SUMMARY:The zucker diabetic fatty (ZDF-fa/fa) rat is one of the attractive models for type II diabetes based on impaired glucose tolerance caused by the inherited insulin-resistance gene fa. Characterization of nephropathy in this model may provide useful insights into the mechanism of the progression of diabetic nephropathy. The present study analyzed the pathophysiology of diabetes and nephropathy, including the process of glomerulosclerosis in this model by biochemical and morphometric analyses. In addition, we conducted studies in podocytes in culture to examine the direct effects of high glucose on podocytes. ZDF-fa/fa rats showed overt diabetes despite hyperinsulinemia as early as 3 months of age. Blood glucose levels increased further with a considerable decrease of insulin levels at 5 months. Glomerular filtration rate (GFR) was significantly elevated until 3 months, but fell to the level seen in lean rats by 7 months. Proteinuria started to rise during the period of increased GFR, and increased further after GFR had fallen to within the normal range. Renal fibronectin, collagen iv, and vascular endothelial growth factor mRNA levels were increased at 7 months. Glomerulosclerosis commenced as early as 5 months of age, and was associated with glomerular hypertrophy and mild mesangial expansion with evidence of accentuated podocyte injury, as revealed by increased expression of desmin. Electron microscopy suggested that degeneration of podocytes and the development of tuft adhesions were responsible for the glomerular sclerosis in this model. In addition, glomeruli from the diabetic rats showed up-regulation of the cyclin kinase inhibitors, p21 and p27. Further studies suggested that the increase in p27 expression was predominantly caused by podocytes, because predominant immunolocalization of p27 in podocytes in diabetic rats and high glucose medium induced cell hypertrophy accompanied by p27 up-regulation in differentiated podocyte cell lines. In conclusion, progressive diabetic nephropathy in ZDF-fa/fa rats is associated with evidence of podocyte injury. High concentrations of ambient glucose induced podocyte hypertrophy and stress in vitro, suggesting that the podocyte is a likely target of the diabetic milieu. (Lab Invest 2002, 82:25-35).

Abstract-An antibody was raised in rabbits against SFFLRNPSEDTFEQF peptide, which is an NH 2 -terminal peptide of the thrombin-cleaved rat thrombin receptor. In vitro, the antibody inhibited rat smooth muscle cell proliferation but had no effect on rat platelet aggregation or clotting time. These data indicate that the antibody is a specific blocker of the thrombin receptor-signaling pathway in rat smooth muscle cells but does not work as a blocker in rat platelets, suggesting the existence of a second thrombin receptor in the platelets. Using an in vivo balloon catheter-induced injury model in rats, we examined the effect of the anti-rat thrombin receptor IgG on intimal smooth muscle cell accumulation 2 weeks after angioplasty. Analysis of the ratio of intimal to medial cross-sectional areas showed that injection of immune IgG resulted in 43.7% and 53.1% reduction (PϽ0.01) of neointimal smooth muscle cell accumulation compared with saline and nonimmune IgG treatment, respectively. Moreover, the injection of immune IgG caused a significant decrease of thrombin receptor mRNA expression and also 40.5% and 43.0% decreases (PϽ0.01) of the proliferating cell nuclear antigen (PCNA) index in the intima compared with the PCNA index after saline and nonimmune IgG treatment, respectively. The suppression of the PCNA index was also observed in the immune IgG-treated group at an early stage after angioplasty. These results suggest that thrombin receptor activation is involved in the proliferation and accumulation of neointimal smooth muscle cells induced by balloon injury. (Circ Res. 1998;82:980-987.)Key Words: smooth muscle cell proliferation Ⅲ thrombin Ⅲ thrombin receptor Ⅲ restenosis Ⅲ angioplasty T hrombin, a serine protease derived from its precursor, prothrombin, plays an important role in cellular responses.1,2 Stimulation of vascular endothelial cells by thrombin leads to the release of several vasoactive substances, such as prostacyclin, platelet-activating factor, and PDGF, increases [Ca 2ϩ ] i and also induces translocation of the neutrophiladhesive molecule (P-selectin) to the endothelial cell surface.3,4 Thrombin also has a strong mitogenic effect on vascular SMCs.5 These diverse cellular responses to thrombin may orchestrate the hemostatic, inflammatory, and proliferative responses to vascular injury. 6 In vitro and in vivo studies have implicated bFGF and PDGF as well as thrombin in these responses.7-12 However, thrombin may be especially important as a mediator of vascular lesion formation, since it is present after balloon catheter injury. 10The thrombin receptor has been cloned, and it has been proposed that thrombin binds to its receptor and cleaves it after Arg41 in the receptor's NH 2 -terminal portion, thereby exposing a new NH 2 terminus, which functions as a tethered ligand for the receptor. 13 The thrombin receptor can mediate the mitogenic effect of thrombin, 5 and increased thrombin receptor expression was observed in neointimal cells throughout vascular lesion formation. 12 However, the role of...

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