaAnita Limanjaya More articles by this author , Kalyan GhatakKalyan Ghatak More articles by this author , Nguyen Nhat MinhNguyen Nhat Minh More articles by this author , Soo-Hwan ParkSoo-Hwan Park More articles by this author , Won-Hee ParkWon-Hee Park More articles by this author , Jiyeon OckJiyeon Ock More articles by this author , Guo Nan YinGuo Nan Yin More articles by this author , Ji-Kan RyuJi-Kan Ryu More articles by this author , Jun-Kyu SuhJun-Kyu Suh More articles by this author , and In Rae ChoIn Rae Cho More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.2462AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Induction of neovascularization is a promising strategy to treat diabetic erectile dysfunction (ED). Here we for the first time report the unexpected role of vasohibin-1, which is previously defined as an anti-angiogenic factor, in the restoration of erectile function by enhancing cavernous angiogenesis in an animal model of diabetic ED. METHODS Diabetes was induced by intraperitoneal injection of streptozotocin into 8-week-old C57BL/6 male mice. At 8 weeks after the induction of diabetes, the animals were divided into 4 groups: controls, streptozotocin-induced diabetic mice receiving repeated intracavernous injections of PBS (days -3 and 0; 20 μL), or vasohibin-1 protein (days -3 and 0; 1 μg and 4 μg in 20 μL of PBS, respectively). Two weeks after treatment, we measured erectile function by electrical stimulation of the cavernous nerve. The penis was harvested for histologic and biochemical studies. RESULTS The cavernous expression of vasohibin-1 was down-regulated in diabetic mice and in patients with diabetic ED; vasohibin-1 was mainly expressed in endothelial cells. Vasohibin-1 knockout mice revealed decrease in cavernous endothelial and pericyte content compared with wild type mice, which resulted in deterioration of erectile function. Local delivery of vasohibin-1 into the corpus cavernosum of diabetic mice induced significant restoration of erectile function, which reached up to 85% of control values at the concentration of 4 μg/20 μL. Vasohibin-1 significantly increased cavernous endothelial cell and induced eNOS phosphorylation (Ser1177). Vasohibin-1 decreased extravasation of oxidized-LDL by restoring pericyte content and endothelial cell-cell junction proteins in the diabetic mice. By using proteome profiler array kit, we found that the induction of angiogenic factors, including angiopoietin-1, vascular endothelial growth factor, and basic fibroblasts growth factor, mainly in fibroblasts is a major molecular mechanism responsible for vasohibin-1-mediated cavernous angiogenesis and subsequent restoration of erectile function. CONCLUSIONS Our findings suggest that vasohibin-1 is proangiogenic in diabetic penis and is a promising therapeutic target for ED from vascular causes. © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e1137 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Woo Jean Kim More articles by this author Kang-Moon Song More articles by this author Jin-Mi Park More articles by this author Anita Limanjaya More articles by this author Kalyan Ghatak More articles by this author Nguyen Nhat Minh More articles by this author Soo-Hwan Park More articles by this author Won-Hee Park More articles by this author Jiyeon Ock More articles by this author Guo Nan Yin More articles by this author Ji-Kan Ryu More articles by this author Jun-Kyu Suh More articles by this author In Rae Cho More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

This Month in Investigative Urology

No AccessJournal of UrologyThis Month in Investigative Urology1 Jun 2016This Month in Investigative Urology Karl-Erik Andersson Karl-Erik AnderssonKarl-Erik Andersson More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.03.042AboutFull TextPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail "This Month in Investigative Urology." The Journal of Urology, 195(6), pp. 1637–1638 References 1 : Integrative pathway analysis of metabolic signature in bladder cancer: a linkage to The Cancer Genome Atlas project and prediction of survival. J Urol2016; 195: 1911. Link, Google Scholar 2 : Sac-1004, a pseudo-sugar derivative of cholesterol, restores erectile function through reconstruction of nonleaky and functional cavernous angiogenesis in the streptozotocin induced diabetic mouse. J Urol2016; 195: 1936. Link, Google Scholar 3 : Liposome based intravesical therapy targeting nerve growth factor ameliorates bladder hypersensitivity in rats with experimental colitis. J Urol2016; 195: 1920. Link, Google Scholar © 2016 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 195Issue 6June 2016Page: 1637-1638 Advertisement Copyright & Permissions© 2016 by American Urological Association Education and Research, Inc.MetricsAuthor Information Karl-Erik Andersson More articles by this author Expand All Advertisement PDF downloadLoading ...

Sac-1004, a Pseudo-Sugar Derivative of Cholesterol, Restores Erectile Function through Reconstruction of Nonleaky and Functional Cavernous Angiogenesis in the Streptozotocin Induced Diabetic Mouse

We examined whether and how Sac-1004, a vascular leakage blocker, would restore erectile function in an animal model of diabetic erectile dysfunction. Eight-week-old C57BL/6J mice were used. Diabetes was induced by intraperitoneal injection of streptozotocin. Eight weeks after diabetes induction the animals were divided into 6 groups, including controls, diabetic mice that received repeat intracavernous injections of phosphate buffered saline (20 μl) on days -3 and 0, and diabetic mice that received repeat intracavernous injections of Sac-1004 on days -3 and 0 (1, 2, 5 and 10 μg, respectively, in 20 μl phosphate buffered saline). One week after injection erectile function was measured by cavernous nerve stimulation. The penis was then harvested for histological examinations and Western blot analysis. Local delivery of Sac-1004 in the corpus cavernosum restored erectile function in diabetic mice. The highest erectile response was noted at a dose of 5μg with a response comparable to that in the control group. Sac-1004 significantly increased cavernous endothelial and smooth muscle contents, and induced endothelial nitric oxide synthase phosphorylation (Ser1177). Sac-1004 decreased extravasation of oxidized low density lipoprotein by restoring endothelial cell-cell junction proteins and pericyte content. Sac-1004 also promoted tube formation in primary cultured mouse cavernous endothelial cells invitro. Sac-1004 mediated cavernous angiogenesis and erectile function recovery was abolished by inhibiting angiopoietin-1-Tie2 signaling with soluble Tie2 antibody. With the effects of angiogenesis and antipermeability Sac-1004 reestablishes structural and functional cavernous sinusoids. This is highly promising for future treatment of erectile dysfunction from vascular causes.

Designed angiopoietin-1 variant, COMP-angiopoietin-1, rescues erectile function through healthy cavernous angiogenesis in a hypercholesterolemic mouse.

Despite the advent of oral phosphodiesterase-5 inhibitors, curative treatment for erectile dysfunction (ED) remains unavailable. Recently, the link between ED and cardiovascular disease was unveiled and the main etiology of ED was found to be vasculogenic. Therefore, neovascularization is a promising strategy for curing ED. Angiopoietin-1 (Ang1) is an angiogenic growth factor that promotes the generation of stable and functional vasculature. Here, we demonstrate that local delivery of the soluble, stable, and potent Ang1 variant, COMP-Ang1 gene or protein, into the penises of hypercholesterolemic mice increases cavernous angiogenesis, eNOS phosphorylation, and cGMP expression, resulting in full recovery of erectile function and cavernous blood flow up to 8 weeks after treatment. COMP-Ang1-induced promotion of cavernous angiogenesis and erectile function was abolished in Nos3-/- mice and in the presence of the NOS inhibitor, L-NAME. COMP-Ang1 also restored the integrity of endothelial cell-cell junction by down-regulating the expression of histone deacetylase 2 in the penis of hypercholesterolemic mice and in primary cultured mouse cavernous endothelial cells. These findings constitute a new paradigm toward curative treatment of both cavernous angiopathy and ED.

Intracavernous delivery of stromal vascular fraction restores erectile function through production of angiogenic factors in a mouse model of cavernous nerve injury.

Erectile dysfunction (ED) is a major complication of radical prostatectomy. Men with radical prostatectomy-induced ED respond less positively to oral phosphodiesterase-5 inhibitors. The study aims to examine whether and how stromal vascular fraction (SVF) restores erectile function in mice with cavernous nerve injury (CNI). Twelve-week-old male C57BL/6J mice were used and the animals were distributed into five groups: sham operation group and CNI group receiving a single intracavernous injection of phosphate-buffered saline (PBS) or SVF (1 × 10(4) , 1 × 10(5) , or 3 × 10(5) cells/20 μL, respectively). SVF was isolated from epididymal adipose tissues of green fluorescence protein transgenic mice. Two weeks after injection, erectile function was measured by cavernous nerve stimulation. The penis was stained with antibodies to platelet/endothelial cell adhesion molecule-1, phosphohistone H3, and phosphorylated endothelial nitric oxide synthase (phospho-eNOS). We also performed Western blot for angiopoietin-1 (Ang-1), vascular endothelial growth factor-A, hepatocyte growth factor, phospho-eNOS, and eNOS in the corpus cavernosum tissue. Local delivery of SVF restored erectile function in a dose-dependent manner in CNI mice. The highest erectile response was noted at a dose of 3 × 10(5) cells, for which the response was comparable with that in the sham operation group. Local delivery of SVF significantly increased the expression of angiogenic factor proteins and induced cavernous endothelial cell proliferation and eNOS phosphorylation compared with that in the PBS-treated CNI group. SVF-induced promotion of cavernous angiogenesis and erectile function was diminished in the presence of soluble antibody to Tie2, a receptor tyrosine kinase of Ang-1. Secretion of angiogenic factors from SVF is an important mechanism by which SVF induces cavernous endothelial regeneration and restores erectile function. These findings suggest that cavernous endothelial regeneration by using SVF may represent a promising treatment strategy for radical prostatectomy-induced ED.

COMP-Angiopoietin-1 Promotes Cavernous Angiogenesis in a Type 2 Diabetic Rat Model

Cartilage oligomeric matrix protein-angiopoietin-1 (COMP-Ang1) is an angiogenic factor for vascular angiogenesis. The aim was to investigate the effect of an intracavernosal injection of COMP-Ang1 on cavernosal angiogenesis in a diabetic rat model. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats made up the experimental group (1 yr old) and Long-Evans Tokushima Otsuka (LETO) rats made up the control group. The experimental group was divided into vehicle only, 10 µg COMP-Ang1, and 20 µg COMP-Ang1. COMP-Ang1 was injected into the corpus cavernosum of the penis. After 4 weeks, the penile tissues of the rats were obtained for immunohistochemistry and Western blot analysis. The immunoreactivity of PECAM-1 and VEGF was increased in the COMP-Ang1 group compared with the vehicle only group. Moreover, the expression of PECAM-1 and VEGF was notably augmented in the 20 µg Comp Ang-1 group. In the immunoblotting study, the expression of PECAM-1 and VEGF protein was significantly less in the OLEFT rats than in the control LETO rats. However, this expression was restored to control level after intracavernosal injection of COMP-Ang1. These results show that an intracavernosal injection of COMP-Ang1 enhances cavernous angiogenesis by structurally reinforcing the cavernosal endothelium.

Intracavernous Delivery of Freshly Isolated Stromal Vascular Fraction Rescues Erectile Function by Enhancing Endothelial Regeneration in the Streptozotocin‐Induced Diabetic Mouse

Men with diabetic erectile dysfunction (ED) often have severe endothelial dysfunction and respond poorly to oral phosphodiesterase-5 inhibitors. To examine whether and how freshly isolated stromal vascular fraction (SVF) promotes cavernous endothelial regeneration and restores erectile function in diabetic animals. Eight-week-old C57BL/6J mice were used. Diabetes was induced by intraperitoneal injection of streptozotocin. SVF was isolated from epididymal adipose tissues of green fluorescence protein transgenic mice. At 8 weeks after the induction of diabetes, the animals were divided into six groups: controls, diabetic mice, and diabetic mice treated with a single intracavernous injection of phosphate-buffered saline (PBS) or SVF (1 × 10(4) cells, 1 × 10(5) cells, or 2 × 10(5) cells/20 µL, respectively). Two weeks later, erectile function was measured by cavernous nerve stimulation. The penis was stained with antibodies to CD31, CD34, phosphohistone H3, phospho-endothelial nitric oxide synthase (eNOS), and vascular endothelial growth factor-A (VEGF-A). We also performed Western blot for phospho-eNOS and eNOS, and determined cyclic guanosine monophosphate (cGMP) concentration in the corpus cavernosum tissue. Significant improvement in erectile function was noted in diabetic mice treated with SVF at concentrations of 1 × 10(5) and 2 × 10(5) cells, which reached up to 82% of the control values. Local delivery of SVF significantly increased cavernous endothelial cell proliferation, eNOS phosphorylation, and cGMP expression compared with that in the untreated group and the PBS-treated diabetic group. Intracavernous injection of SVF increased cavernous VEGF-A expression and induced recruitment of CD34(+)CD31(-) progenitor cells. Some SVF underwent differentiation into cavernous endothelial cells. SVF-induced promotion of cavernous angiogenesis and erectile function was abolished in the presence of VEGF-Trap, a soluble VEGF-A neutralizing antibody. The results support the concept of cavernous endothelial regeneration by use of SVF as a curative therapy for diabetic ED.

Intracavernous Delivery of a Designed Angiopoietin-1 Variant Rescues Erectile Function by Enhancing Endothelial Regeneration in the Streptozotocin-Induced Diabetic Mouse

OBJECTIVEPatients with diabetic erectile dysfunction often have severe endothelial dysfunction and respond poorly to oral phosphodiesterase-5 inhibitors. We examined the effectiveness of the potent angiopoietin-1 (Ang1) variant, cartilage oligomeric matrix protein (COMP)-Ang1, in promoting cavernous endothelial regeneration and restoring erectile function in diabetic animals.RESEARCH DESIGN AND METHODSFour groups of mice were used: controls; streptozotocin (STZ)-induced diabetic mice; STZ-induced diabetic mice treated with repeated intracavernous injections of PBS; and STZ-induced diabetic mice treated with COMP-Ang1 protein (days −3 and 0). Two and 4 weeks after treatment, we measured erectile function by electrical stimulation of the cavernous nerve. The penis was harvested for histologic examinations, Western blot analysis, and cGMP quantification. We also performed a vascular permeability test.RESULTSLocal delivery of the COMP-Ang1 protein significantly increased cavernous endothelial proliferation, endothelial nitric oxide (NO) synthase (NOS) phosphorylation, and cGMP expression compared with that in the untreated or PBS-treated STZ-induced diabetic group. The changes in the group that received COMP-Ang1 restored erectile function up to 4 weeks after treatment. Endothelial protective effects, such as marked decreases in the expression of p47phox and inducible NOS, in the generation of superoxide anion and nitrotyrosine, and in the number of apoptotic cells in the corpus cavernosum tissue, were noted in COMP-Ang1–treated STZ-induced diabetic mice. An intracavernous injection of COMP-Ang1 completely restored endothelial cell-cell junction proteins and decreased cavernous endothelial permeability. COMP-Ang1–induced promotion of cavernous angiogenesis and erectile function was abolished by the NOS inhibitor, N-nitro-L-arginine methyl ester, but not by the NADPH oxidase inhibitor, apocynin.CONCLUSIONSThese findings support the concept of cavernous endothelial regeneration by use of the recombinant Ang1 protein as a curative therapy for diabetic erectile dysfunction.

Intracavernous Delivery of Synthetic Angiopoietin-1 Protein as a Novel Therapeutic Strategy for Erectile Dysfunction in the Type II Diabetic db/db Mouse

Patients with erectile dysfunction (ED) associated with type II diabetes often have impaired endothelial function and tend to respond poorly to oral phosphodiesterase type 5 inhibitors. Therefore, neovascularization is a promising strategy for curing diabetic ED. To determine the effectiveness of a soluble, stable, and potent angiopoietin-1 (Ang1) variant, cartilage oligomeric matrix protein (COMP)-Ang1, in promoting cavernous angiogenesis and erectile function in a mouse model of type II diabetic ED. Methods. Sixteen-week-old male db/db mice (in which obesity and type II diabetes are caused by a mutation in the leptin receptor) and control C57BL/6J mice were used and divided into four groups (N=14 per group): age-matched controls; db/db mice receiving two successive intracavernous injections of phosphate-buffered saline (PBS) (days -3 and 0; 20 µL); db/db mice receiving a single intracavernous injection of COMP-Ang1 protein (day 0; 5.8 µg/20 µL); and db/db mice receiving two successive intracavernous injections of COMP-Ang1 protein (days -3 and 0; 5.8 µg/20 µL). Two weeks later, erectile function was measured by electrical stimulation of the cavernous nerve. The penis was then harvested and stained with antibodies to platelet/endothelial cell adhesion molecule-1 (PECAM-1) (endothelial cell marker), phosphohistone H3 (PH3, a nuclear protein indicative of cell proliferation), phospho-endothelial nitric oxide synthase (eNOS), and eNOS. Penis specimens from a separate group of animals were used for cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) quantification. Local delivery of COMP-Ang1 protein significantly increased eNOS phosphorylation and cGMP and cAMP expression compared with that in the group treated with PBS. Repeated intracavernous injections of COMP-Ang1 protein completely restored erectile function and cavernous endothelial content through enhanced cavernous neoangiogenesis as evaluated by PECAM-1 and PH3 immunohistochemistry and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling assay, whereas a single injection of COMP-Ang1 protein elicited partial improvement. Cavernous neovascularization using recombinant Ang1 protein is a novel therapeutic strategy for the treatment of ED resulting from type II diabetes.

Characterization of VEGF and Angiopoietins Expression in Human Corpus Cavernosum during Aging

Erectile dysfunction (ED) is a highly prevalent and age-related disease, caused by endothelial dysfunction and impaired cavernous angiogenesis. However, cellular and molecular changes involved in erectile pathophysiology in aging male remain to be elucidated. To characterize the vascular organization, concomitantly with analysis of the expression of vascular endothelial growth factor (VEGF), Angiopoietin 1 (Ang1) and Angiopoietin 2 (Ang2) in young and aged human corpus cavernosum. Human penile fragments were removed from patients submitted to penile deviation surgery (11 cases; 58-70 years) and from potential organ donors (four cases; 18-28 years) without ED or risk factors for ED. Smooth muscle and connective tissue were assessed by Masson's trichrome staining and computer-assisted histomorphometry. Dual immunostaining for specific markers of endothelium (von Willebrand factor) and smooth muscle cell (alpha-actin), VEGF, Ang1 and Ang2 was assayed by fluorescence microscopy. Semi-quantification of expression of angiogenic factors was performed by Western blotting. Expression of VEGF and Angiopoietins in human corpus cavernosum, using a combination of histologic stainings, and molecular biology tools in order to achieve a better understanding of cavernosal tissue remodeling with aging. Aged human corpus cavernosum presented wider sinusoidal spaces, loss of muscle cell bundles, and increased connective tissue content. Ang1 was scarcely expressed in small clusters in smooth muscle cell cytoplasm with identical localization in both studied groups. VEGF expression was abundant in smooth muscle cell and its expression markedly decreased in aged tissue, contrasting with the expression of angiopoietins that increased in the aged corpus cavernosum. Immunoflourescent studies of cellular markers and growth factors help clarifying vascular organization and angiogenesis mechanisms in erectile tissue. Our findings demonstrate that the organization pattern of vascular endothelium and smooth muscle components of cavernosal tissue modifies during aging. Ang1 and Ang2 upregulation in human-aged penile tissue suggest a VEGF-independent vascular remodeling mechanism.

Gene therapy to promote new blood vessel formation in the ischemic heart: where do we stand?

Erectile dysfunction (ED) is a highly prevalent and age-related disease, caused by endothelial dysfunction and impaired cavernous angiogenesis. However, cellular and molecular changes involved in erectile pathophysiology in aging male remain to be elucidated.To characterize the vascular organization, concomitantly with analysis of the expression of vascular endothelial growth factor (VEGF), Angiopoietin 1 (Ang1) and Angiopoietin 2 (Ang2) in young and aged human corpus cavernosum.Human penile fragments were removed from patients submitted to penile deviation surgery (11 cases; 58–70 years) and from potential organ donors (four cases; 18–28 years) without ED or risk factors for ED. Smooth muscle and connective tissue were assessed by Masson's trichrome staining and computer-assisted histomorphometry. Dual immunostaining for specific markers of endothelium (von Willebrand factor) and smooth muscle cell (alpha-actin), VEGF, Ang1 and Ang2 was assayed by fluorescence microscopy. Semi-quantification of expression of angiogenic factors was performed by Western blotting.Expression of VEGF and Angiopoietins in human corpus cavernosum, using a combination of histologic stainings, and molecular biology tools in order to achieve a better understanding of cavernosal tissue remodeling with aging.Aged human corpus cavernosum presented wider sinusoidal spaces, loss of muscle cell bundles, and increased connective tissue content. Ang1 was scarcely expressed in small clusters in smooth muscle cell cytoplasm with identical localization in both studied groups. VEGF expression was abundant in smooth muscle cell and its expression markedly decreased in aged tissue, contrasting with the expression of angiopoietins that increased in the aged corpus cavernosum.Immunoflourescent studies of cellular markers and growth factors help clarifying vascular organization and angiogenesis mechanisms in erectile tissue. Our findings demonstrate that the organization pattern of vascular endothelium and smooth muscle components of cavernosal tissue modifies during aging. Ang1 and Ang2 upregulation in human-aged penile tissue suggest a VEGF-independent vascular remodeling mechanism. Tomada N, Tomada I, Cruz F, Vendeira P, and Neves D. Characterization of VEGF and angiopoietins expression in human corpus cavernosum during aging.

Combined Angiopoietin-1 and vascular endothelial growth factor gene transfer restores cavernous angiogenesis and erectile function in a rat model of hypercholesterolemia

Hypercholesterolemia-related endothelial cell dysfunction and decreased endothelium-derived nitric oxide formation may account for impaired angiogenesis and subsequent erectile dysfunction. Angiopoietin-1 (Ang1) is a critical angiogenic factor for vascular maturation and enhances vascular endothelial growth factor (VEGF)-induced angiogenesis in a complementary manner. We hypothesized that combined adenovirus-delivered human Ang1 (ad-Ang1) and VEGF165 (ad-VEGF165) gene transfer might promote angiogenesis cooperatively in a rat model of hypercholesterolemic erectile dysfunction and result in a recovery of erectile function. Ad-Ang1 and ad-VEGF165 were injected either alone or in combination into the corpus cavernosum of the penis. Combined gene transfer of both ad-Ang1 and ad-VEGF165 significantly increased cavernous angiogenesis, eNOS phosphorylation, and cGMP expression compared with that in the groups treated with either therapy alone. Erectile function, as evaluated by electrical stimulation of the cavernous nerve 2 and 8 weeks after treatment, was completely restored in the combined treatment group, whereas intracavernous injection of either ad-Ang1 or ad-VEGF165 alone elicited partial improvement. The results indicate that combined application of angiogenic factors may enhance cavernous angiogenesis cooperatively by reinforcing the endothelium both structurally and functionally, which results in an additive effect on erectile function in hypercholesterolemic rats.