Empagliflozin reduces kidney fibrosis and improves kidney function by alternative macrophage activation in rats with 5/6-nephrectomy

Background: Sodium glucose cotransporter 2 (SGLT2) inhibitors originally developed for the treatment of type 2 diabetes are clinically very effective drugs halting chronic kidney disease progression. The underlying mecha-nisms are, however, not fully understood.Methods: We generated single-cell transcriptomes of kidneys from rats with 5/6 nephrectomy before and after SGLT2 inhibitors treatment by single-cell RNA sequencing. Findings: Empagliflozin treatment decreased BUN, creatinine and urinary albumin excretion compared to placebo by 39.8%, 34.1%, and 55%, respectively (p < 0.01 in all cases). Renal interstitial fibrosis and glomerulosclerosis was likewise decreased by 51% and 66.8%; respectively (p < 0.05 in all cases). 14 distinct kidney cell clusters could be identified by scRNA-seq. The polarization of M2 macrophages from state 1 (CD206-CD68- M2 macro-phages) to state 5 (CD206+CD68+ M2 macrophages) was the main pro-fibrotic process, as CD206+CD68+ M2 macrophages highly expressed fibrosis-promoting genes and can convert into fibrocytes. Empagliflozin remarkably inhibited the expression of fibrosis-promoting (IFG1 and TREM2) and polarization-associated genes (GPNMB, LGALS3, PRDX5, and CTSB) in CD206+CD68+ M2 macrophages and attenuated inflammatory signals from CD8+ effector T cells. The inhibitory effect of empagliflozin on CD206+CD68+ M2 macrophages polari-zation was mainly achieved by affecting mitophagy and mTOR pathways.Interpretation: We propose that the beneficial effects of empagliflozin on kidney function and morphology in 5/6 nephrectomyiced rats with established CKD are at least partially due to an inhibition of CD206+CD68+ M2 macrophage polarization by targeting mTOR and mitophagy pathways and attenuating inflammatory signals from CD8+ effector T cells.Fundings: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Automated summary

Empagliflozin treatment significantly reduced BUN, creatinine, urinary albumin excretion, renal interstitial fibrosis, and glomerulosclerosis. Single-cell RNA sequencing revealed CD206+CD68+ M2 macrophages highly expressed fibrosis-promoting genes, and Empagliflozin inhibited their polarization by affecting mitophagy and mTOR pathways.