Tubular hypoxia has a major pathogenic role in diabetic kidney disease (DKD). Hypoxia is in strong association with elevated glucose reabsorption and increased protein O-GlcNAcylation which could contribute to renal fibrosis. We recently showed that SGLT2 inhibitors (SGLT2i) are renoprotective in experimental type 1 diabetes. Considering emerging evidence of proximal tubular involvement in DKD and the major role of SGLT2 in glucose metabolism, here we investigated the direct effects of SGLT2i on hypoxia and O-GlcNAcylation.

Diabetes (D) was induced by streptozotocin in adult, male Wistar rats. Rats were treated for six weeks with dapagliflozin (D+DAPA, 1 mg/bwkg/day). Renal function and fibrosis were evaluated. The effect of hyperglycaemia 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. 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 for 2 hours. HIF1-α, EPO, VEGFA and PAI-1 were measured. Immunocytochemistry (ICC) of HIF1-α was performed.

DAPA improved renal function (creatinine clearance: D: 3.8±0.4 vs. D+DAPA: 8.9±1.0 mL/min; p<0.01) and decreased fibrosis. DAPA minimized hyperglycemia-induced total protein O-GlcNAcylation and OGT, while OGA was elevated in HK-2 cells. Hypoxia-induced HIF-1α elevation was suspended by DAPA treatment. Abolishment of HIF-1α upregulation by DAPA was confirmed by ICC staining. EPO, VEGFA and PAI-1 levels were also increased in hypoxia and DAPA prevented EPO and PAI-1 elevation.

Here we identified a novel mechanism of SGLT2i by which the direct reduction of tubular hypoxia and inhibition of OGT by DAPA results in decreased O-GlcNAcylation in proximal tubular cells. These processes contribute to improved renal function and alleviated kidney fibrosis.

Disclosure

D.B. Balogh: None. J. Hodrea: None. L. Lenart: None. A. Hosszu: None. C. Mezei: None. L. Wagner: None. A.J. Szabo: None. A. Fekete: None.

Funding

Hungarian Academy of Sciences (LP008/2017, OTKA-K112629-FK124491-NN-114607, VKE-2017-00006); Semmelweis University; New National Excellence Program of the Hungarian Ministry of Human Capacities (?NKP-18-3)

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