| Objective:（1）Meta-analysis was used to evaluate the effectiveness of common markers in the process of EMT（epithelial to mesenchymal transition）in Hep G2 cells;（2）To study the malignant biological behavior changes of insulin-resistant liver cancer cells,explore the differences in micro RNA（mi RNA）expression,explore the target genes and functions of the core mi RNA with significant changes,and clarify the mechanism leading to the enhancement of malignant biological behavior of insulinresistant liver cancer cells,so as to provide a theoretical basis for the screening of new targets for clinical diagnosis and treatment of liver cancer.Methods:（1）The main English and Chinese databases were searched by computer until December 14,2020.In addition,the references attached to the original literature and review literature are traced to supplement the relevant literature;（2）The changes of biological behavior of insulin-resistant hepatocellular carcinoma cells in vitro and in vivo were studied by molecular biology methods.The mi RNA expression profiling chip was used to screen the mi RNA expression difference between insulinresistant hepatocellular carcinoma cells and their parental cells,and bioinformatics analysis was performed to analyze the related pathways and the interaction relationship.The core differentially expressed mi RNA molecules were determined and verified by PCR（Polymerase Chain Reaction）.Bioinformatics method was used to predict the target gene of mi R-5195-3p,and the target gene was verified by dual luciferase reporter assay.The effect of regulating the expression level of mi R-5195-3p on the malignant biological behavior of liver cancer cells was detected in vitro and in vivo.Results:（1）This study included 61 independent comparisons from 58 articles.Our study showed that E-cadherin responds well to the intervention of medication,gene,hypoxia,and other tumor microenvironments,as well as non-coding RNA（nc RNA）overexpression and silencing.N-cadherin can effectively evaluate the intervention effect of medication,genetic intervention,hypoxia and other tumor microenvironments,as well as nc RNA overexpression.Vimentin reflects the effects of medication,pro-EMT genetic intervention and gene knockout/knockdown,anti-EMT nc RNA overexpression and anti-EMT nc RNA silencing and hypoxia.Snail only responds to the intervention of anti-EMT genetic intervention and gene knockout/knockdown,tumor microenvironments other than hypoxia,anti-EMT nc RNA overexpression and nc RNA silencing;（2）The results of in vitro and in vivo studies showed that Hep G2/IR cells had strong malignant biological behavior.A total of 2006 mi RNAs differentially expressed in Hep G2/IR cells compared with their parental cells were screened by mi RNA expression profile chip technology,including 32 differentially expressed mi RNAs.Bioinformatics analysis showed that some pathways of target gene enrichment were closely related to tumorigenesis and development.After further analysis of the interaction relationship,the core mi RNA molecules were determined and verified.Five mi RNAs represented by mi R-5195-3p were found to be consistent with microarray results.Bioinformatics prediction and dual luciferase reporter assay showed that SOX9（sex determining region Y-box 9）and TPM4（Tropomyosin 4）were the target genes of mi R-5195-3p in Hep G2 cells.Western blot experiment showed that the up-regulation of mi R-5195-3p expression in Hep G2/IR cells could inhibit the expression of SOX9 and TPM4,while the inhibition of mi R-5195-3p expression in Hep G2 cells was the opposite.Functional experiments in vitro and tumorigenicity experiments in nude mice showed that up-regulation of mi R-5195-3p inhibited the proliferation,migration,invasion and drug resistance of Hep G2/IR cells,while inhibition of mi R-5195-3p expression in Hep G2 cells showed the opposite results.Conclusion:（1）The results of meta-analysis showed that the markers for measuring EMT level of Hep G2 cells under different intervention measures in vitro were different,and E-cadherin,N-cadherin and Vimentin could be selected as their marker proteins in drug intervention;E-cadherin and Vimentin can be selected as the marker proteins in gene intervention;E-cadherin and N-cadherin can be selected as the marker proteins in tumor microenvironment intervention;E-cadherin and Snail can be selected as marker proteins when non-coding RNA intervention;（2）mi R-5195-3p may regulate the malignant biological behavior of insulin resistant hepatocellular carcinoma cells by targeting SOX9 and TPM4,which provides new research ideas and targets for reversing drug resistance and improving prognosis of insulin resistant hepatocellular carcinoma. |
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