Combination of Chymostatin and Aliskiren attenuates ER stress induced by lipid overload in kidney tubular cells

Background: Lipotoxicity plays an important role in the pathogenesis of kidney injury. Our previous study demonstrated that activation of local renin-angiotensin system (Ras) was involved in saturated free fatty acids palmitic acid (PA)-induced tubular cell injuries. The current study aims to investigate whether suppression of Ras by combination of direct Renin Inhibitor aliskiren and noncanonical Ras pathway chymase inhibitor chymostatin attenuates PA or Cholesterol induced-endoplasmic reticulum stress (ER stress) and apopotosis in cultured human proximal tubular HK2 cells.

Methods: HK2 cells were treated with saturated fatty acid PA (0.6 mM) for 24 h or Cholesterol (10 μg/ml) for 6d with or without chymostatin and/or aliskiren. Expressions of the ER stress associated proteins and Apoptosis markers were detected by western blotting. The mRNA levels of Ras components were measured by real-time qPCR.

Results: Combination treatment of chymostatin and aliskiren markedly suppressed PA or cholesterol-induced ER stress, as reflected by increased BiP, IRE1α, phosphorylated-eIF2α and ATF4 as well as proapoptotic transcription factor CHOP. The ratio of Bax/Bcl-2 and cleaved Caspase-3, two markers of Apoptosis were upregulated by PA or Cholesterol treatment. PA treatment was also associated with increased levels of Angiotensinogen and angiotensin type 1 receptor (AT1R) mRNA expression. Combination treatment of chymostatin and aliskiren markedly suppressed PA or cholesterol-induced ER stress and Apoptosis. The protective effect of two inhibitors was also observed in primary cultured cortical tubular cells treated with PA. In contrast, chymostatin and/or aliskiren failed to prevent ER stress induced by tunicamycin.

Conclusions: These results suggested that combination treatment of chymostatin and aliskiren attenuates lipid-induced renal tubular cell injury, likely through suppressing activation of intracellular Ras.