Antiferroptotic effect of hydrogen-sulfide donors on the pancreas and liver in type 1 diabetic mice

Abstract

The pathogenesis of diabetes mellitus (DM) involves progressive cell death in islets and diabetes-targeted tissues. Ferroptosis, an iron-driven form of programmed cell death, has emerged as an important antidiabetic target. DM is also associated with decreased systemic and peripheral hydrogen sulfide (H2S) levels that contribute to DM pathology. Although recent findings indicate that H2S protects cells from ferroptosis, its role in DM prevention/amelioration is not elucidated. Thus, we aimed to examine the anti-ferroptotic potential of the donors of H2S and its reactive species on pancreas and early stages liver pathology in diabetic mice. DM was induced in C57BL/6 mice with streptozotocin (STZ, 65 mg/kg/day for 3 days). Synthetic H2S donors (1) GYY4137, (2) sodium sulfide (Na2S4) and (3) cysteine-3-sulfide (Cys-S3) were administered intraperitoneally, starting with the first dose of STZ for 3 weeks. Histologic examinations of the pancreas and liver and immunohistochemical analyses of insulin, 4-hydroxynonenal (4HNE) and glutathione (GSH) peroxidase 4 (GPX4) were performed. The GSH level and antioxidative enzymes activities were measured spectrophotometrically. The

levels of ferroportin (FPN), ferritin (FTH) and ACSL4 were analysed by Western blot. The level of heme oxygenase- 1 (HO-1) was measured by ELISA.

The results suggest that treatment with H2S donors, particularly Cys-S3, prevents the development of DM, as evidenced by lower glucose levels and DM incidence compared to untreated diabetic mice. In the pancreas, treatment with H2S donors improved the overall morphology, islet size, insulin and GPX4 expression in the islets and the amount of GSH. In the liver of diabetic mice early pro-ferroptotic phenotype seen through ferroptosis markers: increased content of free iron and lipid peroxidation product, 4HNE and expression of the main

ferroptosis driver - ACSL4 were reduced with H2S donors. H2S donors also restored the expression of the iron- regulatory proteins FPN1 and FTH1, HO-1, and the activity of GSH-S-transferase and catalase. The beneficial

effects of the H2S donors, particularly Cys-S3, were also evident in improved overall liver appearance, reduced fibrosis and an increased amount of binucleated hepatocytes. These data clearly show the strong antiferroptotic potential of H2S donors and suggest a new approach for the prevention/treatment of diabetes and diabetic diseases based on targeting ferroptosis via H2S signalling.