A nephritogenic antigen for acute poststreptococcal glomerulonephritis (APSGN) was isolated recently from group A streptococcus and termed nephritis-associated plasmin receptor (NAPlr). In vitro experimental data indicate that the pathogenic role of NAPlr occurs through its ability to bind to plasmin and maintain its proteolytic activity. However, the mechanism whereby this antigen induces glomerular damage in vivo has not been fully elucidated. Renal biopsy tissues from 17 patients with APSGN, 8 patients with rapidly progressive glomerulonephritis, and 10 normal kidneys were analyzed in this study. Plasmin-like activity was assessed on cryostat sections by in situ zymography with a plasmin-sensitive synthetic substrate. Serial sections were simultaneously assessed for NAPlr deposition by immunofluorescence staining. Glomerular plasmin-like activity was absent or weak in normal controls and in patients with rapidly progressive glomerulonephritis, although tubulointerstitial activity was occasionally detected. Prominent glomerular plasmin-like activity was found in patients who had APSGN and in whom glomerular NAPlr was positive, whereas it was absent or weak in patients who had APSGN and in whom glomerular NAPlr was negative. The distribution of glomerular plasmin-like activity was identical to that of NAPlr deposition but was generally different from that of fibrin(ogen) deposition as assessed by double staining. The activity was abolished by the addition of aprotinin to the reaction mixture but was not altered by the addition of a matrix metalloprotease inhibitor, a cysteine protease inhibitor, or inhibitors of plasminogen activators. Thus, upregulated glomerular plasmin-like activity in relation to NAPlr deposition in APSGN was identified. This result supports the nephritogenic character of NAPlr and offers insight into the mechanism whereby this antigen induces nephritis.

Cell electrophoretic mobility (EPM) can be used to characterize individual cells. The purpose of this study is to establish reproducible and reliable cell EPM values obtained using microcapillary electrophoresis (microCE) chips. We studied cell electrophoresis on microCE chips through the comprehensive measurement of EPM and zeta potential. The inner wall of microchannels in microCE chips was coated with three kinds of reagents, namely bovine serum albumin (BSA), gelatin, and 2-methacryloyloxyethylphosphorylcholine (MPC) polymer to prevent nonspecific adhesion and interaction between cells and the inner wall. Electrophoresis was conducted in phosphate-buffered saline (pH 4-9) using erythrocytes extracted from sheep whole blood. Electroosmotic flow (EOF) mobility was measured using noncharged particles, and then the true EPM was calculated by subtracting the EOF mobility from the electromigration. MPC polymer coatings in microCE chips reduced the zeta potential of the inner wall and fully prevented nonspecific adhesion. EPM data obtained using microCE chips were almost the same and reproducible over a wide range of pH irrespective of the coating reagent used. In conclusion, reliability in the measurement of cell EPM using microCE chips was realized.

The profibrotic effect of plasminogen activator inhibitor-1 (PAI-1) in renal fibrosis is widely recognized, but its mechanism remains controversial especially in chronic progressive kidney disease. In the present study, pioglitazone (Pio) and candesartan (CD), which are reported to inhibit PAI-1, were administered to spontaneously hypercholesterolemic (SHC) rats, a model of chronic progressive kidney disease. Therapeutic effects and effects on the intrarenal plasmin cascade were examined. Eight-wk-old SHC rats were used as controls. Oral administration of vehicle alone, Pio, or CD was performed starting at 8 wk of age and was continued for 24 wk. The degree of renal fibrosis was evaluated by sirius red staining of kidney sections and by total collagen assay of renal homogenates. The renal PAI-1 protein level was assessed by Western blotting, and plasmin activity was analyzed by chromogenic assay and casein gel zymography. Urinary protein and blood urea nitrogen were significantly increased in the vehicle-treated group, but the increase was attenuated in the Pio- and CD-treated groups. This correlated well with the degree of fibrosis as assessed by sirius red staining and total collagen assay. The PAI-1 protein level was also increased significantly in the vehicle-treated group, and the increase was attenuated in the Pio- and CD-treated groups. Despite the presumed plasmin-inhibitory function of PAI-1, plasmin activity changed in parallel with PAI-1. These results suggest that Pio and CD inhibit PAI-1 and exert renoprotective effects against chronic progressive renal disease, but its action is independent of the regulatory function on plasmin activity.

Twin and family studies had shown that genetic factors are important determinants of bone mass. Multiple genes might be involved. One candidate gene, the reversion-induced LIM gene (RIL), is a PDZ and LIMdomain-containing protein and has been localized within the cytokine cluster of chromosome 5 (5q31.1). In a genetic study of 370 adult Japanese women, we investigated the correlation between radial bone mineral density (BMD) and a genetic variation ()3333T fi C) of the 5'-flanking region of RIL gene. A significant association was identified between the RIL variation )3333T fi C and radial BMD (r=0.15, P=0.003). The variation of the RIL locus may be an important determinant of osteoporosis.Keywords Single-nucleotide polymorphism AE RIL AE Bone mineral density AE Association study IntroductionOsteoporosis is a multi-factorial common disease that is characterized by reduced bone mass and increasing risk of fracture. Genetic and environmental factors play important roles in the determination of bone mineral density (BMD) (Hirota et al. 1992;Suleiman et al. 1997;Pocock et al. 1987;Krall and Dawson-Hughes 1993).Previous studies have examined associations of candidate gene polymorphisms with BMD. Examples are the genes encoding the vitamin D receptor, the estrogen receptor, the apolipoprotein E and type I collagen, combinations of which may determine individual BMD levels (Morrison et al. 1994;Melhus et al. 1994;Greenfield and Goldberg 1997;Kobayashi et al. 1996;Shiraki et al. 1997;Uitterlinden et al. 1998). In addition to these makers, numbers of unidentified polymorphic genes may participate in determining the bone mass of an individual. Candidates might be genes involved in cytokine-signaling pathways, the hormonal regulation of calcium balance and bone mineral, or the cellular function of bone cells.One possible candidate gene localized within the cytokine cluster of chromosome 5 (5q31.1) is the reversion-induced LIM gene (RIL), whose mRNA expression in human bone marrow stromal cells has been strongly detected in a previous study (Bashirova et al. 1998). Although its exact function is not defined as yet, an involvement of osteoblast development/function is implicated.In this study, we have carried out a correlation study of genetic variations in RIL gene for radial BMD levels. Involvement of an RIL gene polymorphism was tested with respect to the regulation of BMD. Subjects and methodsSubjects DNA samples were obtained from peripheral blood of 370 adult Japanese women (Shinohara et al. 2001). BMD was measured in all of these subjects. Mean ages and body mass indices (BMI) with standard deviations (SD) were 58.4±8.6 years (range:

In this cross-sectional study, we investigated the effect of daily walking steps on ultrasound parameters of the calcaneus in elderly Japanese women. The subjects were 143 community-dwelling elderly women aged 61-87 years (mean age 71.4+/-5.5 years). The speed of sound (SOS), broadband ultrasound attenuation (BUA) and the stiffness index (Stiffness) of the calcaneus were measured. Walking steps were recorded using a pedometer for 7 consecutive days as an outcome measure of physical activity. In univariate analyses, steps/day significantly decreased with aging. SOS, BUA and Stiffness showed negative correlations with age and positive correlations with weight. Linear relationships were not seen between any of the ultrasound parameters and daily walking steps. Then, the ultrasound parameters were adjusted for age and/or weight using multiple regression models, and the relationships between the adjusted ultrasound measurements and walking steps were examined using quadratic regression models. Walking activity up to approximately 12,000 steps/day was positively associated with the adjusted ultrasound measurements, above which additional walking steps had no positive effect. We conclude that daily walking steps may be suitable for evaluating the relationship between ultrasound parameters and physical activity in elderly women, but further research is needed to confirm the effect of daily walking steps on the rate of bone loss.