P2X₇ receptor-nitric oxide interaction mediates apoptosis in mouse immortalized mesangial cells exposed to high glucose

Abstract

Introduction:  Diabetes mellitus (DM) is a chronic disease characterized by hyperglycemia that leads to diabetic nephropathy (DN). We showed that P2X₇, a purinergic receptor, was highly expressed in DM; however, when oxidative stress was controlled, renal NO recovered, and the activation of this receptor remained significantly reduced. The aim of this study was to assess the influence of NO on the P2X₇ and apoptosis in mouse immortalized mesangial cells (MiMC) cultured in high glucose (HG) medium.

Methods:  MiMCs were cultured with DMEM and exposed to normal glucose (NG), mannitol (MA), or HG. Cell viability was assessed by an automated counter. Supernatants were collected for NO quantification, and proteins were extracted for analysis of NO synthases (iNOS and eNOS), caspase-3, and P2X₇.

Results:  Cell viability remained above 90% in all groups. There was a significant increase in the proliferation of cells in HG compared to MA and NG. NO, iNOS, caspase-3, and P2X₇ were significantly increased in HG compared to NG and MA, with no changes in eNOS. We observed that there was a strong and significant correlation between P2X₇ and NO.

Discussion:  The main finding was that the production of NO by iNOS was positively correlated with the increase of P2X₇ in MCs under HG conditions, showing that there is a common stimulus between them and that NO interacts with the P2X₇ pathway, contributing to apoptosis in experimental DM. These findings could be relevant to studies of therapeutic targets for the prevention and/or treatment of hyperglycemia-induced kidney damage to delay DN progression.

Keywords: Mesangial Cells; Glucose; Apoptosis; Nitric Oxide Synthases; Receptors, Purinergic; Nitric Oxide