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The Role Of RBM10 In Hepatocellular Carcinoma Was Elucidated By Using Clinical Tissue Samples And CKO Mice

Posted on:2024-01-02Degree:MasterType:Thesis
Country:ChinaCandidate:X X GuanFull Text:PDF
GTID:2544307166952699Subject:Pathology and pathophysiology
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RBM10,whose encoded gene locates on X chromosome p11.23,is an important member of the RNA binding protein family.It contains two RNA recognition motifs that can perform alterative RNA splicing and post-transcriptionally regulate the expression of its target genes.Disrupted expression and function of RBM10 can lead to a variety of diseases,including malignant tumors.RBM10 functions as a tumor suppressor in most tumors.However,recently published literature reveals conflicting results about the function of RBM10 in Hepatocellular carcinoma(HCC).In vitro cell experiments confirmed that RBM10 inhibited HCC cell proliferation and promoted apoptosis,while the results of bioinformatics analysis and detection in a large cohorts of clinical tissues supported that RBM10 may play a possible pro-cancer role in HCC.The reasons for this contradictory result are unclear,and may be due to the differences of the various stresses received by HCC between the in vivo tumor microenvironment and in vitro cell culture conditions,such as ischemia,hypoxia,immune stress,etc.This project aims to elucidate the effects of RBM10 on the development of HCC from the animal level and human tissue specimens.Objectives:To elucidate the correlation between RBM10 and the development and progression of hepatocellular carcinoma,and mine the underlying mechanism that RBM10 involved in the process of HCC development and progession.Methods:1.The differences between the expression of RBM10 in hepatocellular carcinoma and non-cancerous tissues,and the differences in overall survival(OS)and relapse-free survival(RFS)of patients with different expression of RBM10 were mined from the cancer genome atlas(TCGA)database.2.The expression of RBM10 in human HCC tissue samples was detected by immunohistochemistry,and the correlation between the expression and clinicopathologic features was analyzed.3.Hepato-specific RBM10conditional knockout mouse model was used to construct HCC model through intraperitoneal injection of DEN and intragastric administration of CCl4.The knockout of RBM10 was verified by PCR,WB and IHC,and the HCC development was observed by H&E staining and immunohistochemistry.4.The correlation between RBM10 and Ki67 expression in TCGA HCC cases were analyzed by KMplot online software.The expression of RBM10 and cell proliferation marker Ki67 in mouse HCC tissues and human tissue samples were detected by immunohistochemistry to explore the correlation between RBM10and Ki67,thus preliminarily exploring the potential mechanism of RBM10 in the carcinogenesis of HCC.Results:1.The TCGA database mining showed that the expression of RBM10 in HCC tissues was significantly higher than that in non-cancerous tissues.High expression of RBM10 is associated with poor clinical prognosis in HCC cases.2.Immunohistochemical detection of human hepatocellular carcinoma and noncancerous tissue samples showed that RBM10 expression was up-regulated in hepatocellular carcinoma(P<0.05).The expression of RBM10 was significantly correlated with tumor size(T classification)(P=0.001),histological grading/differentiation(G classification)(P=0.023)and TNM Stage(P=0.001),suggesting that RBM10 expression was up-regulated in HCC,and high expression of RBM10 was associated with carcinogenesis and tumor progression in HCC patients.3.PCR,Western blotting and immunohistochemical verification showed that the hepato-specific RBM10knockout mouse model was successfully constructed.HCC model was constructed by intraperitoneal injection of DEN and intragastric administration of CCl4.As compared with wild-type mice,we found that the number of HCC tumors in RBM10 knockout mice decreased,suggesting that RBM10 promoted the development of HCC.4.KMplot database analysis showed that RBM10 was positively correlated with Ki67 expression in TCGA HCC cases.The expressions of Ki67 and RBM10 was positively correlated between mouse HCC tissues and human HCC tissue samples were verifued by immunohistochemistry.The above data suggest that RBM10 may be involved in HCC by promoting cell proliferation.Conclusions:1.Bioinformatic and immunohistochemical data suggested that RBM10 has a potential oncogenic effect on the development and progession of hepatocellular carcinoma.2.Hepato-specific RBM10 conditional knockout inhibits carcinogenesis of HCC in mice,indicating that RBM10 plays an oncogene role in HCC development.3.The expression of RBM10 was positively correlated with the Ki67 proliferation index,indicating that RBM10may be involved in HCC by promoting cell proliferation.
Keywords/Search Tags:RBM10, Gene knockout, Hepatocellular carcinoma, Diethylnitrosamine, Ki67 proliferation index
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