Store-Operated Calcium Entry regulated IL6 expression and matrix proteins in glomerular mesangial cells

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2021

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

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Abstract

Background: Glomerular mesangial cells (MCs) produce various cytokines in response to multiple stimuli. MCs also contribute to matrix expansion in kidney diseases. The aim of this study was to determine whether interleukin-6 (IL6) participated in matrix protein turnover and if Orai1-mediated store-operated calcium entry (SOCE) in MCs regulate IL6. Methods: In cultured human MCs, expression of target proteins was examined in culture media and whole cell lysates using ELISA and Western blot analysis, respectively. Overexpression of IL6 was achieved using the expression plasmid for IL6 while knockdown of Orai1 using Orai1 siRNA. Orai1 in MCs was knocked down using the targeted nanoparticle-siRNA delivery system in wild type C57BLKS/J mice at the age of 16 weeks. Glomerular IL6 expression was evaluated using immunohistochemistry of kidney sections. Results: In human MCs, overexpression of IL6 and its receptor decreased the abundance of fibronectin and collagen type IV in MCs. Thapsigargin (activator of SOCE, 1µM) significantly increased IL6 level in supernatant media and in whole cell lysates of MCs at 15 hrs, and this effect was attenuated by GSK 7975A (selective inhibitor of SOCE). Inhibition of NFкB pathway significantly blunted the thapsigargin-induced increase in IL6 protein abundance. Moreover, thapsigargin stimulated the nuclear translocation of p65 in human MCs. IL6 expression was reduced in the glomeruli of the mice treated with nanoparticle/Orai1 siRNA for 2 weeks compared to control mice. Conclusion: SOCE positively regulates IL6 production by MCs through activation of NFкB pathway and IL6 in turn inhibits matrix proteins.

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