Recent data have indicated the emerging role of glomerular autophagy in diabetic kidney disease. We aimed to assess the effect of the SGLT2 inhibitor empagliflozin, the DPP4 inhibitor linagliptin, and their combination, on glomerular autophagy in a model of type 2 diabetes. Eight-week-old male db/db mice were randomly assigned to treatment with empagliflozin, linagliptin, empagliflozin-linagliptin or vehicle for 8 weeks. Age-matched non-diabetic db/+ mice acted as controls. To estimate glomerular autophagy, immunohistochemistry for beclin-1 and LAMP-1 was performed. Podocyte autophagy was assessed by counting the volume density (Vv) of autophagosomes, lysosomes and autolysosomes by transmission electron microscopy. LC3B and LAMP-1, autophagy markers, and caspase-3 and Bcl-2, apoptotic markers, were evaluated in renal cortex by western blot. Vehicle-treated db/db mice had weak glomerular staining for beclin-1 and LAMP-1 and reduced Vv of autophagosomes, autolysosomes and lysosomes in podocytes. Empagliflozin and linagliptin, both as monotherapy and in combination, enhanced the areas of glomerular staining for beclin-1 and LAMP-1 and increased Vv of autophagosomes and autolysosomes in podocytes. Renal LC3B and Bcl-2 were restored in actively treated animals. LAMP-1 expression was enhanced in the empagliflozin group; caspase-3 expression decreased in the empagliflozin-linagliptin group only. Mesangial expansion, podocyte foot process effacement and urinary albumin excretion were mitigated by both agents. The data provide further explanation for the mechanism of the renoprotective effect of SGLT2 inhibitors and DPP4 inhibitors in diabetes.Int. J. Mol. Sci. 2020, 21, 2987 2 of 22 dedifferentiation. In its turn, the loss or damage of podocytes causes dysfunction of the filtration barrier and increases albuminuria [2]. Some recent studies have indicated an emerging role of autophagy downregulation in diabetic podocytopathy [4][5][6]. Autophagy is a cellular recycling process involving self-degradation and reconstruction of damaged organelles and proteins [6]. The process is vital for highly differentiated post-mitotic cells, such as neurons and podocytes [7]. A growing body of evidence indicates a critical role of autophagy in maintaining podocyte integrity and renal function [6]. Mice with podocyte-specific deletion of autophagic regulators, such as class III PI3K vacuolar protein sorting 34 (Vps34) and the Atg5 gene, develop early proteinuria, progressive glomerulosclerosis, and renal failure [6]. Accordingly, autophagy is considered a potential therapeutic target for renal protection [8,9].Sodium-glucose cotransporter-2 (SGLT2) inhibitors and dipeptidylpeptidase-4 (DPP4) inhibitors are promising antidiabetic agents introduced into clinical practice in the last decade. The antihyperglycemic effect of SGLT2 inhibitors is mediated by increment of glucosuria, while DPP4 inhibitors realize their activity through an increase in the half-life of incretin hormones. Both SGLT2 and DPP4 inhibitors demonstrated renal protective...

BACKGROUND Modern guidelines recommend sodium-glucose cotransporter-2 (SGLT2) inhibitors as the preferred antihyperglycemic agents for patients with type 2 diabetes and chronic kidney disease. However, the mechanisms underlying the renal protective effect of SGLT2 inhibitors are not fully understood. AIM To estimate the effect of the SGLT2 inhibitor, empagliflozin (EMPA), on the structure of podocytes and nephrin expression in glomeruli in db/db diabetic mice. METHODS We treated 8-wk-old male db/db mice with EMPA (10 mg/kg/d) or vehicle for 8 wk. Age-matched male db/+ mice were included as non-diabetic controls. Parameters of body composition, glycemic and lipid control, and plasma concentrations of leptin, insulin and glucagon were assessed. We evaluated renal hypertrophy as kidney weight adjusted to lean mass, renal function as plasma levels of creatinine, and albuminuria as the urinary albumin-to-creatinine ratio (UACR). Renal structures were studied by light and transmission electron microscopy with a focus on mesangial volume and podocyte structure, respectively. Glomerular nephrin and transforming growth factor beta (TGF-β) were assessed by immunohistochemistry. RESULTS Severe obesity and hyperglycemia developed in db/db mice prior to the start of the experiment; increased plasma concentrations of fructosamine, glycated albumin, cholesterol, leptin, and insulin, and elevated UACR were detected. Mesangial expansion, glomerular basement membrane thickening, and increased area of TGF-β staining in glomeruli were revealed in vehicle-treated mice. Podocytopathy was manifested by effacement of foot processes; nephrin-positive areas in glomeruli were reduced. EMPA decreased the levels of glucose, fructosamine and glycated albumin, UACR, kidney hypertrophy, mesangial expansion, glomerular basement membrane thickening, and glomerular TGF-β staining, alleviated podocytopathy and restored glomerular staining of nephrin. CONCLUSION These data indicate that EMPA attenuates podocytopathy in experimental diabetic kidney disease. The anti-albuminuric effect of EMPA could be attributed to mitigation of podocyte injury and enhancement of nephrin expression.