A Simple and Nonenzymatic Method to Isolate Human Corpus Cavernosum Endothelial Cells and Pericytes for the Study of Erectile Dysfunction

Occurrence and molecular characterization of Enterocytozoon bieneusi among long-tailed macaques (Macaca fascicularis) in Hainan Province: High genetic diversity and zoonotic potential

Objective:Microsporidia have been rapidly emerging as pathogens in both immunocompromised and immunocompetent humans. Enterocytozoon bieneusi (E. bieneusi) is the most common microsporidial species found in human. E. bieneusi has also been found in a wide range of animals and is considered to be a potentially important zoonotic pathogen. The epidemiological and genetic characterization of E. bieneusi among long-tailed macaques [Macaca fascicularis (M. fascicularis) is not fully understood. Here, we conducted the first molecular epidemiological investigation of E. bieneusi among M. fascicularis in Hainan Province, the southernmost part of China.Methods:A total of 193 fecal specimens of M. fascicularis were collected from a breeding base housing non-human primates for experimental use in Hainan Province, China. E. bieneusi was identified and genotyped by nested PCR analysis of the internal transcribed spacer (ITS) region of the rRNA gene. Phylogenetic analysis was performed by constructing a neighboring-joining tree of the ITS gene sequences using MEGA6.Results:A total of 59 (30.6%) of the M. fascicularis were PCR-positive for E. bieneusi. All 59 samples were sequenced successfully and 16 ITS genotypes were identified. These included nine known genotypes: Type IV (n=19), D (n=11), CM1 (n=8), PigEBITS7 (n=4), Pongo2 (n=4), Peru 8 (n=3), Peru11 (n=1), WL21 (n=1) and CM2 (n=1). Additionally, seven novel genotypes named as HNM-I to HNM-VII (one each) were identified. Importantly, genotypes D, Type IV, Peru8, PigEBITS7, and Peru11, which were the predominant (38/59, 64.4%) genotypes identified among M. fascicularis in this study, are also well-known human-pathogenic genotypes. All the genotypes of E. bieneusi identified in this study, including the seven novel ones, belonged to zoonotic group 1.Conclusions:This is the first report of the identification of E. bieneusi in M. fascicularis in Hainan Province, China. The findings of numerous known human-pathogenic types and seven novel genotypes (HNM-I to HNM-VII) of E. bieneusi all belong to zoonotic group 1 indicate the possibility of transmission of this important pathogenic parasite between M. fascicularis and humans.

Combination of stromal vascular fraction and Ad-COMP-Ang1 gene therapy improves long-term therapeutic efficacy for diabetes-induced erectile dysfunction / 亚洲男科学杂志(英文版)

Men with diabetic erectile dysfunction (ED) respond poorly to the currently available oral phosphodiesterase-5 inhibitors. Therefore, functional therapies for diabetic ED are needed. Stromal vascular fraction (SVF) and the adenovirus-mediated cartilage oligomeric matrix angiopoietin-1 (Ad-COMP-Ang1) gene are known to play critical roles in penile erection. We previously reported that SVF and Ad-COMP-Ang1 have only a short-term effect in restoring erectile function. Further improvements to ED therapy are needed for long-lasting effects. In the present study, we aimed to test if the combination of SVF and Ad-COMP-Ang1 could extend the erection effect in diabetic ED. We found that the combination therapy showed a long-term effect in restoring erectile function through enhanced penile endothelial and neural cell regeneration. Combination therapy with SVF and Ad-COMP-Ang1 notably restored cavernous endothelial cell numbers, pericyte numbers, endothelial cell-cell junctions, decreased cavernous endothelial cell permeability, and promoted neural regeneration for at least 4 weeks in diabetic mice. In summary, this is an initial description of the long-term effect of combination therapy with SVF and Ad-COMP-Ang1 in restoring erectile function through a dual effect on endothelial and neural cell regeneration. Such combination therapy may have therapeutic potential for the treatment of diabetic ED.

Transforming Growth Factor-beta Type I Receptor Inhibitor Induces Functional and Morphologic Recovery in a Rat Model of Erectile Dysfunction and Cavernous Fibrosis / 대한남성과학회지

PURPOSE: To examine the effectiveness of small-molecule inhibitor of transforming growth factor-beta (TGF-beta) type I receptor, an activin receptor-like kinase 5 (ALK5), on erectile dysfunction (ED) in a rat model of cavernous fibrosis, in which fibrosis was induced by intracavernous injection of adenovirus expressing TGF-beta1 (Ad-TGF-beta1). MATERIALS AND METHODS: Four-month-old Sprague-Dawley rats were divided into four groups (n=10 per group): age-matched controls without treatment, age-matched controls receiving intracavernous injection of LacZ adenovirus, and cavernous fibrosis rats receiving an intracavernous injection of saline or ALK5 inhibitor (5 mg/kg). ALK5 inhibitor or saline was administered on day 5 after injection of Ad-TGF-beta1. On day 30, erectile function was assessed by electrical stimulation of the cavernous nerve and the penis was then harvested for histologic studies (n=6 per group) and for the measurement of the hydroxyproline level (n=4 per group). RESULTS: Ad-TGF-beta1-induced cavernous fibrosis rats treated with saline showed a significant decrease in cavernous smooth muscle and endothelial content, and an increase in collagen deposition, which resulted in profound deterioration of all erectile function parameters, such as the ratios of maximal intracavernous pressure (ICP), total ICP, and slope to mean arterial pressure. ALK5 inhibitor significantly restored erectile function in a rat model of cavernous fibrosis by increasing cavernous smooth muscle and endothelial content, and by blocking cavernous fibrosis. CONCLUSIONS: The results suggest that inhibition of the TGF-beta pathway is a promising therapeutic strategy for the treatment of ED related to cavernous fibrosis from various causes.

Establishment of a Cavernous Fibrosis Model in a Rat Using Adenovirus Expressing Transforming Growth Factor-beta1 / 대한남성과학회지

PURPOSE: Transforming growth factor-beta1 (TGF-beta1) has been implicated in cavernous fibrosis due to a variety of causes of erectile dysfunction (ED), such as diabetes mellitus and post-radical prostatectomy. To examine the role of the TGF-beta signaling pathway in cavernous fibrosis, we established a rat model of cavernous fibrosis by using adenovirus expressing TGF-beta1 (ad-TGF-beta1). MATERIALS AND METHODS: Four-month-old male Sprague-Dawley rats received intracavernous injection of ad-TGF-beta1 (1x10(8), 1x10(9), or 1x10(10) virus particles [vp] in 100 microliter of PBS) and the penis was harvested for histologic examination at 10, 20, or 30 days after injection (n=4 per group and per time point). Based on the initial findings, the animals were divided into three groups (n=6 per group): Group 1, age-matched control; Group 2, intracavernous injection of ad-LacZ (1x10(10) vp/100 microliter); and Group 3, intracavernous injection of ad-TGF-beta1 (1x10(10) vp/100 microliter). At 30 days after injection, erectile function was evaluated during electrical stimulation of the cavernous nerve. The penis was then harvested and stained with Masson's trichrome and antibody to smooth muscle alpha-actin. RESULTS: Masson's trichrome staining revealed that intracavernous delivery of ad-TGF-beta1 sufficiently induced cavernous fibrosis in a dose-dependent manner. The fibrotic scars persisted up to 30 days after injection at the highest dosage (1x10(10) vp/100 microliter), whereas no histologic evidence of cavernous fibrosis was found in the control rats or the ad-LacZ-injected rats. The rats receiving ad-TGF-beta1 showed a higher cavernous collagen content and less smooth muscle content than the control rats or ad-LacZ-injected rats. Erectile function was significantly decreased in rats receiving ad-TGF-beta1 compared with that in controls or rats receiving ad-LacZ. CONCLUSIONS: This model induced by ad-TGF-beta1 may play an important role in understanding the pathophysiologic mechanisms of cavernous fibrosis-associated TGF-beta signaling and the development of new therapeutics targeting this pathway.