Intracellular alkalinization by phosphate uptake via type III sodium-phosphate cotransporter participates in high-phosphate-induced mitochondrial oxidative stress and defective insulin secretion

Elevated plasma levels of inorganic phosphate (P_i) are harmful, causing, among other complications, vascular calcification anddefective insulin secretion. The underlying molecular mechanisms of these complications remain poorly understood. We demonstrated the role of P_i transport across the plasmalemma on P_i toxicity in INS-1E rat clonal β cells and rat pancreatic islet cells. Type III sodium-phosphate cotransporters (NaP_is) are the predominant P_i transporters expressed in insulin-secreting cells. Transcript and protein levels of sodium-dependent phosphate transporter 1 and 2 (PiT-1and-2), isotypes of type III NaP_i, were up-regulated by high-P_i incubation. Inpatch-clamp experiments, extracellular P_i elicited a Na⁺-dependent, inwardly rectifying current, which was markedly reduced under acidic extracellular conditions. Cellular uptake of P_i elicited cytosolic alkalinization; intriguingly, this pH change facilitated P_i transport into the mitochondrialmatrix. Increased mitochondrial P_i uptake accelerated superoxide generation, mitochondrial permeability transition (mPT), and endoplasmic reticulum stress-mediated translational attenuation, leading to reduced insulin content and impaired glucose-stimulated insulin secretion. Silencing of PiT-1/2 prevented P_i-induced superoxide generation and mPT, and restored insulin secretion. We propose that P_i transport across the plasma membrane and consequent cytosolic alkalinization could be a therapeutic target for protection from P_i toxicity in insulin-secreting cells, aswell as in other cell types.