MiR-29c Suppresses The Malignant Phenotype Of Hepatocellular Carcinoma Cells By Mediating TPX2 Associated With Immune Infiltration

Background: Liver cancer remains a global challenge and its incidence continues to grow,especially in Asia and Africa where it has the highest incidence.Among them,hepatocellular carcinoma(HCC)is the main type of primary malignancy of the liver,accounting for about 75% of the total.China has the highest number of cases due to its high incidence rate(18.3 cases per 100,000 people)and the largest population in the world.The main reason why the prognosis of hepatocellular carcinoma remains poor is that most patients with HCC are asymptomatic in the early stages and do not realize the onset of the disease,resulting in a diagnosis that is mostly intermediate to advanced,although liver transplantation and surgical resection are potential curative options for HCC,but the prime time for treatment has been missed,and HCC is significantly resistant to conventional chemotherapy and radiotherapy.Radiofrequency ablation(RFA),a transarterial chemoembolization(TACE)and broad-spectrum tyrosine kinase inhibitors(TKI),such as sorafenib and lenvatinib,as well as combination immunotherapy with antiangiogenic therapy,namely atelelizumab and bevacizumab,have been used for advanced hepatocellular carcinoma,however,with resistance issues and the challenge of disease recurrence,these approaches have struggled to significantly extend life expectancy.Research has intensified and new ways to treat HCC have emerged.In recent years,advanced therapies in the field of cancer targeting specific genes have shown promising results.A large number of immune genebased therapies have been shown to be significant in the treatment of cancer,which means that they also have great potential in the treatment of HCC.miR-29-3p is an important tumor suppressor that has been shown in several studies to inhibit a variety of cancers.TPX2 is a microtubule-associated protein that is highly expressed in a variety of malignancies and promotes cancer cell progression.However,its exact molecular mechanism and biological function in HCC are still not clearly elucidated.If the effects of miR-29-3p and TPX2 on HCC are clarified,it may provide a new direction and theoretical basis for the treatment of hepatocellular carcinoma.Objective: To clarify whether there is a significant difference in the expression of miR-29-3p and TPX2 in HCC and normal tissues;to explore whether miR-29-3p affects the progression of HCC cells by regulating TPX2;and whether TPX2 is associated with immune infiltration.Methods.1,The expression profile data of miR-29c-3p in the LIHC dataset were obtained and downloaded from the TCGA database,the differences in expression were analyzed using the Limma package in R software,and their differential expression was verified by real-time fluorescence quantitative PCR in HCC and normal tissues.2,Obtain and download the respective survival data of miR-29c-3p and TPX2 in the LIHC dataset from the TCGA database,and analyze the relationship between the respective expression and patient prognosis using the survivor package in R software.3,Cellular assays(CCK-8,scratch assay,Transwell,and apoptosis)were performed to investigate whether miR-29c-3p regulates proliferation,migration,invasion,and apoptosis of HCC cells.4,We based on Star Base database to predict,dual luciferase reporter to verify whether miR-29c-3p directly targets TPX2.then we used R software to analyze the expression level of TPX2 in various malignant tumors and explored the correlation between TPX2 expression level and miR-29c-3p expression.5,The GEPIA database,STRING database and Xiantao academic tools were used to synthesize whether the difference of TPX2 expression in LIHC was significant,and also to explore whether the expression status of TPX2 gene would affect the prognosis of HCC patients,and to analyze multiple genes associated with TPX2 expression,as well as the genes that interact with its protein.6.We then used immunohistochemistry to verify whether TPX2 protein differs between HCC and normal tissues at the tissue level.7,We observed whether miR-29c-3p-mediated TPX2 has a role in regulating the biological behavior of HCC cells based on cellular assays(CCK-8,scratch assay,Transwell,and apoptosis).8.We integrated several bioinformatic analysis tools(TIMER2.0,TISIDB data portal and Sento academic tools)to analyze the relationship between TPX2 and immune cells and immune regulators;we performed GO,KEGG enrichment analysis on TPX2 to explore the relationship between TPX2 and immune checkpoints.Results: We found that miR-29-3p was significantly downregulated in HCC,and the prognosis of miR-29-3p low expression group was poor.miR-29-3p overexpression was detrimental to the invasion and migration ability of HCC cells and promoted their apoptosis.We identified the downstream of miR-29c-3p by predictive analysis.TPX2 was significantly upregulated in HCC,and patients with high TPX2 expression had a poor prognosis.TPX2 knockdown somewhat counteracted the promoting effect on hepatocellular carcinoma cells after miR-29-3p was inhibited,and its effect on hepatocellular carcinoma cell biology was similar to miR-29c-3p overexpression.Conclusions: miR-29c-3p,a key gene regulating HCC,is lowly expressed in HCC,and its overexpression can significantly inhibit the biological function of tumor cells.TPX2 gene is highly expressed in HCC and predicts poor patient prognosis;TPX2 is closely associated with immune infiltration in hepatocellular carcinoma.miR-29c-3p can influence the expression of TPX2 by regulating the the progression of hepatocellular carcinoma.