| Hepatocellular carcinoma(HCC)is one of the main causes of malignant tumor death in China.With the emergence of targeted drugs such as sorafenib,its prognosis has been improved,but the 5-year survival rate is still low.The discovery of new treatment modalities relies on a further understanding of the mechanisms involved in the development of liver cancer.Long noncoding RNAs(LncRNAs)are non-coding RNAs over 200 nt in length.Studies have shown that IncRNAs are involved in many physiological processes such as cell proliferation,apoptosis,migration,autophagy,and sternness maintenance.The abnormal expression and function of lncRNAs lead to the occurrence of various tumors by affecting biological behaviors such as cell proliferation,anti-apoptosis,tumor invasion and metastasis,and tumor resistance.So far,the expression,function and mechanism of IncRNAs in liver cancer are still not completely clear,and further research is needed.In view of the important role of LncRNA in cell activity,the relationship between the function of IncRNA related to liver cancer and its relationship with tumorigenesis,explore the possible effector molecules upstream and downstream,and clarify its possible signaling pathway for biotargeted therapy of liver cancer.Providing new therapeutic targets may improve the prognosis of liver cancer.This study is to study the expression,biological function and possible mechanism of lncRNA HOTAIRM1 in liver cancer.Part I:the relationship between LncRNA HOTAIRM1 expression in liver cancer and clinical prognosisObjectiveThe aim of this study was to investigate the expression of non-coding RNA(LncRNA)HOTAIRM1 in liver cancer tissues and cell lines.And the relationship between HOTAIRM1 expression and clinical prognosis.Materials and Methods:Collect liver cancer tissue and corresponding paracancerous tissue specimens.The expression of HOTAIRM1 in hepatocarcinoma tissues and adjacent tissues was statistically analyzed by Q-PCR of collected liver cancer and adjacent normal tissues.Kaplan-Meier was used to test the relationship between HOTAIRM1 and total survival(OS).The expression of HOTAIRM1 in normal liver cell line(L02)and liver cancer cell lines(HepG2,HBG11,HHCC)was analyzed by qPCR.Results:1.Q-PCR results suggest that LncRNA HOTAIRM1 is lowly expressed in liver cancer tissues and cell lines.2.The ROC curve indicates that LncRNA HOTATRM1 has high sensitivity and specificity for HCC diagnosis.3.LncRNA HOTAIRM1 low expression status often indicates poor prognosis.Summary:The HOTAIRM1 gene is poorly expressed in liver cancer tissues and liver cancer cell lines and suggests a poor prognosis.Part Ⅱ:The biological function of LncRNA HOTAIRM1 in liver cancerObjective:Explore the biological function of HOTAIRM1 in liver cancer.Materials and Methods:Overexpression of HOTAIRM1 in hepatocarcinoma cell lines HepG2 and HHCC by overexpression plasmid,and in vitro cell function assay:CCK-8 detection and flow cytometry to detect apoptosis to elucidate the biological behavior of HOTAIRM1 gene on hepatoma cells influences.Results:1.Overexpression of LncRNA HOTAIRM1 significantly reduced the proliferation of liver cancer cells;2.KEGG pathway enrichment analysis suggests that the function of HOTAIRM1 is mainly enriched in the apoptotic pathway;3.Apoptosis detection by flow cytometry suggests an increase in apoptosis after overexpression of HOTAIRM1.Summary:Overexpression of LncRNA HOTAIRM1 reduced proliferation of hepatoma cells and increased apoptosis.Part Ⅲ:Mechanism of Overexpression of LncRNA HOTAIRM1 Gene Inhibiting Proliferation of Hepatoma CellsObjective:The above two parts of our data confirmed that HOTAIRM1 is underexpressed in liver cancer,and overexpression of HOTAIRM1 results in decreased proliferation and increased apoptosis of hepatoma cells.In this section,we further explored the mechanism by which HOTAIRM1 gene is expressed to inhibit the proliferation of hepatoma cells.Materials and Methods:Western blot was used to verify the expression of apoptosis protein and wnt/β-catenin signaling pathway proteins before and after HOTAIRM1 overexpression.The FOP and TOP plasmids were constructed separately,and the double luciferase reporter gene was used to detect the activity of wnt/β-catenin signaling pathway.Results:1.Overexpression of HOTAIRM1 can reduce the expression of apoptosis-inhibiting genes and increase the expression of pro-apoptotic genes.2.The expression of wnt/β-catenin signaling pathway-related proteins was decreased after overexpression of HOTAIRM1.The dual luciferase reporter gene assay results in a decrease in the activity of the wnt/β-catenin signaling pathway.Summary:LncRNA HOTAIRM1 affects apoptosis and regulates cell proliferation by regulating the wnt/β-catenin signaling pathway.Conclusion:LncRNA HOTAIRM1 is underexpressed in HCC tissues and cell lines,and the ROC curve indicates that LncRNA HOTATRM1 has high sensitivity and specificity for HCC diagnosis.And such a low expression state often indicates a poor prognosis.Overexpression of LncRNA HOTAIRM1 significantly reduced the proliferation of hepatocarcinoma cells.The enrichment analysis of KEGG pathway suggested that the function of HOTAIRM1 was mainly enriched in the apoptotic pathway.Apoptosis detection by flow cytometry suggested that apoptosis was increased after overexpression of HOTAIRM1.Overexpression of HOTAIRM1 reduces the expression of apoptosis-inhibiting genes and increases the expression of pro-apoptotic genes.The expression of wnt/β-catenin signaling pathway-related protein was decreased after overexpression of HOTAIRM1,and the detection of dual luciferase reporter gene indicated that the activity of wnt/β-catenin signaling pathway was decreased.In conclusion,LncRNA HOTAIRM 1 may regulate hepatoma cell proliferation through the wnt/β-catenin signaling pathway. |
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