Sodium Citrate Increases Expression and Flux of Mg ²⁺ Transport Carriers Mediated by Activation of MEK/ERK/c-Fos Pathway in Renal Tubular Epithelial Cells

Abstract

A chronic magnesium deficiency may be one of the causes of lifestyle-related diseases such as hypertension and diabetes. Serum Mg ²⁺ concentration is strictly controlled by the reabsorption pathway in the renal tubules, but little is known about how Mg ²⁺ reabsorption is upregulated. We searched for food compounds which can increase the expression levels of Mg ²⁺ transport carriers including transient receptor potential melastatin 6 (TRPM6) channel and cyclin M2 (CNNM2). Sodium citrate (SC) increased the mRNA levels of TRPM6 and CNNM2 in renal tubular epithelial NRK-52E cells. The SC-induced elevation of TRPM6 was inhibited by U0126, a mitogen-activated protein kinase kinase (MEK) inhibitor, but the CNNM2 was not. SC increased the levels of p-ERK1/2 and p-c-Fos, which were inhibited by U0126. SC induced alkalization of culture medium. Both SC and alkalization enhanced Mg ²⁺ influx, which was inhibited by U0126 and introduction of TRPM6 siRNA. The reporter activity of TRPM6 was increased by SC and alkalization, which was suppressed by mutation in an AP-1-binding site. The SC-induced elevation of p-ERK1/2 and p-EGFR was inhibited by diphenylene iodonium, a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor, and erlotinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor. SC did not change the level of acetyl histone H3, but increased the association of c-Fos with the promoter region of TRPM6. These results suggest that SC increases TRPM6 expression and Mg ²⁺ influx mediated by the activation of NADPH oxidase and an EGFR/ERK/c-Fos pathway in the renal tubules.