Efficacy of Voclosporin in Maintaining Podocyte Function and Viability in a Rat Streptozotocin Model of Diabetic Nephropathy Public

Abstract

Diabetic nephropathy is a leading cause of end-stage renal disease. Diabetes stimulates the activity and expression of calcineurin; a serine-threonine phosphatase that is a critical regulator of podocyte function and stability. Because calcineurin has been linked to podocyte apoptosis, Synaptopodin degradation, and slit pore function we investigated the effects of Voclosporin, a third-generation calcineurin inhibitor (CNI), on the progression of diabetic nephropathy. Streptozotocin (60 mg/kg) was administered to male Sprague-Dawley rats to induce diabetes. Animals were divided into three cohorts including normal animals (15), diabetic controls (30), and diabetics treated with oral Voclosporin (30, 5.0 mg/kg) by 2X per day gavage feeding. Animals were euthanized at 6, 9, and 12 weeks and the renal cortex was harvested for histologic and western analysis of calcineurin, WT-1 (Wilms tumor-1), and CTGF (connective tissue growth factor) expression. Streptozotocin-treated animals developed hyperglycemia (BG-500-600 mg/dl) within 30 days and proteinuria by Day 84. After 6 weeks, renal cortical expression of calcineurin in diabetic controls increased by 3.5X fold compared to normal controls while treatment with Voclosporin reduced the increase by 50%. Similarly, Voclosporin blocked the rise in glomerular CTGF while blocking a fall in WT-1 and Synaptopodin expression. Sirius Red staining demonstrated that Voclosporin treatment did not lead to an increase in interstitial fibrosis compared to diabetic controls. This study demonstrates that severe diabetes upregulates calcineurin expression in the renal cortex. Treatment of diabetic animals with Voclosporin reduced podocyte depletion and stabilized Synaptopodin without exacerbation of interstitial fibrosis.

Table of Contents

Table of Content

Introduction

Methods

Animal model

Sample preparation

Western blotting

Histology

Immunohistochemistry

Electron microscopy

Statistics

Results

Blood Glucose, Total Protein, Albumin, and Proteinuria at Baseline

Cortical Glomerular and Interstitial Fibrosis: Effect of Voclosporin in Renal Scarring

Calcineurin Expression Following Induction of Diabetes: Effect of Voclosporin

Effect of Diabetes on Podocyte Foot Plate Processes: Effect of Voclosporin

Synaptopodin Expression Following Induction of Diabetes: Effect of Voclosporin

Effect of Diabetes on Glomerular CTGF Expression: Effect of Voclosporin

Effect of Diabetes on Glomerular WT-1 Expression: Effect of Voclosporin

Discussion

Voclosporin Blocks the Diabetes Induced Rise in Calcineurin Expression

Voclosporin Blocks Diabetes Induced Reductions in Synaptopodin Diabetic Glomeruli

Voclosporin Blocks Diabetes Upregulation of Connective Tissue Growth Factor (CTGF)

References

Acknowledgments

Manuscript in Progress

About this Honors Thesis

Rights statement

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School	Emory College
Department	Biology
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Anatomy
Mot-clé	Podocyte Diabetes Voclosporin
Committee Chair / Thesis Advisor	Janet D. Klein, Emory University
Committee Members	Cesar Romero, Emory University Patrick Cafferty, Emory University Jeff Sands, Emory University

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