Date of Graduation
School of Medicine
Physiology, Pharmacology & Neuroscience
Robert W. Brock.
C-peptide, a by-product of insulin biosynthesis, has been shown to ameliorate diabetes-induced renal impairment. Yet, the mechanisms underlying this protective benefit remain unclear. Our studies have shown that C-peptide improved renal peritubular capillary blood flow and reduced vascular oxidants. NADPH (reduced nicotinamide adenine dinucleotide phosphate), appears to be critical to this effect via the actions of numerous vasoprotective systems. Further, our experiments in type I diabetic mice resulted in a significant reduction in renal endothelial NADPH that is subsequently restored with C-peptide. We hypothesized that C-peptide provides protection to renal cortical endothelial cells during type I diabetes by restoring the activity of glucose-6-phosphate dehydrogenase (G6PD), the principal source of NADPH synthesis. Our data show that renal cortical endothelial cells (RCE) from diabetic mice have diminished G6PD activity and their treatment with C-peptide restores its activity back to control levels. These changes in G6PD activity occurred with concomitant alterations in NADPH. Using 2-D gel electrophoresis of RCE lysate coupled with immunoblotting for G6PD, we demonstrate that diabetic RCEs have a significant increase in G6PD post-translational modification (PTM). C-peptide treatment reduced the magnitude of this PTM in diabetic RCEs, which occurred concomitant to restored G6PD activity. These results suggest that improving the activity of endothelial G6PD, by preventing its PTM, may be a potential mechanism by which C-peptide confers protection to renal cortical endothelial cells during type I diabetes. Hyperglycemia-mediated microvascular damage has been proposed to originate from excessive generation of mitochondrial superoxide in endothelial cells and is the suggested mechanism by which the pathogenesis of diabetes-induced renal damage occurs. To determine whether C-peptide affords protection to renal microvascular endothelial cell mitochondria during hyperglycemia we exposed conditionally immortalized murine renal microvascular endothelial cells (MEC) to low or high glucose (25 mM) media with either C-peptide (6.6 nM) or its scrambled sequence control peptide for 24- or 48-hours. Respiratory control ratio, a measure of mitochondrial electrochemical coupling, was significantly higher in high glucose treated renal MECs treated with C-peptide than those of high glucose alone. C-peptide also restored high glucose-induced renal MEC mitochondrial membrane potential changes back to their basal low glucose state. Moreover, C-peptide prevented the excessive mitochondrial superoxide generation and concomitant reductions in mitochondrial complex I activity that are mediated by the exposure of the renal MECs to high glucose. Together, these data demonstrate that C-peptide protects against high glucose- induced generation of mitochondrial superoxide in renal MECs via restoration of basal mitochondrial function. Although interest in the physiologic benefits of C-peptide has persisted for more than two decades, C-peptide has yet to make its way into standard treatment regimens for various diabetic complications. The findings from our work have provide proof-of-principle evidence in support of the inclusion of C-peptide to the existing therapeutic regimen for treatment of diabetic complications, specifically those related to diabetes-induced renal impairment.
Vejandla, Himani, "C-peptide Improves Hyperglycemia-induced Endothelial Redox Balance by Preventing Mitochondrial Oxidative Stress and Enhancing NADPH Synthesis" (2012). Graduate Theses, Dissertations, and Problem Reports. 3565.