Expression And Preliminary Functional Researches Of CHD1L Gene In Intrahepatic Cholangiocarcinoma

Background and Purposes:Intrahepatic Cholangiocarcinoma(ICC)is the second most common primary hepatobiliary cancer after hepatocellular carcinoma and accounts for about 10-15% of primary hepatocellular carcinoma.Although overall advances in the diagnosis and treatment of patients with ICC,the prognosis of this disease remains poor.One of the most important reasons influencing the dismal prognosis of ICC is on the basis of the late diagnosis at an advanced phase of disease,when cancer cell invasion into the blood and lymphatic vessels has resulted in metastatic spread and complete curative resection can no longer be achieved.Therefore,the discovery of specific markers for cancer's early diagnosis,early treatment and assessment of patient's prognosis is great important.CHD1L(chromodomain helicase/ATPase DNA binding protein 1-like gene)is a newly identified oncogene located at Chr1q21 that is frequently amplified in hepatocellular carcinoma(HCC).CHD1 L protein is comprised of two helicase domains,a C-terminal macro domain,nuclear localization and two putative phosphorylation sites.CHD1 L belongs to the SNF2-like family,which plays important roles in DNA repair,maintenance of chromosome integrity and transcriptional regulation.A number of studies have found that amplification of CHD1 L is very frequent in many solid tumors,including hepatocellular carcinoma,breast cancer,stomach cancer,and is closely related to tumor invasion and metastasis.However,the molecular mechanisms underlying and driving invasion of CHD1 L remain unclear in ICC.The purpose of this topic is found the correlation with clinical parameters and prognosis of the ICC patients through the detection of CHD1 L gene expression in the tissues of ICC and its cell lines.Moreover,we observe the effect of the proliferation,migration and invasion of intrahepatic cholangiocarcinoma RBE cell line by down-regulated CHD1 L expression.Methods:1.We investigated the expression of CHD1 L in 80 cases of ICC tissues and 30 cases of adjacent non-tumor tissues by immunohistochenmical and analysed the relationship between CHD1 L expression and the clinicopathological characteristics and prognosis of the ICC patients.2.We detected the protein and mRNA expression of CHD1 L in four intrahepatic cholangiocarcinoma cell lines(RBE?HCCC-9810?HUCCT1?QBC939)and normal intrahepatic bile duct cell HIBEpic by Western blot and real time qPCR.3.After we constructed and transfected interference plasmids CHD1L-RNAi with ICC cell line RBE,Western blot was used to detect the expression of CHD1 L,and CCK-8 test,cell scratch test,transwell migration and invasion experiments were used to discovered abilities of RBE cell line's proliferation,migration and invasion.4.Statistical analysis was performed by using SPSS19.0 statistical software,measurement data was represented by using(x±S),the experimental data were compared by t-test analysis,Chi-square test,survival analysis and Cox regression analysis,P<0.05 was that the difference was statistically significant.Results:1.Immune histochemical methods examined 80 cases of ICC tissues and 30 cases of adjacent non-tumor tissues of paraffin embedding CHD1 L protein expression in the organization,and CHD1 L is mainly lied on nuclear,relative to adjacent normal tissue,the expression of cancerous samples is higher.We also find the expression of CHD1 L is closely related to pathological grading(P =0.011),vascular invasion(P=0.002),lymph node metastasis(P=0.008)and TNM stage(P = 0.001);Survival analysis revealed that the higher expression of CHD1 L may independently be an adverse prognostic factors for ICC patients.2.Compared with HIBEpic,the protein and mRNA expression of CHD1 L in ICC cell line have different degrees of ascent,especially in RBE cell.3.When we constructed interference plasmids CHD1L-RNAi with RBE cell,CHD1L's protein expression,proliferation,migration and invasion ability of RBE was both reduced.Conclusion:CHD1L plays an important role in the development and progression of ICC,and can be used as an important target for prognosis of ICC patients.