A Novel Potential of SGLT2 Inhibition: Antifibrotic Effect in Diabetes

Abstract

Background and aimsIdentification of novel therapeutic strategies that reduce the risk of diabetic kidney disease (DKD) is a research priority. Clinical data suggest that improved renal outcomes by sodium‐glucose cotransporter 2 inhibitors (SGLT2i) are partly beyond their glucose lowering effects; however, mechanisms are still elusive. Enhanced glucose reabsorption in diabetes leads to tubular hypoxia playing part in fibrogenesis. Hyperglycemia is in strong association with increased protein O‐GlcNAcylation, a post‐translational modification contributing to renal fibrosis. Considering the proximal tubular involvement in DKD pathogenesis and the key role of SGLT2 in glucose metabolism, here we investigated the effects of SGLT2i on tubular hypoxia and O‐GlcNAcylation.MethodsDiabetes (D) was induced by streptozotocin (65 mg/bwkg, ip.) in adult, male Wistar rats. Following the onset of diabetes rats were treated for six weeks with dapagliflozin (D+DAPA, 1 mg/bwkg/day, po.). Metabolic parameters and renal function were evaluated. Novel urinary biomarkers of extracellular matrix remodeling (Pro‐C3, uC3M, tumstatin) and profibrotic growth factors (TGF‐β, CTGF, PDGF) were determined. Histological evaluation of glomerular damage (PAS) and tubulointerstitial fibrosis (Masson’s trichrome, Picrosirius red, fibronectin) were performed. The effect of hyperglycemia was tested in human proximal tubular epithelial cells (HK‐2) kept under normal glucose (5.5 mM), high glucose (35 mM) or high mannitol (osmotic control, 35 mM) conditions for 24 hours. HG cells were treated with 10 μM DAPA. O‐GlcNAc, O‐GlcNAc transferase (OGT) and O‐GlcNAcase (OGA) were measured. To test the effect of hypoxia cells were treated with 10 μM DAPA and were placed in a hypoxic chamber (1% O2) for 2 hours. HIF‐1α, EPO, VEGFA and profibrotic factors were measured. Immunocytochemistry (ICC) staining for HIF‐1α was performed.ResultsDAPA decreased blood glucose levels (D: 37±2.7 vs. D+DAPA: 18±5.6 mmol/L; p<0.05) and improved renal function (creatinine clearance: D: 3.8±0.4 vs. D+DAPA: 8.9±1.0 mL/min; p<0.01). Subsequently novel fibrosis biomarkers, profibrotic growth factor expressions and renal fibrotic tissue accumulation were reduced. DAPA minimized hyperglycemia‐induced total protein O‐GlcNAcylation in HK‐2 cells. Hypoxia‐induced HIF‐1α elevation was suspended by DAPA treatment. EPO, VEGFA and profibrotic factor levels were also increased in hypoxia and DAPA prevented EPO and CTGF elevation.ConclusionsThese data highlight the role of ameliorated O‐GlcNAcylation and diminished tubular hypoxia as important benefits of SGLT2i treatment. Our results support the link between glucose toxicity, tubular hypoxia and fibrosis, a vicious trio that could be targeted by SGLT2i. All these mechanisms are important parts in the puzzle of the complex system behind the protective effect of SGLT2i.Support or Funding InformationOTKA‐FK124491, VKE‐2017‐00006, FIKP, UNKP‐19‐3‐III‐SE‐6, EFOP‐VEKOP‐16‐2017‐0009