Expression of cytochrome P450 3A4 and its clinical significance in human prostate cancer.

OBJECTIVES: To evaluate CYP3A4 expression in human prostrate cancer (PCa) tissues. Enzymes of the cytochrome P450 (CYP) family are key inactivators of testosterone in the liver and prostate. We previously reported that CYP2B6 is a growth-inhibitory and prognostic factor in human PCa; however, the status of CYP3A4 in PCa remains unclear. METHODS: We used immunohistochemistry to analyze CYP3A4 expression in 107 human PCa specimens obtained by radical prostatectomy. Stained slides were evaluated for the proportion and staining intensity of positively stained cells. Total immunoreactivity scores (0-8) were obtained as the sum of the proportion and intensity scores. In addition, we estimated the relationship between CYP3A4 status and clinicopathologic features. RESULTS: CYP3A4 immunoreactivity was identified in the cytoplasm of prostate cells. The CYP3A4 immunoreactive PCa score (3.6+/-2.6) was significantly lower than that of benign epithelium (4.5+/-2.1; P < .0001). In addition, CYP3A4 immunoreactivity correlated inversely with the Gleason score (P < .0001). Decreased CYP3A4 immunoreactivity was significantly related to a poor prognosis in human PCa (P = .0175). CONCLUSIONS: We demonstrated differential CYP3A4 expression in prostatic tissues, indicating that decreased CYP3A4 expression may contribute to the development of PCa.

Estrogen receptor-binding fragment-associated gene 9 expression and its clinical significance in human testicular cancer.

OBJECTIVES: We previously demonstrated that estrogen receptor-binding fragment-associated gene 9 (EBAG9) is a tumor promoting factor in renal cell carcinoma (Ogushi T, Cancer Res. 2005; 65: 3700). Here, we evaluated EBAG9 expression and its clinical significance in normal and malignant human testicular tissues. METHODS: We investigated the expression of EBAG9 in 90 testicular specimens (28 benign testicular tissue and 62 testicular germ cell tumor samples) by immunohistochemistry using rabbit polyclonal anti-EBAG9 antibody. RESULTS: Positive immunostaining of EBAG9 in the cytoplasm was found in 32 (52%) cancerous lesions, whereas the immunoreactivity of EBAG9 was weak in benign testicular tissues. Serum lactate dehydrogenaze (LDH) level was significantly higher in EBAG9-positive cases (715.0 +/- 727.3) compared with the negative cases (221.4 +/- 126.8) (P = 0.0016). The EBAG9-positive cases among the patients with advanced clinical stage (Stage II and III) more frequently belonged to the intermediate or poor risk group in the International Germ Cell Consensus Prognostic Classification System (IGCCPCS), compared with the EBAG9-negative cases (P = 0.0012). CONCLUSIONS: These findings suggest that increased expression of EBAG9 may play a significant role in cancer progression and aggressiveness in testicular germ cell tumors.

EBAG9 is a tumor-promoting and prognostic factor for bladder cancer.

Upregulation of EBAG9 expression has been observed in several malignant tumors such as advanced breast and prostate cancers, indicating that EBAG9 may contribute to tumor proliferation. In the present study, we assess the role of EBAG9 in bladder cancer. We generated human bladder cancer EJ cells stably expressing FLAG-tagged EBAG9 (EJ-EBAG9) or empty vector (EJ-vector), and investigated whether EBAG9 overexpression modulates cell growth and migration in vitro as well as the in vivo tumor formation of EJ transfectants in xenograft models of BALB/c nude mice. EBAG9 overexpression promoted EJ cell migration, while the effect of EBAG9 to cultured cell growth was rather minimal. Tumorigenic experiments in nude mice showed that the size of EJ-EBAG9-derived tumors was significantly larger than EJ-vector-derived tumors. Loss-of-function study for EBAG9 using small interfering RNA (siRNA) in xenografts with parental EJ cells showed that the intra-tumoral injection of EBAG9 siRNA markedly reduced the EJ tumor formation compared with control siRNA. Furthermore, immunohistochemical study for EBAG9 expression was performed in 60 pathological bladder cancer specimens. Intense and diffuse cytoplasmic immunostaining was observed in 45% of the bladder cancer cases. Positive EBAG9 immunoreactivity was closely correlated with poor prognosis of the patients (p = 0.0001) and it was an independent prognostic predictor for disease-specific survival in multivariate analysis (p = 0.003). Our results indicate that EBAG9 would be a crucial regulator of tumor progression and a potential prognostic marker for bladder cancer.

[Hyaline vascular type of Castleman's disease confined to the kidney].

A 59-year-old female was referred to our department for a renal mass which was incidentally found during pneumonia treatment. Computed tomography and magnetic resonance imaging showed a 1.5-cm enhancing mass in the posterior midportion of the left kidney. Subsequently, an open partial nephrectomy was performed on the diagnosis of renal cell carcinoma. Pathological examinations revealed a hyaline-vascular type of Castleman's disease. Castleman's disease confined to the kidney is exceptionally rare. We present this rare case and review the literature concerning this unusual disorder.

[Laser hair removal for urethral hair after hypospadias repair].

A 56-year-old male was admitted for induration of ventral side of the penile shaft. Computed tomography showed a large urethral calculus in the distal urethra. About 50 years previously, he had undergone multi-staged urethroplasty for hypospadias. He had also suffered from recurrent urethral calculi managed by urethrolithotomy 5 and 2 years before the admission. Urethrolithotomy revealed hair-bearing urethral calculus. Instillation of depilating agent containing thioglycolate into the neourethra for preventing hair regrowth was ineffective. Transurethral laser hair removal of neourethra was subsequently performed. All the neourethral follicles were ablated with GaAlAs diode laser (wave length 810 nm; at a power of 15W for 2 seconds) through a side-firing laser fiber. Another three operations were performed for a few regrown hairs at a power of 20-30W. Convalescence was uneventful. The patient is free of hair regrowth except for a hair at five months of follow-up.

[Angiomyolipoma in the renal pelvis].

A 54-year-old male presented with massive hematuria with bladder tamponade. Computed tomography and magnetic resonance imaging showed a 7.5 x 6.5 x 4.5 cm. mass centered in the left renal pelvis, which is composed of macroscopic fat and soft-tissue component. Differential diagnosis included liposarcoma, lipoma and angiomyolipoma. Urothelial carcinoma was also undeniable because urine cytology was equivocal. Subsequently, a left nephroureterectomy was performed. Pathological examinations revealed a renal angiomyolipoma protruding into the renal pelvis. We present a rare case of angiomyolipoma in the renal pelvis and review the literature concerning these unusual findings of this common neoplasm.

Cytochrome P450 2B6 is a growth-inhibitory and prognostic factor for prostate cancer.

BACKGROUND: Cytochrome P450s (CYPs) influence the biological effects of carcinogens, drugs and hormones including testosterones. Among them, Cytochrome P450 2B6 (CYP2B6) plays a critical role in the deactivation of testosterone. In the present study, we examined CYP2B6 expression in human prostate tissues and prostate cancer. METHODS: Immunohistochemical analysis was performed in 98 benign and 106 malignant prostate tissues and patients' charts were reviewed for clinical, pathologic and survival data. We also investigated whether stable expression of CYP2B6 in LNCaP (human prostate cancer cell line) influences cellular proliferation. RESULTS: CYP2B6 was abundantly expressed in the normal epithelial cells compared to the prostate cancer cells. Significant immunostaining of CYP2B6 was found in 75 of 106 samples (71%), in the cytoplasm of cancerous tissue samples. CYP2B6 immunoreactivity was inversely correlated with high Gleason score (P < 0.001). Decreased immunoreactivity of CYP2B6 significantly correlated with poor prognosis (P < 0.0001). Univariate and multivariate hazard analyses revealed a significant correlation of decreased CYP2B6 expression with poor cancer-specific survival (P = 0.0028 and 0.0142, respectively). Furthermore, overexpression of CYP2B6 in LNCaP cells significantly decreased testosterone-induced proliferation. CONCLUSIONS: These results demonstrated that decreased expression of CYP2B6 might play a role in the development of prostate cancer, and be useful as the prognostic predictor for human prostate cancer.

Increased expression of estrogen-related receptor alpha (ERRalpha) is a negative prognostic predictor in human prostate cancer.

The nuclear receptor ERRalpha (estrogen-related receptor alpha) is known to modulate the estrogen-signaling pathway, but the biological significance of ERRalpha in the prostate remains unclear. We investigated the expression of ERRalpha in human prostate tissues and cancer cell lines to evaluate the potential roles of the receptor in prostate cancer (PC). Western blot analysis of ERRalpha was performed in three cell lines of human PC (LNCaP, DU145 and PC-3). The expressions of ERRalpha in cancerous lesions (n = 106) and benign foci (n = 99) of 106 surgically obtained prostate specimens were evaluated by immunohistochemistry. The relationships between the ERRalpha expression and clinicopathological features were evaluated. Western blot analysis using the polyclonal anti-ERRalpha antibody detected a 52 kD band in all three PC cell lines. Positive immunostaining of ERRalpha in the nuclei was found in 73 (69%) cancerous and 47 (47.5%) benign epithelium, whereas the stromal tissues were negative for ERRalpha. The mean immunoreactivity score (IR score) of the cancerous lesions (3.5 +/- 2.6) was significantly higher than that of the benign foci (1.8 +/- 2.1) (p < 0.0001). The IR score of the cancerous lesions significantly correlated with the Gleason score (p = 0.0135). Univariate and multivariate hazard analyses revealed significant correlations between elevated ERRalpha expression and poor cancer-specific survival (p = 0.0141 and 0.0367, respectively). The enhanced expression of ERRalpha might play a role in the development of human PC and serve as a significant prognostic factor for the disease.

Effect of Chinese herbal medicine on overactive bladder.

Gosha-jinki-gan (GJG), a traditional Chinese medicine, is known to be potentially effective for urinary disturbance. For the clinical evaluation of Gosha-jinki-gan, we administered GJG for 6 weeks to elderly male patients with overactive bladder (OAB) and assessed its efficacy and tolerability. In this study, 30 male patients with over 6 months of OAB symptoms had received 2.5 g GJG mixture x 3/day. After 6 weeks of treatment, the efficacy, safety, and tolerability were assessed. We evaluated International Prostate Symptom Score (I-PSS), Overactive Bladder Symptom Score (OABSS), quality of life (QOL), maximal urinary flow rate (Qmax), average urinary flow rate (Qave), incidence of urinary incontinence, and post-void residual before and after treatment. We observed significant improvements in I-PSS (15.2 +/- 1.0 vs. 12.0 +/- 0.9, p < 0.0001), OABSS (7.5 +/- 0.6 vs. 4.9 +/- 0.5, p < 0.0001), and QOL score (4.4 +/- 1.0 vs. 3.3 +/- 1.1, p < 0.0001, Wilcoxon rank sum test). GJG was significantly effective in improving urgency, micturition frequency, nocturia, and urinary incontinence (p < 0.05). However, Qmax, Qave, and post void residual did not significantly change. Mild adverse effects were observed in 3 cases. The symptoms were diarrhea, nausea, and urinary frequency. These data suggest that Gosha-jinki-gan may be a new potential therapeutic agent for OAB without deterioration of voiding function in men with benign prostatic obstruction (BPO).

Periurethral injection of sustained release basic fibroblast growth factor improves sphincteric contractility of the rat urethra denervated by botulinum-a toxin.

PURPOSE: We evaluated the effects of sustained release basic fibroblast growth factor injection in rat urethra denervated by botulinum-A toxin (Wako Life Science, Osaka, Japan). MATERIALS AND METHODS: A total of 30 female Sprague-Dawley rats underwent periurethral injection of 10 U botulinum-A toxin to induce chemical denervation of the urethral sphincter. Leak point pressure in the waking state was determined and a significant decrease in leak point pressure vs that in control rats was confirmed (mean +/- SD 58.7 +/- 6.2 vs 120.7 +/- 13.0 cm H(2)O, p <0.0001). Two weeks later 0, 50 and 200 microg basic fibroblast growth factor incorporating 200 microl gelatin hydrogels in 10 rats each were injected into the urethral sphincter, enabling sustained release of basic fibroblast growth factor for 2 weeks. Four weeks later injection leak point pressure measurement and histological evaluation of the urethra were performed. RESULTS: Leak point pressure in rats with 50 and 200 microg basic fibroblast growth factor injection was significantly higher than in rats with the 0 microg injection (82.7 +/- 9.0 vs 95.1 +/- 6.2 and 119.3 +/- 8.1 cm H(2)O, p = 0.0021 and <0.0001, respectively). Maximum cross-sectional area of the urethral smooth muscle layer in the 50 and 200 microg groups significantly increased compared with that in the urethra in the 0 micro group, which was considered 100% (114.1% +/- 15.8% and 132.5% +/- 13.4%, p = 0.029 and <0.0001, respectively). Similarly the cross-sectional area of the striated sphincter in the 50 and 200 microg groups was greater than the 100% in the 0 microg group (112.3% +/- 15.6% and 124.3% +/- 14.1%, p = 0.069 and 0.0007, respectively). Vascular density in the urethral peri-atrophic zone in the 50 and 200 microg groups was significantly higher than in the 0 microg group (p = 0.027 and <0.0001, respectively). CONCLUSIONS: Sustained release basic fibroblast growth factor injection in the chemically denervated urethral sphincter facilitates regeneration of the urethral muscles and improves sphincteric contractility. Endoscopic periurethral injection of basic fibroblast growth factor incorporating gelatin hydrogels may be an attractive therapy for stress urinary incontinence.

Function of the lower urinary tract in mice lacking alpha1d-adrenoceptor.

PURPOSE: alpha1-Adrenoceptor (AR) blockers are known to relieve not only voiding symptoms, but also storage symptoms in elderly men. We investigated lower urinary tract function in mice lacking alpha1d-AR using frequency/volume analysis and filling cystometry. MATERIALS AND METHODS: A total of 10, 12-week-old female alpha1d-knockout (KO) mice and 10 age matched female wild-type (WT) mice were studied. Each mouse was placed in a metabolic cage connected to a digital scale and personal computer. Under a 12/12-hour dark/light photocycle voiding frequency and volume were recorded for 48 hours. After frequency/volume analysis filling cystometry was performed with the mice awake and without restraint. The expression of alpha1-AR subtype mRNA in the bladder of mice in each group was quantified using real-time polymerase chain reaction. RESULTS: Mean daily voiding frequency +/- SD in alpha1d-KO mice was 9.0 +/- 2.1, significantly lower than 15.9 +/- 5.2 in WT mice (p = 0.0048). Mean volume per void in alpha1d-KO mice was significantly larger than in WT mice (0.24 +/- 0.02 vs 0.16 +/- 0.03 ml, p = 0.0096). Similarly cystometric analysis demonstrated larger bladder capacity (140%, p = 0.0008) and voided volume (146%, p = 0.0048) in alpha1d-KO mice compared with those in WT mice. No significant difference in maximum pressure at void was observed between the 2 groups. In WT mice the amount of alpha1a, alpha1b and alpha1d-AR subtype mRNA in the bladder was 5.2 +/- 0.7, 1.0 +/- 0.1 and 6.3 +/- 0.7 gene copies per ng total RNA, respectively. In contrast, alpha1d-AR transcript was not detectable in alpha1d-KO mice but alpha1a and alpha1b-AR expression was similar to that in WT mice. CONCLUSIONS: The results demonstrate that the alpha1d-AR subtype has an important role in regulating bladder function. They theoretically support a clinical finding that alpha1-blockers with significant affinity for alpha1d-AR are effective for treating storage symptoms associated with benign prostatic obstruction.

Estrogen receptor-binding fragment-associated antigen 9 is a tumor-promoting and prognostic factor for renal cell carcinoma.

The estrogen receptor-binding fragment-associated antigen 9 (EBAG9) has been identified as a primary estrogen-responsive gene in human breast cancer MCF7 cells. A high expression of EBAG9 has been observed in invasive breast cancer and advanced prostate cancer, suggesting a tumor-promoting role of the protein in malignancies. Here we show that intratumoral (i.t.) administration of small interfering RNA against EBAG9 exerted overt regression of tumors following s.c. implantation of murine renal cell carcinoma (RCC) Renca cells. Overexpression of EBAG9 did not promote the proliferation of culture Renca cells; however, the inoculated Renca cells harboring EBAG9 (Renca-EBAG9) in BALB/c mice grew faster and developed larger tumors compared with Renca cells expressing vector alone (Renca-vector). After renal subcapsular implantation, Renca-EBAG9 tumors significantly enlarged compared with Renca-vector tumors in BALB/c mice, whereas both Renca-EBAG9 and Renca-vector tumors were developed with similar volumes in BALB/c nude mice. No apparent difference was observed in specific cytotoxic T-cell responses against Renca-EBAG9 and Renca-vector cells; nonetheless, the number of infiltrating CD8+ T lymphocytes was decreased in Renca-EBAG9 subcapsular tumors. Furthermore, immunohistochemical study of EBAG9 in 78 human RCC specimens showed that intense and diffuse cytoplasmic immunostaining was observed in 87% of the cases and positive EBAG9 immunoreactivity was closely correlated with poor prognosis of the patients. Multivariate analysis revealed that high EBAG9 expression was an independent prognostic predictor for disease-specific survival (P = 0.0485). Our results suggest that EBAG9 is a crucial regulator of tumor progression and a potential prognostic marker for RCC.

Interleukin-2 blocks the antitumour activity caused by depletion of CD25 cells in a murine renal adenocarcinoma model.

OBJECTIVE: To test the effectiveness of antimouse CD25 monoclonal antibody (mAb) against murine renal adenocarcinoma (RENCA) cells, as immunoregulatory/suppressor cells are known to be involved in tumour development in vivo, but the functions of these cells are not yet clear, and eliminating naive CD25 (interleukin-2 receptor alpha)-positive T cells elicits potent immune responses to syngeneic tumours in vivo. MATERIALS AND METHODS: Aliquots of 1 x 10(4) or 1 x 10(5) RENCA cells were implanted into the subcapsule of the left kidney of syngeneic male Balb/c mice. Mice were injected with 125 micro g of antimouse CD25 mAb to deplete CD25(+) cells before RENCA implantation. Then 10(4) units of recombinant human interleukin-2 (rhIL-2) were subcutaneously injected twice daily for 7 days. Fourteen or 25 days later the tumour size was determined by laparotomy, and cells sorted using two-colour flow cytometry. RESULTS: Depletion of naive CD25(+) cells with anti-CD25 mAb and rhIL-2 administration effectively induced anti-RENCA tumour activity in Balb/c hosts. However, co-administration of anti-CD25 mAb and rhIL-2 abrogated this significant suppression of RENCA tumour growth. RENCA implantation reduced the proportion of CD4(+) cells among splenocytes, whereas anti-CD25 mAb treatment increased it. The proportion of CD25(+)CD8(+) cells among splenocytes and that of CD25(+) cells among CD8(+) cells were markedly reduced by co-administration of anti-CD25 mAb and rhIL-2 with RENCA implantation. Both CD4(+) and CD8(+) cells were stained around the remnant microscopic RENCA tumour after anti-CD25 mAb treatment. CONCLUSION: Either depletion of naive CD25(+) cells or rhIL-2 administration suppressed RENCA tumour growth in murine hosts. However, co-administration of anti-CD25 mAb and rhIL-2 abrogated this significant suppression of RENCA tumour growth.