Aims. In this report, it was investigated that hepatoma cells can cause downregulation of cytotoxic T lymphocyte (CTL) function and tea polyphenols (TPs) can reverse downregulation of CTL function. Methods. The expression of GRP78, PD-1, and TIM-3 was detected by western blotting in CTLL-2 cocultured with Hepa1-6 cells. Moreover, perforin (PRF1) and granzyme B (GzmB) protein levels and ER morphology were examined by ELISA and TEM, respectively. After 4-phenylbutyric acid (4-PBA) or tunicamycin (TM) treatment, programmed cell death protein 1 (PD-1), and mucin domain 3 (TIM-3), PRF1, and GzmB were measured by western blotting and ELISA. After sh-CHOP or GSK2656157 (PERK inhibitor) stimulation, the activation of the PERK-CHOP pathway was detected in CTLL-2 cells. Finally, changes in PD-1, TIM-3, PRF1, and GzmB levels were detected to verify the reversal of CTL depletion by TP. Results. The expression of GRP78, PD-1, and TIM-3 clearly increased, and swelling was observed for the endoplasmic reticulum (ER) in CTLL-2 cells cocultured with hepatoma cells. Concurrently, the levels of PRF1 and GzmB decreased. CTLL-2 depletion was induced after stimulation with TM and differed from 4-PBA stimulation. Treatment with sh-CHOP or GSK2656157 caused a decrease in PD-1 and TIM-3 expression, whereas the expression of PRF1 and GzmB clearly increased. After adding TP, the function of CTLs increased markedly. Conclusion. Hepatoma cells induced the depletion of CTLs through the ER stress PERK-CHOP pathway, and TP reversed this depletion by downregulating ER stress.

中文翻译：

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通过 ER 应激依赖性 PERK-CHOP 信号通路消除和逆转诱导 CTL 的肝细胞癌

目标。在本报告中，研究了肝癌细胞可引起细胞毒性T淋巴细胞（CTL）功能的下调，茶多酚（TPs）可逆转CTL功能的下调。方法. 通过蛋白质印迹法检测与Hepa1-6细胞共培养的CTLL-2中GRP78、PD-1和TIM-3的表达。此外，穿孔素 (PRF1) 和颗粒酶 B (GzmB) 蛋白水平和 ER 形态分别通过 ELISA 和 TEM 检测。在 4-苯基丁酸 (4-PBA) 或衣霉素 (TM) 处理后，通过蛋白质印迹和 ELISA 测量程序性细胞死亡蛋白 1 (PD-1) 和粘蛋白结构域 3 (TIM-3)、PRF1 和 GzmB。在 sh-CHOP 或 GS​​K2656157（PERK 抑制剂）刺激后，在 CTLL-2 细胞中检测到 PERK-CHOP 通路的激活。最后，检测到 PD-1、TIM-3、PRF1 和 GzmB 水平的变化，以验证 TP 对 CTL 消耗的逆转。结果. 与肝癌细胞共培养的 CTLL-2 细胞中 GRP78、PD-1 和 TIM-3 的表达明显增加，并且观察到内质网 (ER) 肿胀。同时，PRF1 和 GzmB 的水平下降。CTLL-2 耗竭是在用 TM 刺激后诱导的，与 4-PBA 刺激不同。用 sh-CHOP 或 GS​​K2656157 治疗导致 PD-1 和 TIM-3 表达降低，而 PRF1 和 GzmB 的表达明显增加。添加TP后，CTLs的功能明显增强。结论。肝癌细胞通过 ER 应激 PERK-CHOP 通路诱导 CTL 耗竭，而 TP 通过下调 ER 应激来逆转这种耗竭。