Localization of cyclophilin A and cyclophilin C mRNA in murine kidney using RT-PCR

Localization of cyclophilin A and cyclophilin C mRNA in murine kidney using RT-PCR. Cyclosporin A (CsA), which is widely used as an immunosuppressant, has a nephrotoxic side effect. The mechanism of this nephrotoxicity is not well understood; however, recent studies suggest that cyclophilin (cyp) is responsible for mediating the immunosuppressive action of CsA through the interaction with the Ca2+ - and calmodulin-dependent phosphatase, calcineurin. While cyp A mRNA is expressed ubiquitously, cyp C mRNA has been shown to be topically expressed, including in the kidney. We examined: (1) distribution of cyp A and cyp C mRNA in microdissected murine nephron segments, using a combination of reverse transcription and polymerase chain reaction (RT-PCR) techniques, and (2) the effect of CsA administration on cyp C mRNA expression in proximal convoluted tubule. Among the nephron segments examined, large signals for cyp C PCR product were detected in proximal convoluted tubule and proximal straight tubule. Our data showed that the distribution of cyp C mRNA was uneven, and it mainly existed in segments that are relatively sensitive to CsA toxicity. In contrast, cyp A mRNA was found to be distributed almost equally along the nephron segments examined. By CsA administration, the signal for cyp C mRNA PCR product was increased. These results suggest that cyp C may play some role in the renal tubular disorder observed in CsA nephrotoxicity

Regulation of Adrenal Aldosterone Production by Serine Protease Prostasin

A serine protease prostasin has been demonstrated to have a pivotal role in the activation of the epithelial sodium channel. Systemic administration of adenovirus carrying human prostasin gene in rats resulted in an increase in plasma prostasin and aldosterone levels. However, the mechanism by which the elevation of prostasin levels in the systemic circulation stimulated the plasma aldosterone levels remains unknown. Therefore, we examined if prostasin increases the aldosterone synthesis in a human adrenocortical cell line (H295R cells). Luciferase assay using CYP11B2 promoter revealed that prostasin significantly increased the transcriptional activity of CYP11B2. Prostasin significantly increased both CYP11B2 mRNA expression and aldosterone production in a dose-dependent manner. Surprisingly, treatment with camostat mesilate, a potent prostasin inhibitor, had no effect on the aldosterone synthesis by prostasin and also a protease-dead mutant of prostasin significantly stimulated the aldosterone production. A T-type/L-type calcium channel blocker and a protein kinase C (PKC) inhibitor significantly reduced the aldosterone synthesis by prostasin. Our findings suggest a stimulatory effect of prostasin on the aldosterone synthesis by adrenal gland through the nonproteolytic action and indicate a new role of prostasin in the systemic circulation

Impact of home and community-based services on hospitalisation and institutionalisation among individuals eligible for long-term care insurance in Japan

Abstract Background This population-based retrospective cohort study aimed to clarify the impact of home and community-based services on the hospitalisation and institutionalisation of individuals certified as eligible for long-term care insurance (LTCI) benefits. Methods Health insurance data and LTCI data were combined into a database of 1,020 individuals in two farming communities in Hokkaido who were enrolled in Citizen's Health Insurance. They had not received long-term care services prior to April 1, 2000 and were newly certified as eligible for Long-Term Care Insurance benefits between April 1, 2000 and February 29, 2008. The analysis covered 565 subjects who had not been hospitalised or institutionalised at the time of first certification of LTCI benefits. The adjusted hazard ratios (HRs) of hospitalisation or institutionalisation or death after the initial certification were calculated using the Cox proportional hazard model. The predictors were age, sex, eligibility level, area of residence, income, year of initial certification and average monthly outpatient medical expenditures, in addition to average monthly total home and community-based services expenditures (analysis 1), the use or no use of each type of service (analysis 2), and average monthly expenditures for home-visit and day-care types of services, the use or no use of respite care, and the use or no use of rental services for assistive devices (analysis 3). Results Users of home and community-based services were less likely than non-users to be hospitalised or institutionalised. Among the types of services, users of respite care (HR: 0.71, 95% confidence interval [CI]: 0.55-0.93) and rental services for assistive devices (HR: 0.70, 95% CI: 0.54-0.92) were less likely to be hospitalised or institutionalised than non-users. For those with relatively light needs, users of day care were also less likely to be hospitalised or institutionalized than non-users (HR: 0.77, 95% CI: 0.61-0.98). Conclusions Respite care, rental services for assistive devices and day care are effective in preventing hospitalisation and institutionalisation. Our results suggest that home and community-based services contribute to the goal of the LTCI system of encouraging individuals certified as needing long-term care to live independently at home for as long as possible.</p

mRNA distribution and membrane localization of the OAT-K1 organic anion transporter in rat renal tubules

AbstractOAT-K1, a renal organic anion transporter, which mediates methotrexate uptake, was analyzed for mRNA distribution along microdissected nephron segments and the immunolocalization in isolated plasma membranes from rat kidney. By using a reverse transcription-coupled PCR, OAT-K1 mRNA was detected predominantly in the superficial and juxtamedullary proximal straight tubules. Western blotting with antiserum for OAT-K1 revealed that the transporter protein with the apparent molecular mass of 40 kDa was expressed exclusively in the brush-border membranes from rat kidney. These findings suggest that the OAT-K1 is localized in the brush-border membranes of the renal proximal straight tubules

Abnormal development and differentiation of macrophages and dendritic cells in viable motheaten mutant mice deficient in haematopoietic cell phosphatase

In mice homozygous for the ‘viable motheaten’ (mev) mutation, numbers of macrophage progenitor cells, particularly monocytes, were markedly increased in the bone marrow and spleen. Increased mobilization of these precursor cells to peripheral tissues and their differentiation to macrophages were evidenced by striking increases in macrophage numbers. Immunohistochemical double staining of tissue sections and flow cytometry analyses of single cell suspensions from these mice demonstrated CD5 (Ly-1)-positive macrophages in the peritoneal cavity, spleen and other tissues. Ly-1-positive macrophage precursor cells were demonstrated in the peritoneal cavity of the mev mice and developed in the omental milky spots. The development of marginal metallophilic and marginal zone macrophages was poor in the splenic white pulp and related macrophage populations were absent in the other lymphoid tissues. The numbers of epidermal Langerhans cells in the skin and T cell-associated dendritic cells in the thymic medulla, lymph nodes, and the other peripheral lymphoid tissues were decreased. However, increased numbers of dendritic cells accumulated in the lungs, liver, and kidneys. These abnormalities in development and differentiation of macrophages and dendritic cells may be ascribed to the deficiency in haematopoietic cell SHP-1 tyrosine phosphatase or may be a secondary consequence of abnormal microenvironments, (either constitutive or in response to inflammatory stimuli) in the haematopoietic and lymphopoietic organs and tissues of these mice