Papaverine selectively inhibits human prostate cancer cell (PC-3) growth by inducing mitochondrial mediated apoptosis, cell cycle arrest and downregulation of NF-κB/PI3K/Akt signalling pathway.

PURPOSE: The main objective of the current research work was to investigate the antitumor effects of papaverine in PC-3 human prostate cancer cells along with testing its toxicity in the normal human fibroblast (NHF) cells. METHODS: The cytotoxic effects of papaverine were examined by the MTT cell viability assay. Flow cytometry using annexin V-FITC/PI was used to study the effects on apoptosis, including its quantification. Effects on cell cycle progression were analyzed by flow cytometry while as effects on apoptosis-related proteins, NF-kB and PI3K/Akt pathways were estimated by Western blot assay. RESULTS: The results indicated that papaverine could induce significant, highly selective and dose-dependent cytotoxic effects in PC-3 cells without causing too much toxicity in normal cells. Papaverine also led to induction of early and late apoptosis along with inducing sub-G1 cell cycle arrest in a dose-dependent manner. Papaverine induced a dose-dependent reduction in the expression levels of Blc-2 proteins and a dose-dependent increase in the expression levels of Bax protein. The expression levels of NF-kB were decreased markedly in comparison to the untreated control. Papaverine treatment also led to a dose-dependent downregulation of PI3K and phospho-Akt expression. CONCLUSION: Papaverine showed selective antitumor properties against PC-3 human prostate cancer cells by inducing early and late apoptosis, sub-G1 cell cycle arrest, modulation of apoptosis-related proteins like Bcl-2, Bax, Bid, XIAP and cytochrome C along with downregulation of NFkB, PI3K/Akt signalling pathway.

[Platelet-derived growth factor-BB induces phenotypic transformation of corpus cavernosum smooth muscle cells in SD rats].

OBJECTIVE: To evaluate the effect of the platelet-derived growth factor-BB (PDGF-BB) on the phenotypic transformation of corpus cavernosum smooth muscle cells (CCSMC) in SD rats. METHODS: CCSMCs were primarily cultured in the modified tissue sticking medium and subjected to immunofluorescence assay. The cells were divided into a blank control and four PDGF-BB groups, the latter exposed to 5, 10, 20, and 40 ng/ml of PDGF-BB, respectively, for 24 hours, and the cells in the 20 ng/ml PDGF-BB group treated for 24, 48, and 72 hours. The the relative expressions of α-SMA, SMMHC, calponin, and OPN mRNA were determined by real-time fluorescence quantitative RT-PCR (qRT-PCR). RESULTS: The α-SMA positive rate of the CCSMCs was over 95%. Compared with the blank control group, the expression levels of α-SMA, SMMHC, and calponin mRNA were significantly decreased (P < 0.05) while that of OPN mRNA remarkably increased (P < 0.05) in the PDGF-BB groups. The 20 ng/ml PDGF-BB group also showed significantly downregulated expressions of α-SMA, SMMHC, and calponin mRNA (P < 0.05) and upregulated expression of OPN mRNA (P < 0.05) at 24, 48, and 72 hours. CONCLUSION: PDGF-BB can induce the transformation of the phenotype of CCSMCs in SD rats from the contractile to the synthetic type.

[Transperitoneal versus extraperitoneal laparoscopic radical prostatectomy for localized prostate cancer: a meta analysis].

OBJECTIVE: To evaluate the effects and safety of transperitoneal laparoscopic radical prostatectomy (TLRP) and extraperitoneal laparoscopic radical prostatectomy (ELRP) in the treatment of localized prostate cancer. METHODS: We searched the Cochrane Library, Medline, Chinese Journal Full-text Database, Wanfang and CBM for clinical controlled trials addressing TLRP and ELRP in the treatment of localized prostate cancer. Two independent reviewers extracted comparable data from eligible studies and performed meta-analysis with the Statal 2.0 software on the relevant indexes of operation time, intraoperative blood loss, postoperative catheterization, postoperative intestinal function recovery, and postoperative hospital stay. RESULTS: Nine clinical controlled trials with 942 cases were included in this analysis, 492 treated by TLRP and the other 450 by ELRP. Meta-analysis showed no statistically significant differences between the TLRP and ELRP groups in operation time (SMD = 0.60, 95% CI: -0.06,1.26), intraoperative blood loss (SMD = 0.01, 95% CI: -0.35, 0.36) , postoperative catheterization time (SMD = 0.10, 95% CI: -0.21, 0.40) and postoperative hospital stay (SMD = 0.45, 95% CI: -0.01, 0.91), except in the time of postoperative intestinal function recovery, which was significantly shorter in the ELRP than in the TLRP group (SMD = 1.18, 95% CI: 0.26, 2.10). CONCLUSION: For the treatment of localized prostate cancer, ELRP is similar to TLRP with respect to operation time, intraoperative blood loss, postoperative catheterization and postoperative hospital stay, but superior to the latter in postoperative intestinal function recovery.

N-acetylcysteine ameliorates erectile dysfunction in rats with hyperlipidemia by inhibiting oxidative stress and corpus cavernosum smooth muscle cells phenotypic modulation.

Hyperlipidemia is a major risk factor for erectile dysfunction (ED). Oxidative stress and phenotypic modulation of corpus cavernosum smooth muscle cells (CCSMCs) are the key pathological factors of ED. N-acetylcysteine (NAC) can inhibit oxidative stress; however, whether NAC can alleviate pathological variations in the corpus cavernosum and promote erectile function recovery in hyperlipidemic rats remains unclear. A hyperlipidemia model was established using 27 eight-week-old male Sprague-Dawley (SD) rats fed a high-fat and high-cholesterol diet (hyperlipidemic rats, HR). In addition, 9 male SD rats were fed a normal diet to serve as controls (NC). HR rats were divided into three groups: HR, HR+normal saline (NS), and HR+NAC (n = 9 for each group; NS or NAC intraperitoneal injections were administered daily for 16 weeks). Subsequently, the lipid profiles, erectile function, oxidative stress, phenotypic modulation markers of CCSMCs, and tissue histology were analyzed. The experimental results revealed that erectile function was significantly impaired in the HR and HR + NS groups, but enhanced in the HR + NAC group. Abnormal lipid levels, over-activated oxidative stress, and multi-organ lesions observed in the HR and HR + NS groups were improved in the HR + NAC group. Moreover, the HR group showed significant phenotypic modulation of CCSMCs, which was also inhibited by NAC treatment. This report focuses on the therapeutic effect of NAC in restoring erectile function using a hyperlipidemic rat model by preventing CCSMC phenotypic modulation and attenuating oxidative stress.

[Calcitonin gene-related peptide induces phenotypic transformation of corpus cavernosum smooth muscle cells in diabetic rats with erectile dysfunction].

OBJECTIVE: To explore the effect of the calcitonin gene related peptide (CGRP) on the phenotypic transformation of corpus cavernosum smooth muscle cells (CCSM) in diabetic rats with erectile dysfunction (ED). METHODS: Models of diabetes and diabetic ED were established in male Sprague-Dawley rats by administration of streptozotocin, and CCSMs were primarily cultured and subjected to immunocytochemical assay. The cells were divided into a diabetic ED and a normal control group, and exposed to 0, 10, 60 and 100 nmol/L of CGRP for 24 hours. Then the relative expressions of calponin 1 (Cnn1) and osteopontin (OPN) mRNA were determined by real-time fluorescence quantitative RT-PCR (qRT-PCR). RESULTS: The rate of SMalpha-actin positive cells in the CCSMs was (95.94 +/- 0.03) %. The expression of Cnn1 mRNA was significantly lower while that of OPN mRNA remarkably higher in the diabetic ED rats (4.41 +/- 0.29 and 5.28 +/- 0.32) than in the normal controls (10.35 +/- 0.62 and 1.32 +/- 0.24) (P < 0.01). Exposure to 100 nmol/L of CGRP significantly upregulated the expression of Cnn1 mRNA and downregulated that of OPN mRNA as compared with the unexposed rats (6.9 +/- 0.22 vs 4.41 +/- 0.29 and 3.26 +/- 0.31 vs 5.28 +/- 0.32, P < 0.01). CONCLUSION: CGRP can transform the phenotype of CCSMs in diabetic ED rats from contractile to synthetic type.

In vivo tracking on longer retention of transplanted myocardin gene-modified adipose-derived stem cells to improve erectile dysfunction in diabetic rats.

BACKGROUND: Stem cell therapy has revealed a promising future for treating erectile dysfunction (ED), but the fate and curative mechanism of intracavernosal transplanted stem cells are under further exploration. This study aimed to demonstrate the effects of myocardin gene modification on improving erectile function and prolonging the retention of implanted adipose-derived stem cells (ASCs) using in vivo small animal imaging. METHODS: ASCs were isolated, cultured, and identified by flow cytometry and osteogenic and adipogenic induction. The effects of gene modification on cell proliferation, apoptosis, and contraction were determined by CCK-8, EdU, flow cytometry, and collagen gel lattice contraction assays as well as confocal microscopy. A total of 20 normal and 60 diabetes mellitus ED to (DMED) Sprague-Dawley rats were recruited to the 7 day and 21 day groups. Each group contained subgroups of 10 rats each: the negative control (NC), DMED + ASCs plus Ad-Luc-Myocardin, DMED + ASCs plus Ad-Luc, and DMED + phosphate buffer solution (PBS) groups. Erectile function was evaluated with the intracavernosal pressure/mean arterial pressure (△ICP/MAP) ratio. In vivo small animal imaging and an EdU cell tracking strategy were introduced to detect the transplanted ASCs, and IHC and WB were performed to assess smooth muscle cell protein levels. RESULTS: The ASCs expressed high CD29 and CD90 and scant CD45, while the multi-induction potential was verified by oil red O and alizarin red staining. Gene transfection of myocardin had no significant influence on ASC apoptosis but inhibited cell proliferation and promoted cell contraction. Myocardin combined with ASCs enhanced the therapeutic potential of ASCs for improving the △ICP/MAP ratio as well as α-SMA and calponin expression. In vivo imaging confirmed that ASCs resided within the cavernous body in 21 days, while only a few red EdU dots were detected. CONCLUSIONS: Myocardin induced ASC differentiation towards smooth muscle-like cells and enhanced the therapeutic potential of ASCs for ameliorating ED in STZ-induced diabetic rats. Notably, in vivo small animal tracking was an effective strategy for monitoring the implanted stem cells, and this strategy might have advantages over traditional EdU assays.

[Phenotype modulation of smooth muscle in corpus cavernosum in penis tunica albuginea in diabetes mellitus with erectile dysfunction: experiment with rats].

OBJECTIVE: To investigate the phenotype modulation of smooth muscle of corpus cavernosum in diabetes mellitus with erectile dysfunction. METHODS: Thirty SD rats were randomly divided into 2 equal groups: 4-week group and 7-week group, injected with streptozocin to cause diabetes mellitus (DM), and then subdivided into DM group [with DM and without erectile dysfunction. (ED)], DM + ED group (with DM and ED), and group of failure to cause DM (Group None) according whether DM was induced. Another 10 rats without STZ injection were divided into 4-week and 7-week control groups. The penis was resected and immunohistochemistry and color image analysis were used to observe the expression of alpha-actin and desmin in the corpus cavernosum. In situ hybridization was used to detect the mRNA expression of osteopontin (OPN), characteristic of noncontractil phenotype. RESULTS: The alpha-actin expression of smooth muscle in corpus cavernosum in the DM + ED group was significantly lower than those of the other groups (all P < 0.05). No significant difference existed between the 7 week group and 4 week group (F = 3.801, P = 0.62), and there was significant interaction (F = 1.549, P = 0.225). There was no significant difference in the desmin expression of smooth muscle in corpus cavernosum among different groups (F = 0.045, P = 0.987) and there was not significant interaction (F = 0.572, P = 0.639). The OPN mRNA expression of smooth muscle in corpus cavernosum of the DM + ED group was significantly higher than those of the other subgroups (F = 156.439, P = 0.000). No significant difference existed between the 7 week group and 4 week group (F = 1.288, P = 0.266), and there was no significant interaction (F = 1.819, P = 0.168). CONCLUSION: Phenotype modulation of smooth muscle in corpus cavernosum can be caused by DM. ED can be caused by phenotype modulation of smooth muscle in corpus cavernosum.

Corrigendum to "Combination of low-energy shock-wave therapy and bone marrow mesenchymal stem cell transplantation to improve the erectile function of diabetic rats" by Hai-Tao Shan et al.

[This corrects the article DOI: 10.4103/1008-682X.184271.].

Combination of low-energy shock-wave therapy and bone marrow mesenchymal stem cell transplantation to improve the erectile function of diabetic rats.

Stem cell transplantation and low-energy shock-wave therapy (LESWT) have emerged as potential and effective treatment protocols for diabetic erectile dysfunction. During the tracking of transplanted stem cells in diabetic erectile dysfunction models, the number of visible stem cells was rather low and decreased quickly. LESWT could recruit endogenous stem cells to the cavernous body and improve the microenvironment in diabetic cavernous tissue. Thus, we deduced that LESWT might benefit transplanted stem cell survival and improve the effects of stem cell transplantation. In this research, 42 streptozotocin-induced diabetic rats were randomized into four groups: the diabetic group (n = 6), the LESWT group (n = 6), the bone marrow-derived mesenchymal stem cell (BMSC) transplantation group (n = 15), and the combination of LESWT and BMSC transplantation group (n = 15). One and three days after BMSC transplantation, three rats were randomly chosen to observe the survival numbers of BMSCs in the cavernous body. Four weeks after BMSC transplantation, the following parameters were assessed: the surviving number of transplanted BMSCs in the cavernous tissue, erectile function, real-time polymerase chain reaction, and penile immunohistochemical assessment. Our research found that LESWT favored the survival of transplanted BMSCs in the cavernous body, which might be related to increased stromal cell-derived factor-1 expression and the enhancement of angiogenesis in the diabetic cavernous tissue. The combination of LESWT and BMSC transplantation could improve the erectile function of diabetic erectile function rats more effectively than LESWT or BMSC transplantation performed alone.

[Phenotypic modulation of bladder smooth muscle in diabetic rats].

OBJECTIVE: To investigate whether phenotypic modulation of bladder smooth muscle occurs in diabetic rats. METHODS: Thirty-two male SD rats were randomly assigned into diabetic group and control group. Diabetic rat models were established by a single intraperitoneal injection of streptozotocin (60 mg/kg). Nine weeks later, the bladder tissues of the rats were examined for structural changes using HE and Masson's trichrome staining , and the expressions of myocardin, α-SMA, and SMMHC in bladder smooth muscles were detected with RT-PCR and Western blotting. RESULTS: Compared with the control group, the diabetic rats showed obvious polydipsia and polyuria with significantly increased collagenous fibers and lowered expressions of myocardin, α-SMA, and SMMHC in the bladder tissue (P<0.05). CONCLUSION: s In rats at 9 weeks after diabetic model establishment, phenotypic transition of the bladder smooth muscles occurs to cause bladder contractile dysfunction, which may play an important role in the pathology of diabetic bladder dysfunction.

[Advances in researches on the relationship between prostatic diseases and erectile dysfunction].

Prostatic diseases and erectile dysfunction (ED) are common diseases in urology and andrology. Basic and clinical studies have proved that there is a close relationship between the two. This article reviews the mechanism, diagnosis and treatment of ED caused by several prostatic diseases, such as acute prostatitis, chronic prostatitis, benign prostate hyperplasia and prostate cancer.

Total triterpenoids from Ganoderma Lucidum suppresses prostate cancer cell growth by inducing growth arrest and apoptosis.

In this study, one immortalized human normal prostatic epithelial cell line (BPH) and four human prostate cancer cell lines (LNCaP, 22Rv1, PC-3, and DU-145) were treated with Ganoderma Lucidum triterpenoids (GLT) at different doses and for different time periods. Cell viability, apoptosis, and cell cycle were analyzed using flow cytometry and chemical assays. Gene expression and binding to DNA were assessed using real-time PCR and Western blotting. It was found that GLT dose-dependently inhibited prostate cancer cell growth through induction of apoptosis and cell cycle arrest at G1 phase. GLT-induced apoptosis was due to activation of Caspases-9 and -3 and turning on the downstream apoptotic events. GLT-induced cell cycle arrest (mainly G1 arrest) was due to up-regulation of p21 expression at the early time and down-regulation of cyclin-dependent kinase 4 (CDK4) and E2F1 expression at the late time. These findings demonstrate that GLT suppresses prostate cancer cell growth by inducing growth arrest and apoptosis, which might suggest that GLT or Ganoderma Lucidum could be used as a potential therapeutic drug for prostate cancer.

[Role of miR-145 in erectile dysfunction in diabetic rats].

OBJECTIVE: To investigate the role of miR-145 in the corpus cavernosum smooth muscle tissue in the pathogenesis of erectile dysfunction (ED) in diabetic rats. METHODS: The total RNA was extracted from the corpus cavernosum of a diabetic rat model with ED, diabetic rats with normal erectile function and normal rats, and the expression levels of miR145 were compared between the groups. RESULTS: The expression of miR-145 was decreased in the corpus cavernosum of diabetic rats with ED. CONCLUSION: Diabetes mellitus can cause ED in rats, in which process decreased expression of miR145 in the corpus cavernosum may play a role.

[Microarray data analysis of genes related with erectile dysfunction in diabetic rats].

OBJECTIVE: To investigate the changes of gene expression profiles associated with erectile dysfunction in diabetic rats. METHODS: Affymetrix Gene Chip arrays from the Gene Expression Omnibus (GEO) were used to examine the alterations in the gene expression profiles between streptozotocin-induced diabetic rats and littermate controls, and the data were analyzed with GeneSifter microarray analysis software. RESULTS: A total of 661 differentially expressed genes were identified, including 280 up-regulated and 381 down-regulated ones. Among the differentially expressed genes, kruppel-like factor 5 (klf5) was upregulated by 4.01 folds and ceruloplasmin(cp) by 5.14 folds; collagen, type XI, alpha1 was down-regulated by 5.84 folds and collagen, type I, alpha1 by 5.77 folds. The 661 differentially expressed genes involved such functional processes as glycoprotein biosynthesis, collagen fibril organization, angiogenesis in wound healing, triglyceride metabolism, cell proliferation and other important biological processes, and some pathways also involved such as fatty acid metabolism, neurodegenerative disorders, and ECM-receptor interactions. CONCLUSION: Some genes such as klf5, cp, and collagen play important roles in the pathophysiology of diabetes-induced erectile dysfunction. Bioinformatic approaches offer a new means for identifying candidate genes and pathways relevant to the pathophysiology of diabetes-induced erectile dysfunction, highlighting also the potential complexity of this disorder.

[Retroperitoneal laparoscopic surgery combined with ureteroscopic lithotomy for treatment of renal and ureteral calculi].

OBJECTIVE: To evaluate the feasibility of retroperitoneal laparoscopic surgery combined with ureteroscopic lithotomy through the pelvis for treatment of renal and ureteral calculi. METHODS: In February 2010, 2 patients with renal and ureteral calculi underwent retroperitoneal laparoscopic surgery combined with ureteroscopic lithotomy through the pelvis. RESULTS: The operation time in these two cases was 70 and 80 min, and the volume of intraoperative blood loss was about 20 ml. The exposure was excellent, and the patient recovered rapidly without complications or residual calculi. CONCLUSION: Retroperitoneal laparoscopic surgery combined with ureteroscopic lithotomy through the pelvis is feasible for treatment of renal and ureteral calculi.

[A modified tissue culture method for culturing corpus cavernosum smooth muscle cells from the penis of Sprague-Dawley rats].

OBJECTIVE: To culture rat corpus cavernosum smooth muscle cells in vitro using a modified tissue culture method. METHODS: Fifteen male rats were randomized into 3 equal groups, namely enzyme digestion group, tissue culture group, and modified tissue culture group. The penis of the rats was separated carefully and cut into small pieces, and seeded onto culture flasks and cultured in complete medium consisting of DMEM containing 20% fetal calf serum at 37 degrees C; in a humidified atmosphere with 5% carbon dioxide. The cells growth was observed under phase contrast microscope and the smooth muscle cell specific proteins alpha-SM-actin and desmin were identified immunohistochemically. RESULTS: The alpha-SM-actin-positive cell rate was 96.3% in rat corpus cavernosum smooth muscle and 23.8% in the fibroblasts, and the corpus cavernosum smooth muscle contained 74.4% desmin-positive cells while the fibroblasts showed no desmin positivity. Significant difference was found in the positive cell rate for desmin among the 3 groups, with the highest positive cell rate occurred in modified tissue culture group. CONCLUSION: Desmin may serve as a marker for identifying corpus cavernosum smooth muscle cells. The modified tissue culture method can result in highly purified corpus cavernosum smooth muscle cells with intact structure and functions.

[Establishment of a cell culture model of corpus cavernosum smooth muscle cells derived from diabetic rats with erectile dysfunction].

OBJECTIVE: To investigate the method for culturing corpus cavernosum smooth muscle cells (CCSMs) derived from diabetic rats with erectile dysfunction (ED) for the study of ED caused by diabetes. METHODS: CCSMs were isolated from the corpus cavernosum of diabetic rats with ED and cultured using a modified method of adherent tissue culture. The cultured cells were identified by immunohistochemistry and the cell morphology and proliferation were observed. RESULTS: The primary culture of CCSM was performed successfully, and the cells were seen to migrate from the small tissue pieces 3 days later, reaching nearly confluence in 16-18 days. A typical "hill-valley" growth pattern was noted in the cell passaging. Immunohistochemical staining for alpha-smooth muscle actin (alpha-SM-actin) and desmin yielded positive results in the cells. CONCLUSION: The modified method for adherent tissue culture is convenient and reliable in establishing the in vitro cell culture model of CCSMs from diabetic rats with ED, and the cultured CCSMs display a faster proliferation than normal CCSMs. No obvious differences in the cell morphology can be found between diabetic and normal CCSMs under light microscope.

[Phenotypic transformation of corpus cavernosum smooth muscle cells in hypertensive rats].

OBJECTIVE: To validate the hypothesis that the phenotypic transformation occurs in the smooth muscle cells in the corpus cavernosum of hypertensive rat and explore its impact on the erectile function of rats. METHODS: Eighteen 16-week-old male spontaneously hypertensive rats (SHR) and 10 syngeneic normotensive rats (WKY) were used in this experiment. After measurement of systolic blood pressure of the caudal artery and examination of the erectile function with subcutaneous injection of apomorphine (APO), the rats were divided into 3 groups, namely hypertensive with erectile dysfunction (HBP-ED) group (n=6), hypertensive (HBP) group (n=12) and control group (n=10). Immunohistochemical staining and color image analysis system were used to observe expression of calponin 1 and osteopontin (OPN) in rat corpus cavernosum. Real-time quantitative RT-PCR was used to determine the expression of calponin 1 and OPN mRNAs in different groups. RESULTS: The expressions of calponin 1 protein and mRNA were the highest in the control group and the lowest in HBP-ED group, while the expressions of OPN protein and mRNA were the highest in HBP-ED group and the lowest in the control group. CONCLUSION: The smooth muscle cells may transform from the contractile phenotype into synthetic phenotype in the corpus cavernosum of the hypertensive rats, resulting ultimately in erectile dysfunction.

[Establishment of a rat model of diabetic erectile dysfunction].

OBJECTIVE: To establish a rat model of diabetic erectile dysfunction (ED) with streptozotocin (STZ) injection. METHODS: Thirty male rats were randomized equally into 5 groups (control group and STZ 40 mg/kg, 60 mg/kg, 80 mg/kg, and 100 mg/kg groups). All the rats were examined at 4 days and 1, 2, and 3 weeks after STZ injection for fasting blood glucose, erectile frequency induced by apomorpHine (APO) and body weight changes. RESULTS: Significant difference occurred in the fasting food glucose among the groups at different time points (P=0.001), and also in APO-induced erectile frequency, fasting blood glucose and body weight between the groups with STZ injection at different doses (P<0.001, P=0.045 and P<0.001, respectively). No significant difference was found in induced erectile frequency and body weight between different time points (P=0.306 and P=0.628). CONCLUSION: The optimal dose of STZ for establishing diabetic ED model is 60 mg/kg, and two weeks after the injection can be the optimal time for evaluating model establishment by means of APO-induced penis erection.