Store‐operated calcium entry mediated high glucose‐induced podocyte injury and mitochondrial impairment

Abstract

Purpose Diabetic Nephropathy is one of the major complications of diabetes mellitus. Hyperglycemia is a known initiator of diabetes mellitus. Accumulating evidence suggests that podocyte injury is tightly associated with the onset and progression of diabetic nephropathy. However, the mechanisms underlying podocyte injury induced by hyperglycemia or high glucose (HG) is poorly understood. Store-operated calcium entry (SOCE) is a multifunctional signaling pathway in many cell types. However, its role in podocyte injury in the settings of diabetes is not known. The aim of the present study was to examine if SOCE mediated HG-induced podocyte injury by impairing mitochondrial function. Methods All experiments were carried out in cultured immortalized human podocytes. Western blot was conducted to evaluate protein abundance of Orai1 (the channel protein mediating SOCE) and nephrin (a podocyte specific protein). Calcium imaging was used to analyze SOCE. TMRE fluorescence was used to probe the mitochondria membrane potential (MMP). Results HG (25mM) significantly increased podocyte Orai1 protein abundance for time periods ranging from 2 to 12 hours. This HG effect was dose dependent. Consistently, Ca2+ imaging experiments showed that HG treatment (25 mM for 12 hours) significantly enhanced podocyte SOCE. Furthermore, the abundance of nephrin protein decreased in podocytes after exposure to HG, indicating podocyte injury in the presence of ambient HG. However, the HG-induced decrease of nephrin was blunted by BTP2 (an SOCE inhibitor, 4 µM). Moreover, HG treatment (25 mM for 24 hours) decreased MMP, indicating damage of mitochondria by HG. The decrease of MMP was prevented by BTP2, suggesting the contribution of SOCE to the detrimental effect of HG treatment. Conclusion The present study suggests that upregulated SOCE contributes to HG-induced podocyte injury, possibly by impairing mitochondrial function.