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目的:探讨p38MAPK信号转导通路参与蛋白酶体抑制剂诱导甲状腺癌细胞凋亡的作用机制。方法:选取甲状腺癌细胞系,利用丝裂原活化蛋白激酶(MAPK)的三种抑制剂抑制磷酸化途径,利用微小RNA干扰技术转染细胞,封闭p38MAPK。用蛋白质印迹法检测红系衍生的核因子2相关因子2(Nrf2)及p38MAPK的表达,实时定量PCR检测细胞内GCLC mRNA的表达,测定还原型谷胱甘肽的含量,流式细胞仪法检测甲状腺癌细胞凋亡率。结果:在甲状腺癌细胞系8305C细胞中,与对照组相比,p38MAPK的抑制剂SB203580和sip38MAPK均能够抑制蛋白酶体抑制剂诱导的Nrf2细胞核易位(P<0.01),并使随后的GCLC mRNA表达减少、GSH含量减少(P<0.01),显著提高蛋白酶体抑制剂诱导的甲状腺癌细胞凋亡率(P<0.01),而其他2种抑制剂和随机序列核酸siRNA无上述作用。结论:蛋白酶体抑制剂通过p38MAPK途径激活Nrf2的细胞核易位,进而导致GCLC的诱导作用和谷胱甘肽生成的连锁反应,成为蛋白酶体抑制剂在甲状腺癌细胞中的一种抗凋亡机制。
AIM: To investigate the mechanism of p38 MAPK signal transduction pathway involved in proteasome inhibitor-induced apoptosis of thyroid cancer cells. Methods: Thyroid cancer cell lines were selected. Three inhibitors of mitogen-activated protein kinase (MAPK) were used to inhibit the phosphorylation pathway. The p38MAPK was transfected into the cells by microRNA interference technique. The expression of erythroid-derived nuclear factor 2 (NF-2) and p38MAPK were detected by Western blotting. The expression of GCLC mRNA was detected by real-time PCR. The content of reduced glutathione Thyroid cancer cell apoptosis rate. Results: Compared with the control group, p38MAPK inhibitor SB203580 and sip38MAPK all inhibited the proteasome inhibitor-induced Nrf2 nuclear translocation (P <0.01) and the subsequent GCLC mRNA expression in the thyroid cancer cell line 8305C (P <0.01). The apoptosis rate of thyroid cancer cells induced by proteasome inhibitor was significantly increased (P <0.01), while the other two inhibitors and random sequence siRNA did not. CONCLUSION: Proteasome inhibitor activates the nuclear translocation of Nrf2 through the p38MAPK pathway, leading to the induction of GCLC and the chain reaction of glutathione, which is an anti-apoptotic mechanism of proteasome inhibitor in thyroid cancer cells.