Effects of testosterone replacement therapy on metabolic syndrome among Japanese hypogonadal men: A subanalysis of a prospective randomised controlled trial (EARTH study).

We investigated the effects of testosterone replacement therapy (TRT) on metabolic factors among hypogonadal men with a metabolic syndrome. From the study population of the EARTH study, which was a randomised controlled study in Japan, 65 hypogonadal patients with a metabolic syndrome, comprising the TRT group (n = 32) and controls (n = 33), were included in this study analysis. The TRT group was administered 250 mg of testosterone enanthate as an intramuscular injection every 4 weeks for 12 months. Waist circumference, body mass index, body fat volume and blood pressure were measured in all patients at baseline and at 12 months. In addition, blood biochemical data, including total cholesterol, triglyceride (TG), HDL cholesterol, fasting plasma glucose (FPG) and haemoglobin A1c (HbA1c) levels, were also evaluated. Changes in these categories from baseline to 12 months were compared between the TRT and control groups, with significant differences observed in waist circumference, body fat percentage, FPG, TG and HbA1c levels. No significant differences were observed in other parameters. TRT for 1 year was associated with improvements in some metabolic factors among Japanese men with hypogonadism and metabolic syndrome.

Increased prevalence of MEFV exon 10 variants in Japanese patients with adult-onset Still's disease.

Autoinflammatory diseases include a large spectrum of monogenic diseases, e.g. familial Mediterranean fever (FMF), as well as complex genetic trait diseases, e.g. adult-onset Still's disease (AOSD). In populations where FMF is common, an increased MEFV mutation rate is found in patients with rheumatic diseases. The aim of this study was to examine MEFV mutations in Japanese patients with AOSD. Genomic DNA was isolated from 49 AOSD patients and 105 healthy controls, and exons 1, 2, 3 and 10 of the MEFV gene genotyped by direct sequencing. MEFV mutation frequencies in AOSD patients were compared with controls. We found no significant difference in overall allele frequencies of MEFV variants between AOSD patients and controls. However, MEFV exon 10 variants (M694I and G632S) were significantly higher in AOSD patients than controls (6.1 versus 0%). In addition, there was no significant difference between MEFV variant carriers and non-carriers with clinical manifestations, but the monocyclic clinical course of the AOSD disease phenotype was observed less frequently in patients without MEFV variants. AOSD patients had significantly higher frequencies of MEFV exon 10 mutations, suggesting that low-frequency variants of MEFV gene may be one of the susceptibility factors of AOSD.

MEFV gene polymorphisms and TNFRSF1A mutation in patients with inflammatory myopathy with abundant macrophages.

Inflammatory myopathy with abundant macrophages (IMAM) has recently been proposed as a new clinical condition. Although IMAM shares certain similarities with other inflammatory myopathies, the mechanisms responsible for this condition remain unknown. Patients with familial Mediterranean fever (FMF) and tumour necrosis factor receptor-associated periodic syndrome (TRAPS) also often develop myalgia. We therefore investigated the polymorphisms or mutations of MEFV and TNFRSF1A genes in patients with IMAM to identify their potential role in this condition. We analysed the clinical features of nine patients with IMAM and sequenced exons of the MEFV and TNFRSF1A genes. The patients with IMAM had clinical symptoms such as myalgia, muscle weakness, erythema, fever and arthralgia. Although none of the patients were diagnosed with FMF or TRAPS, seven demonstrated MEFV polymorphisms (G304R, R202R, E148Q, E148Q-L110P and P369S-R408Q), and one demonstrated a TNFRSF1A mutation (C43R). These results suggest that MEFV gene polymorphisms and TNFRSF1A mutation are susceptibility and modifier genes in IMAM.

Changes in penile length after radical prostatectomy: effect of neoadjuvant androgen deprivation therapy.

Although reports have shown evidence for penile length (PL) shortening after radical prostatectomy (RP), the association between neoadjuvant androgen deprivation therapy (NADT) and PL after RP has yet to be determined. This study evaluates chronological changes in PL after NADT and RP. Stretched PLs (SPLs) of 143 patients, 41 of whom had undergone NADT, were measured before, 10 days after, and 1, 3, 6, 9, 12, 18, and 24 months after RP. Chronological erectile function and testosterone levels were then evaluated. SPL was shortest 10 days after RP in both the NADT (-) and NADT (+) groups and gradually recovered in length thereafter. SPL in the NADT (-) group was significantly longer than that in the NADT (+) group before RP. However, no significant differences in SPLs were found between both groups 6 months after RP. Although all subjects in the NADT (+) group had testosterone levels of <50 ng/dL before RP, such levels increased after RP. Before RP, the NADT (-) group was found to have significantly better erectile function than the NADT (+) group. However, differences in erectile function between the NADT (-) and NADT (+) groups after RP were not significant. This report is the first to show that among patients with prostate cancer, those who underwent NADT had greater PL recovery after RP than those who did not. Data regarding PL recovery after NADT and RP obtained in this study could be useful for patients with prostate cancer who plan to undergo such procedures.

FK506 augments activation-induced programmed cell death of T lymphocytes in vivo.

FK506 is an immunosuppressive drug that inhibits T cell receptor-mediated signal transduction. This drug can induce immunological tolerance in allograft recipients. In this study, we investigated the in vivo effects of FK506 on T cell receptor-mediated apoptosis induction. Injection of anti-CD3 antibody (Ab) in mice resulted in the elimination of CD4+ CD8+ thymocytes by DNA fragmentation. FK506 treatment significantly augmented thymic apoptosis induced by in vivo anti-CD3 Ab administration. Increased thymic apoptosis resulted in the disappearance of CD4+ CD8+ thymocytes after anti-CD3 Ab/FK506 treatment. DNA fragmentation triggered by FK506 was induced exclusively in antigen-stimulated T cells, since enhanced DNA fragmentation induced by in vivo staphylococcal enterotoxin B (SEB) injection was confirmed in SEB-reactive V beta 8+ thymocytes but not in SEB-nonreactive V beta 6+ thymocytes. In addition to thymocytes, mature peripheral T cells also die by activation-induced programmed cell death. A similar effect of FK506 on activation-induced programmed cell death was observed in SEB-activated peripheral spleen T cells. In contrast, cyclosporin A treatment did not enhance activation-induced programmed cell death of thymocytes and peripheral T cells. Apoptosis is required for the generation and maintenance of self-tolerance in the immune system. Our findings suggest that FK506-triggered apoptosis after elimination of antigen-activated T cells may represent a potential mechanism of the immunological tolerance achieved by FK506 treatment.

Osteoprotegerin inhibits prostate cancer-induced osteoclastogenesis and prevents prostate tumor growth in the bone.

Prostate cancer (CaP) forms osteoblastic skeletal metastases with an underlying osteoclastic component. However, the importance of osteoclastogenesis in the development of CaP skeletal lesions is unknown. In the present study, we demonstrate that CaP cells directly induce osteoclastogenesis from osteoclast precursors in the absence of underlying stroma in vitro. CaP cells produced a soluble form of receptor activator of NF-kappaB ligand (RANKL), which accounted for the CaP-mediated osteoclastogenesis. To evaluate for the importance of osteoclastogenesis on CaP tumor development in vivo, CaP cells were injected both intratibially and subcutaneously in the same mice, followed by administration of the decoy receptor for RANKL, osteoprotegerin (OPG). OPG completely prevented the establishment of mixed osteolytic/osteoblastic tibial tumors, as were observed in vehicle-treated animals, but it had no effect on subcutaneous tumor growth. Consistent with the role of osteoclasts in tumor development, osteoclast numbers were elevated at the bone/tumor interface in the vehicle-treated mice compared with the normal values in the OPG-treated mice. Furthermore, OPG had no effect on CaP cell viability, proliferation, or basal apoptotic rate in vitro. These results emphasize the important role that osteoclast activity plays in the establishment of CaP skeletal metastases, including those with an osteoblastic component.

Identification of 3',5'-cyclic adenosine monophosphate response element and other cis-acting elements in the human androgen receptor gene promoter.

Androgen and androgen receptor (AR) play an important role in sexual differentiation and prostate proliferation. To investigate AR gene transcriptional regulation, a 2.3-kilobase AR gene promoter region was isolated, sequenced, and characterized. Chloramphenicol acetyltransferase (CAT) assay and sequence homology search of AR gene promoter among human, rat, and mouse revealed some potential cis-acting elements, including a GC box, a suppressor region, and a purine-rich element. Deletion analysis and gel retardation assay using a 50-base pair (bp) double-strand purine-rich element showed that this purine-rich element can bind to specific proteins in nuclear extract of LNCaP and HeLa cells and may be essential for AR gene transcription. Furthermore, to investigate the effect of cAMP on AR gene transcription, we treated LNCaP and HeLa cells with 10 mM (Bu)2cAMP after transfection with CAT gene reporter plasmids linked to the AR gene promoter. This treatment induced several folds of CAT activity in LNCaP cells only, and the induction was further confirmed at AR mRNA level by Northern blot analysis and reverse transcription-polymerase chain reaction assay. Deletion analysis of the AR gene promoter showed that a region between 530 bp and 380 bp upstream of AR gene transcription initiation site, which includes one potential cAMP response element (CRE), is responsible for cAMP induction. Gel retardation analysis using this CRE (AR/CRE1) showed that AR/CRE1 can bind to specific proteins in nuclear extract of LNCaP cells, which appears to form a different binding complex compared to somatostatin/CRE.

Androgen receptor: an overview.

The action of androgens in regulating development and growth is mediated by androgen receptor (AR). AR is a member of the steroid hormone receptor superfamily, a class of receptors that function through their ability to regulate the transcription of specific genes. The AR is located in various target tissues, with its levels and activity altered with the onset of various cellular events (e.g., sexual development, malignant transformation). The modulation of AR levels occurs through a number of mechanisms, including transcription, and is regulated by various factors (e.g., androgens). The ability of AR to modulate gene transcription is through its interaction with specific DNA sequences located near or within the target gene promoter. The importance of the AR in reproductive physiology has been emphasized by the finding of AR mutations, leading to a variety of disorders, including testicular feminization syndrome. In this article, we review the structure and function of AR and the role AR plays in the function of the mammalian system.

Immunocytochemical localization of androgen receptor with polyclonal antibody in paraffin-embedded human tissues.

We investigated the immunohistochemical localization of androgen receptor (AR) using a polyclonal antibody for 55 KD recombinant human AR in human tissues fixed with 4% paraformaldehyde solution and embedded in paraffin. Immunoreactive AR was restricted to the nuclei of various tissues. Among the well-known androgen target organs, secretory cells and basal cells of the prostate, spermatogonia, spermatocytes, Sertoli cells and Leydig cells of the testis, epithelial cells of the rete testis, fibroblasts in the whole organ, squamous cells, sweat gland and hair follicle cells of the skin, and hepatocytes of the liver were positive for AR. In addition, smooth muscle cells of the prostate, uterus, urinary bladder, gastrointestinal tract, arteries, and arterioles were strongly immunoreactive for AR. Cardiac muscle and striated muscle of psoas were positive for AR. Acinar cells, ductal cells, and myoepithelial cells of the breast, endocervical and endometrial cells of the uterus, cyto- and syncytiotrophoblast of the chorionic villi, and tubules of the kidney were also positive for AR. Most FSH, LH, and some GH endocrine cells in the anterior and posterior lobes of the pituitary gland, follicular cells of the thyroid gland, and adrenocortical cells were positive for AR. Cells immunoreactive for AR were ubiquitously distributed throughout the entire body. The present study demonstrated the diversity of androgen effects on many target tissues.

Quantitative-analysis of immunohistochemical androgen receptor and ki-67 in prostatic-carcinoma and its relationship to histologic grade.

Androgen receptor and Ki-67 were immunohistochemically investigated in biopsy specimens taken from 50 cases of prostatic carcinoma and the correlation between histologic grade and AR and Ki-67 was examined using a computer-assisted image analysis system. When the immunoreactive intensity of AR of tumor cells was the same or stronger than that of stromal cells in the same section, the tumor cells were defined AR-positive. An immunohistochemical study revealed that AR positive cases-were significantly higher in well differentiated type (86%) and moderately differentiated type (85%) than in poorly (38%) differentiated type. The expression of Ki-67 was significantly higher in poorly and moderately differentiated types than in well, differentiated type. There was a significant correlation between the histologic grade, and AR and Ki-67; however, there was no correlation between the clinical stage and AR or Ki-67.

A case of cytophagic histiocytic panniculitis: successful treatment of recurrent attacks with steroid pulse therapy and oral cyclosporin A.

We report a 35-year-old man, who had been diagnosed with Weber-Christian disease, presented with acute onset of high fever, malaise, jaundice and hepatosplenomegaly with subcutaneous nodules. Laboratory tests showed elevated serum ferritin and liver enzymes, especially lactate dehydrogenase (LDH), with pancytopenia and coagulation abnormalities. Peripheral blood and bone marrow examinations showed erythro-, leuko- and thrombo-phagocytic histiocytes and macrophages. The patient developed the same clinical features seven years ago. Based on diagnosis of cytophagic histiocytic panniculitis, the patient was treated with steroid pulse therapy and oral cyclosporin A. The combination therapy caused a marked improvement in the clinical condition.

Immunolocalization of glutathione-peroxidase and androgen receptors in the rat dorsolateral prostate.

The effects of testosterone (T) and 17 beta-estradiol (E2) on the dorsolateral prostate of castrated rats were investigated by histopathological and immunohistochemical procedures. A significant increase in the prostatic weight of the castrated rats occurred after treatment with testosterone alone, or in combination with E2. Histologically, glandular hypertrophy was observed and fibromuscular stromal proliferation was also seen in the rats receiving the combination treatment. Immunohistochemical localization of glutathione-peroxidase (GSH-PO), an enzyme which effectively reduces the lipid peroxides, was clearly detected in the glandular epithelial of the dorsolateral prostate following testosterone alone or testosterone plus E2-treatment. Therefore, it appeared that GSH-PO protein synthesis in the glandular epithelium of the dorsolateral prostate can be enhanced (induced?) by sex hormones such as testosterone and E2. In the dorsolateral prostate of intact rats, positive staining for androgen receptors (AR) was observed in nuclei of the glandular epithelial cells. In addition, immunodetectable AR decreased within 2 days after castration, but returned to intact levels after administration of testosterone. These findings agree with previous work on the prostate. It is concluded that immunohistochemical analysis is very useful in studying the effects of androgen treatment on the prostate.

[Prospects for molecular research in urological oncology: bladder cancer].

This report consists of a description of our research findings relating to the mechanism of cancer metastasis and target molecules for early diagnosis or cancer therapy. First, we investigated the significance of metastasis-related genes expressed to various extents in three human bladder cancer cell lines using two in vivo models. The relationship between the gene expression pattern and the behavior of cancer cells implicated a loss of E-cadherin expression as a critical factor in facilitating the progression of bladder cancer. Second, we examined the expression of human telomerase reverse transcriptase (hTERT) mRNA in voided urine samples in patients with bladder cancer. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed a higher positive rate as compared with cytological examination, suggesting that the expression of hTERT in urine samples may be a useful diagnostic marker for bladder cancer. Finally, we searched for a molecule to which antisense can be applied as a treatment modality. The 150 kDa oxygen regulated protein (ORP 150), a kind of heat shock proteins, functions as a molecular chaperone in the endoplasmic reticulum. We demonstrated that the adenoviral-mediated antisense ORP150 cDNA transfer resulted in the suppression of vascular endothelial growth factor (VEGF) expression and tumor growth in vivo. In addition, the significant correlation between ORP150 and matrix metalloproteinase 2 (MMP-2) expression was observed in bladder cancer, suggesting that ORP150 functions as a molecular chaperon to MMP-2 secretion for tumor invasion. Anti-sense ORP150 may therefore have a potentially stronger antitumor effect because of its multitargeting capability as a molecular chaperone.

[Production of polyclonal antibody against human androgen receptor and immunohistochemical study of human androgen receptor in prostatic tissues].

We produced polyclonal antibody against human androgen receptor (hAR) by means of immunizing a rabbit with hAR fusion protein that was expressed in E. coli. In Western blot analysis, this antibody, NH27, recognized two protein bands at the site of 110 kDa and 107 kDa in androgen-independent human prostatic cancer cells (PC-3), transfected with full-length hAR expression plasmid DNA and at the site of 114 kDa and 108 kDa in androgen-dependent human prostatic cancer cells (LNCaP). In immunohistochemical examination with NH27, the nuclei of epithelial and stromal cells in human benign prostatic hyperplasia were mainly stained as did with AN1-15, commercially available hAR monoclonal antibody. Titer of NH27, however, was about five times more high than that of AN1-15. In prostatic cancer cells the nuclei were stained with NH27 as did with AN1-15. Intensity of staining was various between the nuclei of cancer cells. The polyclonal antibody, NH27, produced in the present study is useful in investigating the characterization of AR in androgen-dependent and -independent prostatic cancers.

[A male case of primary Sjögren's syndrome with interstitial pneumonitis and interstitial tubulo-nephritis in the absence of dry eye and dry mouth: parotid gland MRI is a useful diagnostic method for Sjögren's syndrome].

Here we report a case of primary Sjögren's syndrome with hilar lymphadenopathy, interstitial pneumonitis and interstitial tubulo-nephritis. A 60-year old man was admitted to our hospital in May 1993 because of general fatigue and fever. He was noted to have hypergammaglobulinemia and had positive antibodies to nuclear antigens since 1990 in the absence of clinical manifestations. Since 9 months before admission, he presented with general fatigue, low grade fever and uveitis. On admission, chest X-ray and CT scan showed bilateral hilar lymphadenopathy and interstitial pneumonitis. The negative results for both serum angiotensin converting enzyme and histological findings of the cervical lymph node and the lung excluded the diagnosis of sarcoidosis. Serological examination exhibited marked elevation of polyclonal IgG level and anti-nuclear antibody, but neither anti-SS-A (Ro) nor anti-SS-B (La) antibody was detected. He did not have symptoms of xerophthalmia and xerostomia. Keratoconjunctivitis sicca was diagnosed by positive Schirmer's and rose bengal tests. His labial gland biopsy demonstrated severe mononuclear cell infiltration around the ducts. MRI findings of the parotid glands revealed heterogeneous and dotted high signal intensity similar to those in fat tissues in the T1- and T2-weighted images. These findings depicted that bilateral parotid gland was extensively destructed and was replaced by lipid tissue. Renal biopsy showed interstitial tubulo-nephritis. On the basis of the above findings, he was diagnosed to have primary Sjögren's syndrome and uveitis. Therefore, MR image of the parotid gland is considered to be a noninvasive and useful method for diagnosis of Sjögren's syndrome.

[HTLV-I infection in primary Sjögren's syndrome--epidemiological, clinical and virological studies].

The HTLV-I seroprevalence rate among the patients with Sjögren's syndrome (SjS, 23.0%) was significantly higher than that among blood donors (3.4%). The age-adjusted summary odds ratio of HTLV-I infection among SjS patients as compared with blood donors was 3.1. The etiologic fraction, i.e., the proportion of SjS in the study population that are attributable to HTLV-I infection, was estimated to be 17.6%. Titers of serum antibodies to HTLV-I in the seropositive SjS patients were significantly higher than those among healthy carriers. IgM class antibodies were commonly detected in sera of SjS patients. Salivary IgA class antibodies were common among seropositive SjS patients, but not in HAM patients or in healthy subjects. The findings strongly suggest that HTLV-I is involved in the pathogenesis of the disease in a subset of patients with SjS in endemic areas.

Impact of pre-treatment prostate tissue androgen content on the prediction of castration-resistant prostate cancer development in patients treated with primary androgen deprivation therapy.

Great advances in tissue androgen analysis using liquid chromatography-tandem mass spectrometry (LC-MS/MS) have made it possible to evaluate the tissue androgen content from a single needle prostate biopsy specimen. In this study, we investigated if pre-treatment androgen content in prostate biopsy specimens could predict their response to primary androgen deprivation therapy (ADT) and future castration-resistant prostate cancer (CRPC). One-hundred and sixty-five prostate cancer patients who received primary ADT were enrolled. They had received multiple core prostate needle biopsy at diagnosis, and an additional one needle biopsy specimen was obtained for tissue androgen determination using LC-MS/MS. The patients' prostate specific antigen (PSA) values were periodically followed during the treatment and patients were determined to have CRPC when their PSA value increased continuously to 25% above the nadir and a 2.0 ng/mL increase. A significant correlation was found between PSA value decline velocity (PSA half-time) after ADT and pre-ADT tissue androgen content. Twenty-three patients were determined to have CRPC. These CRPC patients had a significantly high concentration of tissue T (p < 0.01) and low concentration of tissue 5α-dihydrotestosterone (DHT) (p < 0.01), resulting in a higher tissue T/DHT ratio (p < 0.001). A multivariate Cox proportional hazard model revealed the pre-ADT tissue T/DHT ratio and Gleason score as independent predictors for CRPC development. By using the two statistically significant variables, the relative risk of CRPC development could be calculated. The results of this study suggest that the evaluation of prostate androgen content in a single needle biopsy specimen may be useful to predict future CRPC development after primary ADT. Further studies are required for the clinical application of T/DHT ratio evaluation.

Risedronate prevents persistent bone loss in prostate cancer patients treated with androgen deprivation therapy: results of a 2-year follow-up study.

Androgen deprivation therapy (ADT) for prostate cancer (PCa) causes bone loss. Although we reported previously that risedronate significantly recovers bone mineral density (BMD) for up to 12 months, there have been no reports with longer follow-up periods to date. This study extended our earlier series extending the follow-up period to 24 months. Eligible patients had histologically confirmed PCa without lumbar spine metastasis and underwent ADT. Lumbar spine BMD, urinary deoxypyridinoline (uDPD) and serum bone alkaline phosphatase were measured at 6, 12 and 24 months. Among the total of 96 patients, we analyzed 26 and 18 patients in risedronate administration and control groups, respectively. BMD relative to the young adult mean ratio, uDPD and serum bone alkaline phosphatase of the risedronate administration group recovered significantly after 24 months compared with the control group (P<0.0001, P=0.0001, and P<0.0001, respectively). Transient blurred vision, malaise and vertigo were observed in 1 patient each among the 46 patients treated with risedronate within 28 days after first administration. Oral administration of risedronate is safe and effective for the recovery of ADT-induced bone loss in PCa patients even at 24 months after commencement of treatment.

miR-135a contributes to paclitaxel resistance in tumor cells both in vitro and in vivo.

Cancer cell resistance to paclitaxel continues to be a major clinical problem. In this study, we utilized microRNA (miRNA) arrays to screen for differentially expressed miRNAs in paclitaxel-resistant cell lines established in vitro. We observed concordant upregulation of miR-135a in paclitaxel-resistant cell lines representing three human malignancies. Subsequently, the role of miRNA-135a was evaluated in an in vivo model of paclitaxel resistance. In this model, mice were inoculated subcutaneously with a non-small cell lung carcinoma cell line and treated with paclitaxel for a prolonged period. In paclitaxel-resistant cell lines, established either in vitro or in vivo, blockage of miR-135a sensitized resistant cell lines to paclitaxel-induced cell death. We further demonstrated a correlation between paclitaxel response and miR-135a expression in paclitaxel-resistant subclones that were established in vivo. The paclitaxel-resistant phenotype of these subclones was maintained upon retransplantation in new mice, as shown by decreased tumor response upon paclitaxel treatment compared with controls. Upregulation of miR-135a was associated with reduced expression of the adenomatous polyposis coli gene (APC). APC knockdown increased paclitaxel resistance in parental cell lines. Our results indicate that paclitaxel resistance is associated with upregulation of miR-135a, both in vitro and in vivo, and is in part determined by miR-135a-mediated downregulation of APC.