基于GRP78-ATF6通路探讨沉默SAT1对顺铂诱导耳蜗毛细胞损伤的影响OA

Objective To investigate the role of spermidine/spermine N1-acetyltransferase 1(SAT1)in cisplatin-induced endoplasmic reticulum stress in mouse-derived immortalized auditory hair cell-like HEI-OC1 cells,and to define the impact of SAT1 on cisplatin-induced damage to cochlear hair cells.Methods HEI-OC1 cells were divided into four groups:siRNA negative control group(siNC),SAT1 gene silencing group(siSAT1),negative control with cisplatin treatment group(siNC+DDP),and SAT1 gene silencing with cisplatin treatment group(siSAT1+DDP).The cells were transfected with SAT 1-targeting siRNA and subsequently treated with cisplatin.Cell proliferation from 0 to 72 h was assessed using the CCK-8 assay;apoptosis and ultrastructural changes were analyzed by flow cytometry and transmission electron microscopy;endoplasmic reticulum stress markers(IRE1α,XBP1s,GRP78,ATF6)and apoptotic protein Caspase-12 were detected by Western blotting.Results All four SAT1-siRNA treatments effectively silenced SAT1 gene expression 24 hours post-transfection,with the most pronounced reduction observed in the siSAT1-740 group(P＜0.05).Consequently,the siSAT1-740 group was selected for subsequent experiments.Western blotting results showed that the protein expression level of SAT1 in the siNC group was significantly higher than that in the siSAT1 group(P＜0.05).Flow cytometry and transmission electron microscopy revealed that SAT1 knockdown not only significantly reduced DDP-induced early,late,and total apoptosis rates(P＜0.05),but also markedly improved apoptotic features such as chromatin condensation and nuclear fragmentation in morphological terms.Western blotting results indicate that the expression levels of SAT1,IRE1α,XBP1s,GRP78,Caspase-12,and ATF6 proteins exhibited statistically significant differences(P＜0.05)across different treatment groups.Under DDP-induced endoplasmic reticulum stress conditions,silencing SAT1 upregulates GRP78 protein expression(P＜0.05)while downregulating IRE1α,ATF6,Caspase-12 and XBP1s expression(P＜0.05).Conclusion The study revealed that silencing the SAT1 gene may counteract cisplatin-induced cellular damage by regulating the GRP78/ATF6 pathway,thus identifying SAT1 as a potential novel therapeutic target for preventing and treating drug-induced ototoxicity.CCK-8 assays demonstrated that SAT1 knockdown significantly alleviated the proliferation-inhibitory effect of DDP on HEI-OC1 cells(P＜0.05).