Study On The Role Of LINC-PINT In Cholangiocarcinoma And Preliminary Exploration Of Its Molecular Mechanism

Objective: cholangiocarcinoma(CCA)is the most common primary biliary malignancy and the second most common primary hepatic malignancy in humans.At present,radical resection is the best cure for cholangiocarcinoma.However,due to the lack of obvious early symptoms and early diagnosis methods,most patients have developed to advanced stage and lost the opportunity for surgery.Recent studies have found that the expression levels of some Lnc RNAs in tumor tissues are significantly higher than those in normal tissues.These Lnc RNAs with specificity may become important indicators for diagnosis and prognosis prediction of tumors.It has been reported that LINC-PINT is involved in the occurrence and development of some tumors,but the association and effect between LINC-PINT and cholangiocarcinoma have not been reported yet.The purpose of this study was to investigate the effects of LINC-PINT on the proliferation,migration,invasion and mitotic cycle of bile duct cancer cells,and to conduct a preliminary exploration of the molecular mechanism of LINC-PINT.Methods: 1.The expression of LINC-PINT in cholangiocarcinoma cell lines RBE and QBC939 was detected by q RT-PCR;2.Using RNA interference technology to knock down LINC-PINT in cholangiocarcinoma cell lines RBE and QBC939 to determine the transfection effect by observing green fluorescence under fluorescence microscope,and to detect the knockdown effect by q RT-PCR;3.CCK-8 assay was used to investigate the effect of LINC-PINT on the proliferation of cholangiocarcinoma cells;4.Wound healing assay was used to investigate the effect of LINC-PINT on the migration of cholangiocarcinoma cells;5.Transwell assay was used to investigate the effect of LINC-PINT on the invasion of cholangiocarcinoma cells;6.The effect of LINC-PINT on the mitotic cycle of cholangiocarcinoma cells was investigated by FACS flow cytometry;7.Bioinformatics analysis technology was used to predict the co-expressed genes of LINC-PINT,and GO enrichment analysis and KEGG pathway enrichment analysis were performed;8.Lnc Locator was used to predict the subcellular localization of LINC-PINT.Results: LINC-PINT was highly expressed in QBC939 and RBE cell lines,which was consistent with our previous results in cholangiocarcinoma tissues.The knockdown of LINC-PINT caused a marked decrease in the proliferation,migration,and invasion capabilities of QBC939 and RBE cells.Flow cytometry revealed a marked decrease in the amount of cells in the S stage of the cell division cycle after LINC-PINT was knocked down,when compared to the control group.In addition,GO enrichment analysis showed that LINC-PINT was mainly involved in the activation of white blood cells and the activation of cells involved in immune response.In terms of cellular localization,most of them were located in the cytoplasm.In terms of molecular function,more cellular components are involved.KEGG pathway enrichment analysis showed that it was involved in MAPK signaling pathway,Rap1 signaling pathway,and transcriptional dysregulation in cancer.Lnc Locator predicts that LINC-PINT might exist both inside and outside the nucleus.Conclusions: LINC-PINT is highly expressed in cholangiocarcinoma cells.The proliferation,migration and invasion of cholangiocarcinoma cells are obviously inhibited after LINC-PINT knockdown.It may regulate the expression of genes in the nucleus or regulate the post-transcriptional level of genes in the cytoplasm by interacting with DNA,RNA and proteins in the nucleus.LINC-PINT has the potential to become an effective biomarker for early diagnosis and treatment of cholangiocarcinoma,and its specific molecular mechanism remains to be further explored.