MO264: Selective Endothelin a Receptor Antagonist Atrasentan Attenuates Mesangial Cell Injury, Proteinuria and intra-renal proliferative, inflammatory and Fibrotic Transcriptional Networks in a Rat Model of Mesangioproliferative Glomerulonephritis

Abstract

Abstract BACKGROUND AND AIMS Mesangial cell (MC) activation, characterized by excess proliferation and extra-cellular matrix deposition is the initiating intra-renal event in the pathogenesis of mesangioproliferative glomerulonephritis (MPGN), including IgA nephropathy (IgAN). However, the molecular mechanisms responsible for MC activation and the subsequent podocyte crosstalk resulting in proteinuria have not been well defined. The objective of this study was to determine the role of the endothelin A (ETA) receptor in MC activation, proteinuria and pathogenic intra-renal transcriptional responses in a rat model of MPGN, using atrasentan, a potent and selective ETA antagonist. METHOD MPGN was induced in male Wistar rats by a single IV injection of anti-Thy1.1 antibody (0.5 mg/kg) on day 0. Rats were randomized to receive atrasentan (10 mg/kg, po bid) or vehicle control beginning on Day 1 through Day 7. UPCR was measured on Days 1, 3, 5 and 7; on Day 7, kidneys were fixed, stained and evaluated by light microscopy using a semi-quantitative grading scale by a pathologist, blinded to treatment allocations. RNA-seq was performed on flash-frozen kidney cortex and analyzed using DESeq2 to identify differentially expressed genes followed by gene set enrichment analysis to identify dysregulated transcriptional networks that were cross-validated to the glomerular transcriptome of kidney biopsy samples from IgAN patients (GSE104066). Cell type-specific signatures were derived from a scRNA-seq dataset (GSE171314) from IgAN patients. RESULTS Glomerular injury was characterized histologically by prominent mesangial hypercellularity and matrix expansion, glomerular adhesions, segmental mesangiolysis and glomerulosclerosis, and was significantly attenuated by atrasentan. Tubular injury, including protein casts, tubular degeneration, tubular dilation and interstitial fibrosis, was also observed in the model, although more modest and likely secondary to the glomerular injury, and was also significantly attenuated by atrasentan. The targeted mesangial injury induced by anti-Thy1.1 also resulted in significant proteinuria, which was substantially reduced by atrasentan (Fig. 1). In addition, atrasentan significantly attenuated the increase in kidney weight, likely reflective of the reduced mesangial proliferation, matrix expansion and accompanying inflammation observed histologically. Induction of MPGN in rats was transcriptionally characterized by a downregulation of metabolism gene networks and upregulation of networks associated with proliferation, inflammation and fibrosis, consistent with the hallmark genesets dysregulated in the glomeruli of IgAN patients (Fig. 2). Atrasentan downregulated these intra-renal proliferative, inflammatory and fibrotic transcriptional networks and upregulated metabolism networks (Fig. 2). Analysis using cell-type specific signatures derived from scRNA-seq data from IgAN patients, suggests that atrasentan treatment attenuates anti-Thy1.1-induced gene expression changes in multiple cell types, including mesangial cells, the target cell of injury in MPGN. In addition, although anti-Thy1.1 primarily induces glomerular injury, there was a strong transcriptional signature of failed repair in the proximal tubules, which was attenuated by atrasentan. CONCLUSION This study suggests an important role of the ETA receptor in MC activation, subsequent proteinuria and activation of pathogenic proliferative, inflammatory and fibrotic intra-renal transcriptional networks in MPGN. This further supports the therapeutic potential of atrasentan, a selective ETA receptor antagonist, to attenuate mesangial cell activation, proteinuria and pathogenic intra-renal signaling in MPGNs such as IgAN.