Store-Operated Calcium Entry regulated IL-6 expression and matrix protein changes in glomerular mesangial cells.

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2020

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Ma, Rong
Chaudhari, Sarika
Tao, Yu
Yazdizadeh Shotorbani, Parisa

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Abstract

IL-6 can function as a pro or anti-inflammatory cytokine, depending on the cell type or pathology. Cytokines contribute to the early changes and histological impairment in chronic kidney diseases like diabetic nephropathy (DN). Glomerular mesangial cell (MC) is a major cell type in glomerulus to produce cytokines in response to diabetes and a major contributor to mesangial matrix expansion in DN. This study aimed to determine whether and how Store-Operated Calcium Entry (SOCE) regulated IL-6 production and downstream effects in MCs. Experiments were performed in cultured human MCs. The knockdown of proteins was achieved using a targeted siRNA delivery system, while overexpression of protein was obtained with specific human IL-6 plasmid in MCs. The expression of target proteins was examined either in culture media and cell lysates using ELISA and Western blot analysis, respectively. Thapsigargin (an activator of store-operated Ca2+ entry, SOCE) significantly increased IL-6 level, and this effect was attenuated by GSK 7975-A (a selective inhibitor of SOCE). Silencing Orai1 (the channel protein mediating SOCE) significantly decreased IL-6 protein expression. Inhibition of NFκB pharmacologically and genetically significantly reduced SOCE-induced IL-6 production. Thapsigargin also stimulated the nuclear translocation of the p65 subunit of NFκB. Overexpressing IL-6 and its receptor decreased the expression of fibronectin and collagen IV in MCs. Our results suggest that IL-6 production is positively regulated by Orai1-mediated SOCE via the NFκB pathway in MCs and inhibits ECM protein production by MCs.

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