Mechanisms underlying the inhibitory effect of gliflozins on NHE3 activity in the renal proximal tubule

• Grant number: 24/02263-3
• Support Opportunities: Regular Research Grants
• Start date: July 01, 2024
• End date: June 30, 2026
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Agreement: CNPq
• Principal Investigator: Adriana Castello Costa Girardi
• Grantee: Adriana Castello Costa Girardi
• Host Institution: Instituto do Coração Professor Euryclides de Jesus Zerbini (INCOR). Hospital das Clínicas da Faculdade de Medicina da USP (HCFMUSP). Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated research grant: 21/14534-3 - Pleiotropic effects of antidiabetic agents and their pharmacological targets: renoprotective mechanisms beyond glycemic control, AP.TEM
• Associated scholarship(s): 24/11084-5 - Mechanisms underlying the inhibitory effect of gliflozins on NHE3 activity in the renal proximal tubule, BP.JD

Abstract

Sodium/glucose cotransporter 2 (SGLT2) inhibitors, also known as gliflozins, have emerged as potent therapeutic agents for improving glycemic control by suppressing glucose reabsorption in the renal proximal tubule. Originally designed for managing diabetes, gliflozins exhibit additional benefits, such as reducing cardiovascular mortality and heart failure (HF) hospitalization, irrespective of diabetic status. Notably, recent findings from our research group have unveiled that empagliflozin can restore euvolemia and prevent HF progression in non-diabetic rats, partly attributed to its ability to inhibit sodium reabsorption mediated by sodium/hydrogen exchanger isoform 3 (NHE3) in the renal proximal tubule. Nonetheless, the precise molecular mechanisms underlying the NHE3 inhibition by gliflozins remain incompletely understood. This project aims to unravel the mechanisms underlying the effects of gliflozins on NHE3 within the renal proximal tubule. Two primary hypotheses will be scrutinized: (i) Gliflozins exert their inhibitory effect on NHE3 by directly interacting with the sodium binding site of this transporter. Experimental investigations utilizing HEK cells transfected with NHE3 will elucidate whether gliflozins modulate NHE3 activity directly in SGLT2 non-expressing cells. Additionally, in vivo studies will assess the impact of empagliflozin on diuresis, natriuresis, and bicarbonaturia in both wild-type and SGLT2 knockout mice. (ii) Gliflozins disrupt NHE3 activity by destabilizing the NHE3-PDZK1-MAP17-SGLT2 multimeric complex within the renal proximal tubule. Immunoprecipitation experiments and pull-down assays will be conducted to explore whether physical interactions among these proteins are perturbed in the presence of gliflozins. Subsequent in vivo investigations in PDZK1 knockout mice will validate the significance of the NHE3-PDZK1-MAP17-SGLT2 complex in mediating the inhibitory effects of gliflozins on NHE3. Additionally, this project aims to synthesize fluorescent derivatives of D-glucose to probe the functional interaction between NHE3 and SGLT2. The outcomes of this research endeavor hold promise for advancing our understanding of the cardiorenal protective mechanisms of gliflozins. (AU)