首页|LncRNA SBF2-AS1靶向miR-375对胃癌细胞免疫逃逸及mTOR1信号通路的影响

LncRNA SBF2-AS1靶向miR-375对胃癌细胞免疫逃逸及mTOR1信号通路的影响

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目的 探究LncRNA SBF2-AS1靶向miR-375对胃癌细胞免疫逃逸及mTOR1信号通路的影响.方法 Real-time PCR法检测正常胃上皮细胞及4种胃癌细胞系中LncRNA SBF2-AS1、miR-375表达水平,取胃癌细胞将其分为胃癌细胞(GC)组、胃癌细胞+LncRNA SBF2-AS1-NC(SN)组、胃癌细胞+LncRNA SBF2-AS1激动剂(SM)组、胃癌细胞+LncRNA SBF2-AS1抑制剂(SI)组、胃癌细胞+miR-375-NC(MN)组、胃癌细胞+miR-375抑制剂(MI)组、胃癌细胞+miR-375激动剂(MM)组、胃癌细胞+LncRNA SBF2-AS1抑制剂+miR-375激动剂(IM)组,免疫印迹法检测免疫逃逸相关因子及mTOR1信号通路蛋白表达,双荧光素酶报告实验证实LncRNA SBF2-AS1和miR-375的相互作用.结果 胃癌细胞系中的LncRNA SBF2-AS1 mRNA表达量显著高于正常胃上皮细胞系NGEC(P<0.05),miR-375 mRNA表达量显著低于正常胃上皮细胞系NGEC(P<0.05)其中HGC-27细胞的LncRNA SBF2-AS1、miR-375 mRNA值变化最大(P<0.05),因此选取其作为此次实验细胞;GC组、SN组、MN组,PD-1、PD-L1、mTOR1、p70S6K、4EBP1蛋白表达基本无差异(P>0.05),与与SN组、MN 组相比,SM 组、MI 组 PD-1、PD-L1、mTOR1、p70S6K、4EBP1 蛋白表达明显升高(P<0.05),与 SM 组、MI组相比,SI组、MM组PD-1、PD-L1、mTOR1、p70S6K、4EBP1蛋白表达明显降低(P<0.05),与SI组、MM组相比,IM组PD-1、PD-L1、mTOR1、p70S6K、4EBP1蛋白表达明显降低(P<0.05);双荧光素酶报告结果显示,转染LncRNA SBF2-AS1后可显著降低miR-375-3'-UTR-WT的荧光素酶活性(P<0.05),但对突变基因无显著影响(P>0.05).结论 抑制LncRNA SBF2-AS1可有效抑制胃癌细胞免疫逃逸,并抑制mTOR1信号通路表达,其作用机制可能与靶向激活miR-375有关.
Effects of LncRNA SBF2-AS1 targeting miR-375 on immune escape and mTOR1 signaling pathway of gas-tric cancer cells
Objective To explore the effects of LncRNA SBF2-AS1 targeting miR-375 on immune escape and mTOR1 signaling pathway of gastric cancer cells.Method The expression levels of LncRNA SBF2-AS1 and miR-375 in normal gastric epithelial cells and 4 gastric cancer cell lines were detected by real-time PCR.Gastric can-cer cells were collected and divided into gastric cancer cells(GC)group,gastric cancer cells+LncRNA SBF2-AS1-NC(SN)group,gastric cancer cells+LncRNA SBF2-AS1 agonist(SM)group,and gastric cancer cells+LncRNA SBF2-AS1 inhibitor(SI)group,gastric cancer cells+Mir-375-NC(MN)group,gastric cancer cells+miR-375 in-hibitor(MI)group,gastric cancer cells+miR-375 agonist(MM)group,gastric cancer cells+LncRNA SBF2-AS1 inhibitor+miR-375 agonist(IM)group.Immunoescape related factors and mTOR1 signaling pathway protein expres-sion were detected by Western blot,and the interaction between LncRNA SBF2-AS1 and miR-375 was confirmed by dual luciferase reporting assay.Results LncRNA SBF2-AS1 mRNA expression in gastric cancer cell line was signif-icantly higher than that in normal gastric epithelial cell line NGEC(P<0.05).The mRNA expression of miR-375 was significantly lower than that of normal gastric epithelial cell line NGEC(P<0.05),and the LncRNA SBF2-AS1 and miR-375 mRNA values of HGC-27 cells had the largest changes(P<0.05),so they were selected as the experimen-tal cells.There were basically no differences in the protein expressions of PD-1,PD-L1,mTOR1,p70S6K and 4EBP1 in GC group,SN group and MN group(P>0.05).The protein expressions of PD-1,PD-L1,mTOR1,p70S6K and 4EBP1 in SM group and MI group were significantly increased(P<0.05);compared with SM group and MI group,the protein expressions of PD-1,PD-L1,mTOR1,p70S6K and 4EBP1 in SI group and MM group were significantly decreased(P<0.05).Compared with SI group and MM group,the protein expressions of PD-1,PD-L1,mTOR1,p70S6K and 4EBP1 in IM group were significantly decreased(P<0.05).The results of dual luciferase re-port showed that transfection of LncRNA SBF2-AS1 could significantly reduce the luciferase activity of miR-375-3'-UTR-WT(P<0.05),but had no significant effect on mutated genes(P>0.05).Conclusion Inhibition of LncRNA SBF2-AS1 can effectively inhibit immune escape of gastric cancer cells and inhibit the expression of mTOR1 signal-ing pathway,and the mechanism of action may be related to the targeted activation of miR-375.

Gastric cancerImmune escapeLncRNA SBF2-AS1miR-375mTOR1 signaling pathway

蒋龙超、胡双丽、杨学燕、周欣、郭灏

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泸州市人民医院消化内科,四川泸州 646000

胃癌 免疫逃逸 长链非编码RNA SBF2-AS1 微小RNA-375 哺乳动物雷帕霉素靶蛋白信号通路

2024

解剖学研究
广东省解剖学会 中国解剖学会

解剖学研究

CSTPCD
影响因子:0.327
ISSN:1671-0770
年,卷(期):2024.46(1)
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