Inhibition of Proliferation of Prostate Cancer Cell Line DU-145 in vitro and in vivo Using Salvia miltiorrhiza Bunge / 中国结合医学杂志

OBJECTIVE@#To investigate the antiproliferative activity of Salvia miltiorrhiza Bunge. (SM) on the castration-resistant prostate cancer (CRPC) cell line DU-145, in vitro and in vivo.@*METHODS@#Prostate cancer cell line (DU-145) and normal prostate cell line (RWPE-1) were treated with SM at different concentrations (3.125, 12.5, 25 and 50 μg/mL) to investigate the antiproliferative effects. DNA laddering analysis was performed to investigate the apoptosis of DU-145 cells. Molecular mechanism was investigated by Western blot analysis of p53, Bcl-2, prostate specific antigen (PSA), and androgen receptor (AR). Six-week-old male BALB/c nude mice were randomly divided into normal control group (n=101) and treated group (n=101) which administered 500 mg/kg SM for 2 weeks. Tumor volumes were measured.@*RESULTS@#Treatment with SM resulted in a dose-dependent decrease in cell number of DU-145 cells in comparison with RWPE-1. DNA laddering analysis indicated the apoptosis of DU-145 cells. Treatment with SM increased the expression of p53 and reduced the expression of Bcl-2 proteins. The levels of PSA were considerably reduced in SM-treated group compared to the controls, and a decrease in AR expression was observed when cells were treated with SM in the same pattern as a reduction in PSA. In the tumour xenograft study, SM given once a day for 2 weeks significantly inhibited tumour growth.@*CONCLUSION@#SM might contribute to the anticancer actions such as induction of apoptosis and inhibition of proliferation of prostate cancer cells.

Effect of Korean Herbal Formula (Modified Ojayeonjonghwan) on Androgen Receptor Expression in an Aging Rat Model of Late Onset Hypogonadism

PURPOSE: Testosterone replacement therapy is an effective treatment for late-onset hypogonadism (LOH) despite a few contraindications and side-effects. The aim of this study was to determine whether modified Ojayeonjonghwan (KH-204, Korean herbal formula) improved LOH. KH-204 is a strong antioxidant herbal formula. We evaluated the effect of Korean herbal prescription on androgen receptor (AR) expression in an aged rat model of LOH. MATERIALS AND METHODS: Eighteen-month-old rats were used as aged LOH rat models. Eighteen Sprague-Dawley rats were randomly divided into three equal groups of six animals each and treated with one of the following: 1) normal control group (oral administration with distilled water, n=6), 2) KH-204 200 group (oral administration with 200 mg/kg of KH-204, n=6), and 3) KH-204 400 group (oral administration with 400 mg/kg of KH-204, n=6). After four weeks of treatment (once daily, distilled water or KH-204), serum testosterone levels, changes in testicular and epididymal weight, Western blotting analysis of AR expression and measurement of oxidative stress were examined. RESULTS: Treatment with the herbal formulation KH-204 200 mg/kg and 400 mg/kg (1) increased the weights of testis and epididymis; (2) increased the level of serum testosterone; (3) increased the level of superoxide dismutase and reduced the level of 8-hydroxy-20-deoxyguanosine; and (4) upregulated AR expression in testicular tissue. CONCLUSIONS: KH-204 might be an effective alternative for LOH. It improves antioxidant mechanisms and increases testicular AR expression without side-effects.

Improvement of Persistent Detrusor Overactivity through Treatment with a Phytotherapeutic Agent (WSY-1075) after Relief of Bladder Outlet Obstruction

PURPOSE: Many patients with benign prostatic hyperplasia need treatment for remaining storage symptoms after surgery. Therefore, we evaluated the effect of the phytotherapeutic agent WSY-1075 on persistent detrusor overactivity (DO) after the relief of bladder outlet obstruction (BOO). MATERIALS AND METHODS: Rats were assigned to 3 groups: control (n=6), persistent DO (n=6), and persistent DO treated with the phytotherapeutic agent WSY-1075 (n=6). Persistent DO after relief of partial BOO was generated in the rat model, and 6 of the rats with this condition were orally administered WSY-1075. After 4 weeks of administration, cystometry was performed. Additionally, 8-hydroxy-2-deoxyguanosine and superoxide dismutase were measured to evaluate oxidative stress in the bladder. Pro-inflammatory cytokines, such as interleukin-8 and tumor necrosis factor-α, were analyzed, as were the M2 and M3 muscarinic receptors of the bladder. RESULTS: Significantly increased contraction pressure and a decreased contraction interval were observed in the persistent DO group after relief of BOO. Moreover, oxidative stress, pro-inflammatory cytokines, and M3 muscarinic receptors were significantly increased. After treatment with WSY-1075, significantly reduced DO was observed by cystometry in comparison with the persistent DO group. Additionally, significantly decreased levels of oxidative stress, pro-inflammatory cytokines, and M3 muscarinic receptors in the bladder were observed after treatment with WSY-1075. CONCLUSIONS: Treatment with WSY-1075 improved persistent DO after the relief of BOO mediated by antioxidative and anti-inflammatory effects. Further studies are necessary to identify the exact mechanism of the treatment effect of WSY-1075.

Suppression of Oxidative Stress of Modified Gongjin-Dan (WSY-1075) in Detrusor Underactivity Rat Model Bladder Outlet Induced by Obstruction / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the anti-oxidative stress and preventive effect of modified Gongjin-dan (WSY-1075) in a detrusor underactivity rat model.</p><p><b>METHODS</b>Rats were randomly allocated to three groups: shamoperated (control), bladder outlet obstruction-induced detrusor underactivity (BOO-DU), and BOO-DU with WSY-1075 (WSY) groups. WSY-1075 was orally administrated to rats 200 mg daily for 2 weeks prior to the operation and 4 weeks after the operation. Bladder outlet obstruction was surgically induced in rats by ligation around the urethra avoiding total obstruction. Cystometrography was conducted on rats in each group for examination of bladders.</p><p><b>RESULTS</b>Compared with the control group, bladder outlet obstruction led to a significant increase in oxidative stress with consequent changes to molecular composition, and decrease in maximal detrusor pressure (P<0.05). WSY-1075 treatment significantly suppressed oxidative stress and prevented degenerative and dysfunctional changes in bladder, as compared with BOO-DU group (P<0.05).</p><p><b>CONCLUSION</b>WSY-1075 had beneficial effect on prevention of BOO-DU.</p>

Corrigendum: Improvement of Persistent Detrusor Overactivity through Treatment with a Phytotherapeutic Agent (WSY-1075) after Relief of Bladder Outlet Obstruction

Current affiliation of Su Jin Kim has been changed, but it was not reflected in the process of publishing. The publishing office and editorial office would like to apologize for any inconvenience caused.

Restoration of Spermatogenesis Using a New Combined Herbal Formula of Epimedium koreanum Nakai and Angelica gigas Nakai in an Luteinizing Hormone-Releasing Hormone Agonist-Induced Rat Model of Male Infertility

PURPOSE: We investigated the protective effect of a mixture of 2 herbal extracts, KH-465, which consisted of Epimedium koreanum Nakai and Angelica gigas Nakai, on spermatogenesis in a luteinizing hormone-releasing hormone (LHRH) agonist-induced rat model of male infertility. MATERIALS AND METHODS: Seventy-five 12-week-old male Sprague-Dawley rats were randomly divided into 5 groups, containing 15 rats each: a normal control group that received no treatment and 4 experimental groups (I, II, III, and IV) in which an LHRH agonist was administered for 4 weeks to induce spermatogenic failure. Group I received distilled water, and groups II, III, and IV received 200 mg/kg/day of KH-465, 400 mg/kg/day KH-465, and depo-testosterone for 4 weeks, respectively. Weight changes of the testis and epididymis, sperm count motility, and levels of testosterone (T), free T, follicle-stimulating hormone (FSH), luteinizing hormone (LH), superoxide dismutase (SOD), and 8-hydroxy-2′-deoxyguanosine (8-OHdG) were estimated. RESULTS: Body, testis, and epididymis weight showed no significant differences among the control and experimental groups. Treatment with KH-465 increased the sperm count and motility. Serum hormone levels of T, free T, and FSH were not significantly different in the experimental groups, while the LH level was higher than in the LHRH agonist-induced control group, but not to a significant extent. Levels of SOD were higher and 8-OHdG were lower in the groups that received KH-465 than in the LHRH agonist-induced control group. CONCLUSIONS: Our results suggest that KH-465 increased sperm production via reducing oxidative stress and had a positive effect in a male infertility model.

Development of an Improved Animal Model of Overactive Bladder: Transperineal Ligation versus Transperitoneal Ligation in Male Rats

PURPOSE: We compared a transperineal ligation model and a transperitoneal ligation model in male rats to determine which animal model of overactive bladder (OAB) was more useful based on cystometrography, estimations of oxidative stress, and measurements of pro-inflammatory cytokine levels. MATERIALS AND METHODS: Male rats were randomly divided into three groups (n=15 in each): the control group, the transperineal ligation group, and the transperitoneal ligation group. Four weeks after the ligation procedure, cystometrography was performed and oxidative stress, pro-inflammatory cytokine levels, and histologic changes were evaluated. Oxidative stress was assessed by measuring 8-hydroxy-20-deoxyguanosine and superoxide dismutase, and pro-inflammatory cytokine activity was investigated by measuring levels of interleukin (IL)-6, IL-8, and tumor necrosis factor-α. RESULTS: The transperineal model led to results similar to those observed for the transperitoneal model, namely (1) increased voiding frequency and reductions in the non-voiding contraction interval and the maximal vesical pressure, (2) increased levels of oxidative stress markers, (3) increased pro-inflammatory cytokine levels, and (4) fibrotic changes in the bladder tissue. CONCLUSIONS: We suggest that the transperineal procedure can be used as an alternative OAB model in male rats.

Protective Effects of KH-204 in the Bladder of Androgen-Deprived Rats

PURPOSE: We investigated the protective effects of the herbal formulation KH-204 in the bladder of androgen-deprived rats. MATERIALS AND METHODS: Male rats aged eight weeks were randomly divided into four groups, containing eight rats each: sham operation only (normal control group), androgen-deprived only (androgen-deprived control group), and androgen-deprived followed by treatment with 200 mg/kg or 400 mg/kg of KH-204. After 0.5 mg/kg of leuprorelin was subcutaneously injected in the androgen-deprived groups, the oral administration of either distilled water in the two control groups or KH-204 in the treatment group was continued for four weeks. Serum testosterone levels, RhoGEF levels, nitric oxide (NO)-cyclic guanosine monophosphate (cGMP)-related parameters, oxidative stress, and histologic changes were evaluated after treatment. RESULTS: Treatment with the herbal formulation KH-204 (1) increased serum testosterone levels; (2) restored the expression of RhoGEFs, endothelial NO synthase, and neuronal NO synthase; (3) increased the expression of superoxide dismutase; and (4) decreased bladder fibrosis. CONCLUSIONS: Our results suggest that the positive effects of KH-204 on the urinary bladder may be attributed to its antioxidant effects or to an elevation in NO-cGMP activity.

Anthocyanin Induces Apoptosis of DU-145 Cells In Vitro and Inhibits Xenograft Growth of Prostate Cancer

PURPOSE: To investigate the effects of anthocyanins extracted from black soybean, which have antioxidant activity, on apoptosis in vitro (in hormone refractory prostate cancer cells) and on tumor growth in vivo (in athymic nude mouse xenograft model). MATERIALS AND METHODS: The growth and viability of DU-145 cells treated with anthocyanins were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and apoptosis was assessed by DNA laddering. Immunoblotting was conducted to evaluate differences in the expressions of p53, Bax, Bcl, androgen receptor (AR), and prostate specific antigen (PSA). To study the inhibitory effects of anthocyanins on tumor growth in vivo, DU-145 tumor xenografts were established in athymic nude mice. The anthocyanin group was treated with daily oral anthocyanin (8 mg/kg) for 14 weeks. After 2 weeks of treatment, DU-145 cells (2x106) were inoculated subcutaneously into the right flank to establish tumor xenografts. Tumor dimensions were measured twice a week using calipers and volumes were calculated. RESULTS: Anthocyanin treatment of DU-145 cells resulted in 1) significant increase in apoptosis in a dose-dependent manner, 2) significant decrease in p53 and Bcl-2 expressions (with increased Bax expression), and 3) significant decrease in PSA and AR expressions. In the xenograft model, anthocyanin treatment significantly inhibit tumor growth. CONCLUSION: This study suggests that anthocyanins from black soybean inhibit the progression of prostate cancer in vitro and in a xenograft model.

The Effect of Anthocyanin on the Prostate in an Andropause Animal Model: Rapid Prostatic Cell Death by Apoptosis Is Partially Prevented by Anthocyanin Supplementation

PURPOSE: To evaluate the anti-apoptotic effect of the antioxidant reaction of anthocyanin on the prostate in an andropause animal model. MATERIALS AND METHODS: Sprague-Dawley rats were divided into three groups (n=12 in each): control (Group I), andropause (Group II), andropause treated with anthocyanin (Group III). For induction of andropause, Group II and III underwent bilateral orchiectomy. Group III was treated with daily oral anthocyanin (160 mg/kg) for 8 weeks. After 8 weeks, the rats were sacrificed and their blood and prostates were examined pathohistologically and evaluated for oxidative stress and apoptosis. Oxidative stress was assessed by the activity of superoxide dismutase (SOD) and apoptosis in the prostate was identified by terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling assay. RESULTS: Group II showed markedly increased activity of SOD in serum over that observed in Group I, whereas the rats in Group III showed reduced oxidative stress compared to Group II. Despite no significant differences in prostate weight between Group II and III (p=0.078), the apoptotic index was significantly greater in Group II than Group I, and was significantly lesser in Group III than Group II. CONCLUSIONS: We suggest that the oxidative stress caused by low testosterone may be another inducer of apoptosis, and this apoptosis may partly contribute to the overall apoptosis of the prostate in the andropause animal model. Therefore, anthocyanin supplementation may contribute to preventing excessively rapid cell death by apoptosis in the prostate in an animal model of andropause.

Preliminary Report on the Safety of a New Herbal Formula and Its Effect on Sperm Quality

PURPOSE: Male infertility is a serious problem, and its prevalence has been increasing. Therefore, we investigated the safety of a new herbal formula and its effects on sperm quality. MATERIALS AND METHODS: An in vitro cytotoxicity test in TM3 Leydig cells was performed to evaluate cell viability after administration of five types of herbs separately and of a new herbal formula containing these five. An in vivo test in male mice was performed to evaluate the influence of the new herbal formula on the reproductive organs and sperm quality. After the 8- and 28-day oral administration of the new herbal formula, the weights of the reproductive organs were measured and the sperm count and motility were evaluated. RESULTS: In the in vitro cytotoxicity test, less than 80% cell viability at concentrations of 500 mg/L and 1,000 mg/L of Rubus coreanus Miquel and Cuscuta chinensis Lam was observed. However, more than 80% cell viability was observed at all the tested concentrations of the new herbal formula. After the 8- and 28-day oral administration, there were no considerable changes in body weight. The weights of the testes, epididymis, and seminal vesicles after the 8- and 28-day oral administration were similar to those of the control. The sperm count and activity were significantly improved compared with those of the control group at 8 and 28 days after 100, 200, and 400 mg of oral administration. CONCLUSIONS: The safety of the new formula and its positive effect on the sperm quality were observed after the oral administration of the formula.

The Anti-Inflammatory Effects of a New Herbal Formula (WSY-1075) in a Nonbacterial Prostatitis Rat Model

PURPOSE: The aim of this study was to investigate the anti-inflammatory effects of a new herbal formula (WSY-1075) in a nonbacterial prostatitis rat model. MATERIALS AND METHODS: Prostatitis was induced in male Wistar rats (n=32) by treatment with 17 beta-estradiol and dihydrotestosterone for 4 weeks. After the induction of prostatitis, the rats were randomly divided into one of four treatment groups: control (n=8), ciprofloxacin (n=8), WSY-1075 (100 mg/kg) (n=8), and WSY-1075 (400 mg/kg) (n=8). After 4 weeks of treatment, the prostatic proinflammatory cytokine (tumor necrosis factor-alpha, interleukin [IL]-6, and IL-8) levels and histological findings were noted. RESULTS: The ciprofloxacin and WSY-1075 treatment groups showed significantly decreased proinflammatory cytokine levels compared with the control group. Histologically, treatment with ciprofloxacin and WSY-1075 significantly suppressed the severity of prostatitis lesions compared with those in the control group. No differences in the proinflammatory cytokine levels or histologic findings were observed with the dose dependent treatment of WSY-1075. CONCLUSIONS: The new herbal formula, WSY-1075, showed effective anti-inflammatory activities in the prostate and may be useful for the clinical treatment of nonbacterial prostatitis. Our findings suggest that WSY-1075 has a beneficial effect on the prevention and treatment of nonbacterial prostatitis.

The Effects of New Herbal Formula (KBMSI-1) on the Penile Erection and Corpus Cavernosum in Streptozotocin-induced Diabetic Rats / 대한남성과학회지

PURPOSE: The aim of this study was to investigate the effects of new herbal formula (KBMSI-1) on erectile dysfunction in streptozotocin-induced diabetic rat model. MATERIALS AND METHODS: We used male Sprague-Dawley rats aged 12 weeks and divided into three groups; control (n=8), diabetes (DM) (n=8), DM+KBMSI-1 200 mg/kg treatment (n=8) groups. The DM groups received a single intraperitoneal injection of streptozotocin (STZ). Distilled water was administered in the control and DM group. To investigate the penile erection, intracavernosal pressure (ICP) and intracavernosal pressure/mean arterial pressure (ICP/MAP) were recorded in all groups. Serial sections of the penis were used to perform Masson's trichrome stain. We analyzed the expression of nNOS and eNOS concentration in the isolated corpus cavernosum by western blotting. RESULTS: Peak ICP/MAP ratio was markedly increased in the treatment group with KBMSI-1 compared with DM group (p<0.05). Masson's trichrome staining of corpus cavernosum showed increase in smooth muscle volume and the regular arrangement of collagen fibers in KBMSI-1 treatment group compared with DM group. Western blot analysis revealed that the penile expressions of nNOS and eNOS protein were significantly higher in KBMSI-1-treated group than in DM group. CONCLUSIONS: This study showed that herbal formulation of KBMSI-1 enhances the penile erection and the level of eNOS and nNOS expression of penile corpus cavernosum in streptozotocin-induced diabetic rat model.

The Changes of Testis and the Effects of Anthocyanin on Spermatogenesis in Rat Induced Varicocele / 대한남성과학회지

PURPOSE: Varicocele is known as a main cause of primary male infertility and it supposed to be associated with oxidative stress. Anthocyanin is known as a natural plant pigment and novel antioxidant. This study was designed to investigate the effects of anthocyanin on a rat model of varicocele. MATERIALS AND METHODS: Twenty four male rats, induced varicocele by partial obstruction of left renal vein, were divided into four experimental groups: the group induced varicocele for four weeks without anthocyanin, the group received anthocyanin (80 mg/kg) right after varicocele induction, group induced varicocele for eight weeks without anthocyanin, and the group received anthocyanin (80 mg/kg) after four weeks observation following varicocele induction. After anthocyanin treatment, testes from the rats in all groups were removed, weighed, and subjected to histological examination. Apoptosis in the testes was measured by the TUNEL assay. And the oxidative stress was evaluated by measurement of 8-hydroxy-2'-deoxyguanosine (8-OHdG). RESULTS: Induction of varicocele led to decreasing left testis weight, decreasing spermatogenic cell density significantly (p<0.05). Also it led to increasing apoptotic body counts and increasing concentration of 8-OHdG significantly (p<0.05). However administration of anthocyanin right after varicocele induction prevent this change meaningfully (p<0.05). In group received anthocyanin after four weeks observation following varicocele induction, interestingly, there was no significant difference in testis weight, spermatogenic cell density, apoptotic body count and concentration of 8-OHdG compared to group induced varicocele for eight weeks without anthocyanin administration. CONCLUSIONS: These results suggest that anthocyanin is effective in decreasing the oxidative stress of testis in rat induced varicocele and may be effective in making a healthy sperm in patient of varicocele in early stage. However in patient under way in advanced stage, it is supposed that the anthocyanin cannot help having a protective effect from oxidative stress narrowly unless the condition of oxidative stress by varicocele is corrected. Further studies are needed to better understand the mechanisms and actions of anthocyanin and varicocele, and these studies may lead to the clinical application of anthocyanin in preventing male infertility by varicocele.