| Objective Gastric cancer is one of the major diseases affecting people’s health,and China is one of the areas with high incidence of gastric cancer.Great progress has been made in the comprehensive treatment of gastric cancer,but chemotherapy multidrug resistance still seriously reduces the effect of chemotherapy,so it is necessary to study the causes of chemotherapy multidrug resistance in gastric cancer and its mechanism,and to find targeted methods to reverse drug resistance or delay the its occurrence.MicroRNAs are a class of non-coding RNAs that play important roles in tumor development,progression,invasion and metastasis.MicroRNA-155 is abnormally expressed in a variety of tumors including gastric cancer,and its main function is to post-transcriptionally regulate its target genes and play different roles in pathological processes such as immunity,inflammation,tumor occurrence and progression and multidrug resistance.The purpose of this study was to explore the role of microRNA-155 in cisplatin resistance of gastric cancer cells and its mechanism by establishing cisplatin resistant gastric cancer cell lines and knockout and overexpression of microRNA-155.Methods(1)Gastric cancer tissues and corresponding adjacent tissues were collected,and the expression of microRNA-155 was detected by real-time PCR(QRT-PCR).Rictor and Fos were screened out as target genes of micror NA-155 by bioinformatics method.The expression of microRNA-155,target genes,autophagy and apoptosis-related proteins were detected by western blot and immunohistochemical experiments to explore the effects of microRNA-155 on Rictor and Fos as well as autophagy andapoptosis.(2)Cisplatin-resistant gastric cancer cell lines were established by high dose administration of Cisplatin.Gene knockout and overexpression of microRNA-155,Rictor and Fos were performed by lentiviral transfection method,and the corresponding cell lines were established.CCK8 assay,Transwell invasion assay,high-connotation cell imaging analysis assay,clone formation assay were used to observe the changes of the biological behavior of each cell lines.The expression levels of Rictor,Fos,Beclin1,LC3 II,Caspase3 and Caspase9 in the newly established cell lines were detected by Western blot,the autophagy level was detected by transmission electron microscopy and high-content cell imaging analysis,and the apoptosis level was detected by flow cytometry.(3)NOD-SCID mouse xenograft model was established by subcutaneous injection to observe the tumorigenesis rate,tumor growth rate,and chemotherapy effect.The expression of Rictor,Fos,Beclin1,LC3 II,Caspase3 and Caspase9 in the tumor of the mice were detected by immunohistochemical method.Results(1)The results of biogenic analysis showed that micror NA-155 had many target genes,and Rictor and Fos were selected as research objects.In early gastric cancer microRNA-155 was lowly expressed,Rictor and Fos were highly expressed,Beclin1 and LC3II were lowly expressed,Caspase3 and Caspase9 were highly expressed.In advanced gastric cancer microRNA-155 was highly expressed,Rictor and Fos were lowly expressed,Beclin1 and LC3 II were highly expressed,Caspase3 and Caspase9 were lowly expressed.(2)Cisplatin-resistant gastric cancer cell lines were established by high dose administration of cisplatin,and the drug-resistant gastric cancer cells underwent significant morphological changes.In drug-resistant gastric cancer cells,microRNA-155 was highly expressed,Rictor and Fos were lowly expressed,Beclin1 and LC3II were highly expressed,Caspase3 and Caspase9 were lowly expressed.Autophagosomes was increased in cells,and cell apoptosis was reduced.The proliferation of drug-resistant gastric cancer cells was slowed down and cell cycle was blocked in G1 phase.The invasion ability of drug-resistant gastric cancer cells was enhanced,and the cytoskeleton was changed.Autophagy was decreased when Rictor was overexpressed,and autophagy was increased under drug intervention after Rictor was knocked out.When Fos was overexpressed,apoptosis was increased underdrug intervention,and apoptosis was reduced when Fos was knocked out.(3)The tumorigenesis rate of drug-resistant gastric cancer cells was low when injected subcutaneously in NOD-SCID mice,and tumor growth was slow and cisplatin sensitivity was poor.The variation trend of gene expression levels in tumor tissues was consistent with the results of cell experiments.Conclusion MicroRNA-155 is highly expressed in advanced gastric cancer and drug-resistant gastric cancer cells.MicroRNA-155 inhibits the expression of Rictor and Fos,which are its target genes.Among them,Rictor is involved in the regulation of intracellular autophagy and Fos is involved in the regulation of apoptosis.MicroRNA-155 may induce autophagy and inhibit apoptosis by inhibiting the expression of Rictor and Fos,and participate in the occurrence of multi-drug resistance in gastric cancer cells. |
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