GLOMERULAR CAPILLARY WALL--NORMAL AND PATHOLOGIC

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In diabetes mellitus, nephropathy is one of the major complication that ultimately leads to chronic renal failure. Morphologically, it is characterized by remarkable changes in the extracellular matrices (ECM). The biochemical alterations which correlate with these changes include increased expression of type-IV collagen, & decreased synthesis of proteoglycans (PGs). These changes can be induced by elevating the aldohexose (glucose) concentrations in experimental model systems, and are believed to have some relationship with proteinuria in diabetic patients. in addition to these complications in adults, the offsprings of juvenile diabetics have increased incidence of congenital anomalies affecting heart, skeletal and nervous systems, and genitourinary tract, i.e., caudal regression syndrome. Conceivably, the evolution of such developmental defects is related to the high concentration of glucose, which perturbs the functions of morphogenetic regulators, i.e., ECM & cell adhesion molecules, growth factors, their receptors and protooncogenes. In order to test this contention & possible mechanisms involved in the dysorganogenesis of the embryonic kidneys, we propose to carry out experiments outlined under 3 major objectives by utilizing an established murine metanephric culture system. I. Effect of elevated concentrations of hexoses, including glucose, on the morphogenesis of the embryonic kidneys will be monitored by morphometric analyses, immunohistochemical and DNA replication studies. Following which, de novo synthesis and gene expressions of ECM proteins (type-IV collagen, laminin & PGs) & matrix related receptors (integrin alpha3A, alpha:5, alpha6B & betaFGF-R) will be investigated. Biochemical, immunoprecipitation, translational, transcriptional, solution hybridization, RT-PCR & competitive PCR methods will be used. II. Effect of glucose on the intracellular ATP stores, phosphoinositide metabolism, and the tyrosine phosphorylation mechanisms involved in the signal transduction of various growth factors (IGF-I, insulin & EGF), their receptors (INS-R, IGF-IR) & protooncogenes (c-fos, Egr, c-ret & c- ros) will be investigated. III. Eventually, the known and unknown glucose-induced/suppressed transcripts in the metanephric kidneys will be isolated and characterized by cDNA library subtractive hybridization and mRNA differential display methods.