| Summary: | 博士 === 長庚大學 === 臨床醫學研究所 === 95 === Diabetic nephropathy caused by intensive renal cell depletion and extracellular matrix deposition is a leading cause of end-stage renal diseases in Taiwan. While high glucose and advanced glycation end products (AGEs) have been report as deleterious factors for the pathogenesis of diabetic kindeys, the early cellular signaling events by which these molecules induce fibrosis of mesangial cells are not well defined. We first hypothesized that reactive oxygen radicals might act as early high glucose and AGEs-reponsive mediators to induce fibrosis signaling and matrix accumulation in mesangial cells. We found elevations of TGF-β1 and fibronectin levels in mesangial cell cultures followed high glucose and AGEs stresses, which induced Ras- and NADPH oxidase-dependent superoxide and progressively activated cytosolic ERK and nuclear c-Jun, which was abrogated by Manumycin A and PD98059. Exogenous administration of SOD-PEG alleviated diabetes induction of superoxide, urinary albumin excretion, 8-hydroxy-2’-deoxyguanosine, TGF-β1 and fibronectin expression in renal glomerular mesangial cells in diabetic rats. These findings indicate that superoxide-meidated fibrosis response of mesangial cells regulated the pathogenesis of diabetes-induced early renal injury.
Canonical Wnt/β-catenin signaling molecules have been reported to modulate renal tissue morphogenesis and function. However, the biological role of Wnt/β-catenin signaling pathway in high glucose-stressed glomerular mesangial cells is not well defined. We further hypothesized that Wnt signaling might be involved in regulating the fate of high glucose-stressed mesangial cells. We found that high glucose down-regulated Wnt4 and Wnt5a expression and the subsequent nuclear translocation of β-catenin, while it increased GSK-3β, caspase-3 activities and apoptosis of glomerular mesangial cell. Suppression of GSK-3β activation or increase in nuclear β-catenin by transfecting Wnt4 or Wnt5a or stable β-catenin (S33Y) reversed Akt activation and reduced the high glucose-mediated caspase-3 cleavage and cell apoptosis. Pharmacological inhibition of GSK-3β by recombinant Wnt5a or BIO or LiCl increased Akt phosphorylation, β-catenin translocation and abrogated high glucose-mediated proapoptotic activities. Exogenous BIO treatment reduced phospho-Ser9-GSK-3β, β-catenin expression and apoptosis of cells adjacent to glomeruli in diabetic kidneys and attenuated urinary protein secretion in diabetic rats, suggesting that canonical Wnt/β-catenin signaling is required for protecting glomerular mesangial cells from high glucose-mediated cell apoptosis
Mesangial cell responded to high glucose by raising oxidate stress and down-regulating Wn/β-catenin signaling and progressively induced mesangial cell dysfunction. We futher defined the crosstalk between reactive oxygen radicals and Wnt/β-catenin signaling in modulating apoptosis of high glucose-stressed mesangial cells. Scavenging superoxide by SOD or DPI abrogated high glucose-induced loss of Wnt5a/β-catenin expression, activation of GSK-3β and apoptotic activities of mesangial cells. Transfection of dominant negative Ras(S17N) and Rac1(T17N) and kinase-inactive GSK-3β mutant impaired high glucose-induced superoxide burst, apoptotic programs and reversed Wnt5a/β-catenin signaling. Exogenous SOD treatment abrogated reactive oxygen radical synthesis, apoptosis and increased Wnt5a/β-catenin immunoreactivities of cells adjacent to renal glomeruli and urinary protein secretion in diabetic rats.
Taken together, we provide evidence that Ras induction of oxidative stress induced loss of Wnt/β-catenin-regulated survival and activated ERK-dependent fibrosis factor expression in mesangial cells exposed to high glucose stress. Modulation of redox reactions or Wnt/β-catenin singlaing is presumably benefit for protecting mesangial cell in renal tissue from the deleterious actions of high glucose on.
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