miR-155-5p Promotes Oxalate- and Calcium-Induced Kidney Oxidative Stress Injury by Suppressing MGP Expression

Abstract

Oxalate and calcium are the major risk factors for calcium oxalate (CaOx) stone formation. However, the exact mechanism remains unclear. This study was designed to confirm the potential function of miR-155-5p in the formation of CaOx induced by oxalate and calcium oxalate monohydrate (COM). The HK-2 cells were treated by the different concentrations of oxalate and COM for 48 h. We found that oxalate and COM treatment significantly increased ROS generation, LDH release, cellular MDA levels, and H₂O₂ concentration in HK-2 cells. The results of qRT-PCR and western blot showed that expression of NOX2 was upregulated, while that of SOD-2 was downregulated following the treatment with oxalate and COM in HK-2 cells. Moreover, the results of miRNA microarray analysis showed that miR-155-5p was significantly upregulated after oxalate and COM treated in HK-2 cells, but miR-155-5p inhibitor treatment significantly decreased ROS generation, LDH release, cellular MDA levels, and H₂O₂ concentration in HK-2 cells incubated with oxalate and COM. miR-155-5p negatively regulated the expression level of MGP via directly targeting its 3'-UTR, verified by the Dual-Luciferase Reporter System. In vivo, polarized light optical microphotography showed that CaOx crystal significantly increased in the high-dose oxalate and Ca²⁺ groups compared to the control group. Furthermore, IHC analyses showed strong positive staining intensity for the NOX-2 protein in the high-dose oxalate and Ca²⁺-treated mouse kidneys, and miR-155-5p overexpression can further enhance its expression. However, the expression of SOD-2 protein was weakly stained. In conclusion, our study indicates that miR-155-5p promotes oxalate- and COM-induced kidney oxidative stress injury by suppressing MGP expression.