过表达M2 型肿瘤相关巨噬细胞TIA1 基因通过调控PI3K/AKT 信号通路 抑制胃癌细胞侵袭和迁移

Overexpression of TIA1 gene in M2-type tumor-associated macrophages inhibited invasion and migration of gastric cancer cells by regulating the PI3K/AKT signaling pathway.

Objective: To investigate the effect of overexpression of T cell restricted intracellular antigen 1（ TIA1） gene in M2- type tumor-associated macrophages（ M2-TAMs） on invasion and migration of gastric cancer cells and its mechanism. Methods: Primary TAMs were extracted from gastric cancer tissues and induced to differentiate into M2-TAMs using IL-4 and IL-13. The TIA1 overexpression plasmid （oe-TIA1） and its empty vector were transfected into M2-TAMs. The expression levels of TIA1 mRNA and protein were detected by qRT-PCR and Western blot. The expression levels of CD206 and CD163 were detected by flow cytometry. The levels of IL-10, TGF-β, VEGF-A and Arg-1 in cell culture supernatant were detected by ELISA. The transfected M2-TAMs were co-cultured with gastric cancer BGC-823 cells by Transwell co-cultivation system, and PI3K agonist 740Y-P was used to intervene in parallel. The cell migration ability was detected by scratch assay. Transwell assay was used to detect cell invasion ability. Protein expression levels of PI3K, p-PI3K, AKT, p-AKT, MMP-2 and MMP-9 in the cells were detected by Western blot. Results: ①Compared with primary TAMs, the levels of CD206 and CD163 expression in M2-TAMs cells and IL-10, TGF-β, VEGF-A and Arg-1 in cell culture supernatant were significantly increased （P<0.05）, while the expression levels of TIA1 mRNA and protein were significantly decreased （P< 0.05）. Overexpression of TIA1 gene significantly decreased the expression levels of CD206 and CD163 in M2-TAMs and IL-10, TGF-β, VEGF-A and Arg-1 in cell culture supernatant （P<0.05）. ②Overexpression of TIA1 gene in M2-TAMs could significantly reduce the migration and invasion ability and the expression levels of p-PI3K/PI3K, p-AKT/AKT, MMP-2 and MMP-9 proteins in BGC-823 cells （P<0.05）. ③740Y-P could significantly reverse the inhibitory effects of overexpression of TIA1 gene in M2-TAMs on migration, invasion and PI3K/AKT signaling pathway of BGC-823 cells. Conclusion: Overexpression of TIA1 gene in M2-TAMs can affect the invasion and migration of gastric cancer cells by blocking the PI3K/AKT signaling pathway. [ABSTRACT FROM AUTHOR]

目的: 探讨过表达M2型肿瘤相关巨噬细胞（M2-TAMs）T淋巴细胞内抗原1（TIA1）基因对胃癌BGC-823细胞 侵袭和迁移的影响及其机制。方法: 从胃癌组织中提取原代TAMs, 采用IL-4和IL-13刺激TAMs诱导分化成M2-TAMs, 再将 TIA1 基因过表达质粒（oe-TIA1）及其空载质粒（Vector）转染至M2-TAMs 中, 采用qRT-PCR 和Western blot 检测细胞中TIA1 mRNA 和蛋白表达水平;流式细胞术检测细胞中CD206 和CD163 表达水平;ELISA 检测细胞培养上清液中IL-10、TGF-β、 VEGF-A 和Arg-1 水平。通过Transwell 共培养体系将转染后的M2-TAMs 与胃癌BGC-823 细胞共培养, 并联用PI3K 激动剂 740Y-P进行干预, 采用划痕实验检测细胞迁移能力;Transwell实验检测细胞侵袭能力;Western blot检测细胞中PI3K、p-PI3K、 AKT、p-AKT、MMP-2和MMP-9等蛋白表达水平。结果: ①与原代TAMs比较, M2-TAMs中CD206和CD163表达水平及细胞培 养上清液中IL-10、TGF-β、VEGF-A 和Arg-1 水平均显著升高（P<0.05）, 而TIA1 mRNA 和蛋白表达水平显著降低（P<0.05）。 TIA1基因过表达可显著降低M2-TAMs中CD206和CD163表达水平及细胞培养上清液中IL-10、TGF-β、VEGF-A和Arg-1水平 （P<0.05）。②M2-TAMs中过表达TIA1基因可显著降低BGC-823细胞迁移和侵袭能力及p-PI3K/PI3K、p-AKT/AKT、MMP-2和 MMP-9等蛋白表达水平（P<0.05）。③740Y-P可显著逆转M2-TAMs中过表达TIA1基因对BGC-823细胞迁移、侵袭及PI3K/AKT 信号通路的抑制作用。结论: 过表达M2-TAMs中TIA1基因表达可通过阻断PI3K/AKT信号通路影响胃癌细胞侵袭和迁移。 [ABSTRACT FROM AUTHOR]