Abstract

Drug resistance seriously limits the efficacy of chemotherapy drugs and hinders successful treatment in patients with gastric cancer. Endoplasmic reticulum (ER) and autophagy are recognized to be one of the mechanisms involving the drug resistance of gastric cancer. The mechanisms of action of JAK2/STAT3 pathway were investigated in AGS cells with drug resistance of 5-fluorouracil (5-FU) by corresponding inhibitors. We firstly analyzed the effects of JAK2/STAT3 inhibitor on the expression of drug resistance genes, autophagy markers, and ER stress-related markers on AGS/5-FU cells by Western blot. Whether JAK2/STAT3 pathway regulated the transcription of ATF6 was investigated through luciferase reporter assay. The expression of LC3B was detected by immunofluorescence assay. Next, ER stress inhibitor and ATF6 overexpression plasmid were respectively used to treat AGS/5-FU cells for analyzing whether JAK2/STAT3 pathway regulated ER stress. The results showed that JAK2 inhibitor or STAT3 inhibitor significantly altered the expression of these proteins and suppressed the activities of ATF6 promoter. Intriguingly, ATP6 overexpression could markedly reverse their effects. Moreover, similar effects to JAK2 inhibitor or STAT3 inhibitor appeared in ER stress inhibitor-treated group. These findings indicated that the involvement of JAK2/STAT3 pathway in regulating ER stress affected the 5-FU resistance of AGS cells and autophagy, which was mediated by ATF6. Targeting JAK2/STAT3 pathway could be a potential approach to decrease the 5-FU resistance of gastric cancer and enhance the sensitivity of gastric cancer to 5-FU. Additionally, our study offers new insights into the molecular mechanisms underlying the resistance of gastric cancer to 5-FU.

摘要

耐药性严重限制了化疗药物的疗效，阻碍了胃癌患者的成功治疗。内质网（ER）和自噬被认为是胃癌耐药的机制之一。研究了相应抑制剂对 5-氟尿嘧啶 (5-FU) 耐药的 AGS 细胞中 JAK2/STAT3 通路的作用机制。我们首先通过Western blot分析了JAK2/STAT3抑制剂对AGS/5-FU细胞耐药基因、自噬标志物和ER应激相关标志物表达的影响。通过荧光素酶报告基因分析研究 JAK2/STAT3 通路是否调节 ATF6 的转录。免疫荧光法检测LC3B的表达。下一个，ER应激抑制剂和ATF6过表达质粒分别用于处理AGS/5-FU细胞，分析JAK2/STAT3通路是否调节ER应激。结果表明，JAK2抑制剂或STAT3抑制剂显着改变了这些蛋白质的表达并抑制了ATF6启动子的活性。有趣的是，ATP6 过表达可以显着逆转它们的影响。此外，与 JAK2 抑制剂或 STAT3 抑制剂类似的作用出现在 ER 应激抑制剂治疗组中。这些发现表明 JAK2/STAT3 通路参与调节 ER 应激影响 AGS 细胞的 5-FU 抗性和自噬，这是由 ATF6 介导的。靶向 JAK2/STAT3 通路可能是降低胃癌 5-FU 耐药性和提高胃癌对 5-FU 敏感性的潜在途径。此外，