MO404: Autophagy and Fibrotic Response is Genetically Determined in Mouse Mesangial Cells

Abstract

Abstract BACKGROUND AND AIMS C57Bl6/J (B6) mouse strain is resistant to several experimental models of renal fibrosis. Mesangial cells play an important role in the pathogenesis of glomerulosclerosis. Also, autophagy dysregulation was recently related to fibrosis. However, it is unclear whether autophagy dysregulation could be genetically determined in mice. Thus, we investigated autophagy and fibrotic response in primary mesangial cells isolated from fibrosis resistant (B6) and fibrosis prone (CBA) inbred mouse strains. METHOD Primary mesangial cells were isolated from 6-weeks old B6 and CBA male mice via magnetic bead separation technique. Glomerular outgrowths were maintained in RPMI medium for 21 days when cells were subcultured and characterized for mesangial markers. Mesangial cells with passage numbers P5 to P8 were seeded onto 24-well plates. Gene expressions were analyzed after 48-h of PBS (CTL) or TGF-β (10 ng/mL) treatment (n = 6/group). Data (mean ± SD) were analyzed using Kruskal–Wallis test. RESULTS Tgfb1 and Col1a1 expressions increased by 2.5-fold and 8-fold in TGF-β treated CBA cells as compared to B6 (P < 0.01), respectively. The expression of Egr1 (early growth response factor-1) that participates in both fibrosis and autophagy increased by 2-fold in TGF-β treated CBA cells but remained unchanged in B6 cells (P < 0.05). Similarly, the expression of core autophagy molecule Lc3b did not change in TGF-β treated B6 cells, but increased by 60% in CBA cells (P < 0.05), accompanied by 1.5-fold increase of Sqstm1/p62. On the other hand, the expression of Mtmr14 that negatively regulates autophagy decreased by 30% in TGF-β treated CBA cells but remained unchanged in B6. CONCLUSION We conclude that genetic background of mice highly influences the TGF-β induced fibrotic and autophagy response of mesangial cells. Our results corroborate previous reports on autophagy dysregulation associated to fibrosis. FUNDING: Hungarian Academy of Sciences BO/00 304/20/5, ÚNKP-21–5-SE-1, NTP-NFTÖ-21-B-0045