ORAI store‐operated calcium channels are associated with proximal renal tubule dysfunction in diabetic nephropathy (689.13)

Abstract

Proteinuria is not only an early sign of diabetic nephropathy (DN) but also contributes to the development and progression of the disease. The molecular mechanisms underlying proteinuria remain unclear. Here we explored the role of ORAI store‐operated calcium channels in DN. RT‐PCR and western blot experiments demonstrated expression of all ORAI isoforms (ORAI1‐3) in human kidney. ORAI channels were preferentially expressed in the proximal tubules and intrarenal arteries as assessed by immunohistochemistry; and ORAI channel expression was down regulated in type 1 diabetic patients with DN. Using primary human proximal tubular cells, we showed that high glucose (25 mM) significantly reduced ORAI1‐3 mRNA and protein expression, whereas insulin increased channel expression. Conversely, blockade of insulin signaling using tyrphostin A23 or wortmannin reduced channel expression. We identified a current evoked by thapsigargin and inositol 1,4,5‐trisphosphate (IP3 ) in the proximal tubular cells, sensitive to diethylstilbestrol (DES) and 2‐APB. Using ORAI1‐3/STIM1 overexpressing systems, we found that DES and BTP2 inhibited all three ORAI channels, while 2‐APB selectively blocked ORAI1 and ORAI2, but stimulated ORAI3 channels. Pharmacological inhibition of ORAI channel activity by DES, 2‐APB or BTP2, or silencing of ORAI expression by small interfering RNA significantly reduced FITC‐albumin uptake in proximal tubular cells. These results suggest that down‐regulation of ORAI channel expression or inhibition of channel activity leads to impaired protein reabsorption in the proximal tubules, representing a novel mechanism underlying diabetic proteinuria.Grant Funding Source: British Heart Foundation, Leverhulme Trust, China Scholarship Council; Swedish Research Council