SGLT2 Inhibition Alters Methionine Metabolism in Diabetic Kidney Disease

Background: SGLT2 inhibition protects the kidney although underlying mechanisms are incompletely known. We hypothesized that cell-specific metabolic pathways activated by SGLT2 inhibition in diabetic kidney disease (DKD) underlie benefits. Methods: Kidneys harvested from 10-week-old male Sglt2 mutant (MT) and wildtype (WT) mice, fed with normal or high fat diet (HFD, 60% calories from fat) for eight or eighteen weeks, were analyzed. Single cell RNA sequence (scRNA seq) was performed on libraries prepared from whole kidneys. Metabolomic analysis of renal cortex was conducted by Metabolon, Inc. Methionine Adenosyltransferase 2A inhibitor (MAT2Ai, 10 mg/kg BW), an inhibitor of the methionine cycle, was injected into intraperitoneal cavity of WT/MT mice with prior exposure to HFD for eight weeks. Bulk RNA seq and Cleavage under targets and release using nuclease were performed on libraries prepared from renal cortex. Results: HFD-induced obesity was similar in both MT and WT while compensatory hyperphagia was observed in MT. Glucose intolerance occurred in mice fed HFD (WT>MT). Molecular and functional markers of kidney injury including serum creatinine, KIM-1, number of apoptotic cells and albuminuria were higher in WT-HFD. Analysis of scRNA seq showed emergence of a new class of proximal tubular cells (PTC-H), predominantly found in WT-HFD. PTC-H showed increased expression of genes related to epithelial–mesenchymal transition, apoptosis and inflammation. Analysis of metabolomics uncovered differences in WT-HFD vs MT-HFD; metabolites of methionine cycle including SAM were preferentially increased in MT-HFD. MAT2Ai abrogated renal protection with downregulation of NFκB-target genes in MT. Trimethylation of lysine 27 on histone H3 (H3K27me3), a marker of inactive regions of the genome, within genes related to inflammatory signaling were increased in MT-HFD. Especially, H3K27me3 at the promoters of NFκB-target genes, Fos, Jun, and Junb was enhanced in MT-HFD. H3K27me3 was mostly unchanged at the regions with broad peaks, where this mark is already present in WT. A deposition of the H3K27me3 at the SAM-regulated genes in MT-HFD was decreased by MAT2Ai. Conclusion: SGLT2 inhibition prevents the emergence of a new class of PTC with inflammatory phenotype via region-specific enhancement of repressive histone methylation through methionine metabolic modulation in DKD. Funding: Commercial Support - Takeda Science Foundation