Abstract 002: Macula Densa SGLT1-NOS1-TGF Pathway -- A New Mechanism for Glomerular Hyperfiltration in Diabetes

Abstract

Listen

Translate article

Glomerular hyperfiltration is a common observation in early diabetes and considered as risk factor for diabetic nephropathy. The mechanisms underlying glomerular hyperfiltration have not been fully clarified. In the present study, we tested a novel SGLT1-NOS1 -TGF (sodium-glucose cotransporter 1 - neuronal nitric oxide synthase – tubuloglomerular feedback) pathway with a hypothesis that tubular glucose is sensed by SGLT1 of the macula densa, which enhances macula densa NOS1 activity and blunts TGF response, therefore increasing glomerular filtration rate (GFR). NO generation by the macula densa, measured by DAF-2 DA in isolated perfused juxtaglomerular apparatus (JGA), increased by 78.4±9.1% when tubular glucose (glucose) was increased from 100 mg/dl to 300 mg/dl (n=4, p<0.01 vs 100 mg/dl). A selective SGLT1 inhibitor KGA-2727 (10 -6 M) blocked this glucose-induced NO generation. Human kidney biopsy tissues were cultured in normal glucose (100 mg/dl) and high glucose (300 mg/dl) media for 30 minutes with hanging drop technique. Protein levels of P-NOS1 ser1417 increased by 63.1±9.2% and P-NOS1 ser847 decreased by 47.4±5.9% (n=4, p<0.01 vs normal glucose) with high glucose. TGF response was significantly reduced from 3.8±0.2 μm to 2.4±0.2 μm (n=4, p<0.05 vs 100 mg/dl), measured in isolated and double perfused JGAs when tubular glucose was increased from 100 mg/dl to 300 mg/dl. The glucose-induced TGF inhibition was blocked in the presence of SGLT1 inhibition (n=4). GFR, measured in conscious mice with a single bolus injection of FITC-inulin, increased by 19.1±3.5% in response to an acute intravenous injection of 50 μl 2M glucose in C57BL/6 mice (n=4, p<0.01 vs baseline). Then, we repeated the experiments in macula densa specific NOS1 KO (KO) mice. Both glucose-induced TGF (ΔP sf was from 7.8±1.3 to 7.3±2.1 mmHg) and GFR (from 223±15.7 to 240±6.9 μl/min) (n=5) were blocked in the KO mice. We conclude that increase of tubular glucose concentration activates NOS1 by phosphorylation of NOS1 ser1417 via SGLT1 in the macula densa, which increases GFR by inhibiting TGF response.