Research On The Mechanism Of Long Non-coding RNA In Hgepatocellular Carcinoma Postoperative Recurrence

Introduction : Hepatocellular carcinoma (HCC) is one of the most common malignant tumors with its increasing incidence worldly and it also brings heavy burden to the society.Although surgery is the mainstream of HCC, recurrence after surgery is extremely high,which seriously affect the tumor cure rate. The detection of predictive factors for early recurrence tumor after complete resection is clinically important for increasing the overall survival and making early treatment. Recently, multiple studies have shown that long non-coding RNA (Lnc RNA) plays important role in HCC genesis and development, along with the progress of next-generation sequencing (NGS) of and bioinformatics analysis technology.Study of the Lnc RNA uncertainty nowadays is one of the most important researching fields and it might be the new tumor marker and molecule-targeted drug. As Lnc RNA may take significant effect on the upstream gene regulation, the study of Lnc RNA has few interference. Meanwhile,it might make great contribute to the tumor prognostics and mechanisms, providing new treatment strategies of early recurrence as well.Part1 : The preparation and functional verification of differential Lnc RNA expression between neoplastic tissues and paired normal tissues.Objective: To detect and verify differential Lnc RNA expression between neoplastic tissues and paired normal tissues both in histology and cell lines, furthermore to attain the objective RNA. Methods: Using NGS and bioinformatics analysis technology, we filtered differential Lnc RNA expression and confirmed expression differentiation between normal hepatic cell lines and HCC cell lines. Results: We obtained 632 Lnc RNA and 391 mRNA that were differently expressed between tumor tissue and paired normal tissue. Among them, 7 Lnc RNA (p2609/p33560/ p3816/p40127-v4/p22534/p8751/p42540-v4) in HCC patients were demonstrated the same expression trend with mRNA, 15Lnc RNA being statistically different contrarily. According to the histological and cellular results, we found that lncRNA-C2/ LncRNA-P7and LncRNA-P12 were significantly differed and we chosen LncRNA-P7, the most obviously different express LncRNA, as the objective, and named it LINC RP1130-1. Nucleocytoplasmic separation experiments showed that LINC RP1130-1 was mainly located in cytoplasm. We suggested that LINC RP1130-1 might exert regulatory function through conjugating with some special pathways proteins. Afterwards, we studied mechanisms of LINC RP1130-1 in HCC recurrence and metastasis.Part2: LINC RP1130-1 affect HCC recurrence and metastasis though STAT1-MAPK pathway.Objective: To investigate possible pathways of LINC RP1130-1 in HCC recurrence and development. Methods: To find the LINC RP1130-1 function gene and protein targets, we used plasmid transfection, over expression and silencing gene expression, western blot, cell viability, cell proliferation and apoptosis in gene and protein levels. Results: We showed that LINC RP1130-1 promoted HCC survival and suppressed apoptosis; LINC RP1130-1 regulated ST AT1 gene transcription, and had an impact on the expression of the elated proteins; STAT1 transcription was significantly suppressed and STAT1/ERK1/2 proteins expression were reduced, as the LINC RP1130-1 increased. We suggested that LINC RP1130-1 might play an important role in the HCC recurrence and metastasis though STAT-MAPK signal pathway.Part3: The relevance of LINC RP1130-1 and clinical prognosis.Objective: To find the correlation of LINC RP1130-1 with the clinical information of HCC patient and to investigate meanings and effects of LINC RP1130-1 in clinical treatments.Methods: We detected the differential Lnc RNA expression between neoplastic tissue and paired normal tissue, and divided them into high and low groups. We statistically analyzed the patients clinical differentiation between two groups. Results: We found that LINC RP1130-1 was significantly decreased in HCC tumor tissues, compared with tissues surrounding cancer. We also revealed that LINC RP1130-1 was lower in HCC cell lines than that in in normal hepatocyte. Moreover, the disease free survival (DFS) of LINC RP1130-1-lower-expression group was significantly prolonged, compared with LINC RP1130-1-high-expressin group (P=0.047). In this study, we also found that the expression of LINC RP 1130-1 was closely correlated with tumor stage, cirrhosis, portal vein tumor thrombus,micro vascular tumor thrombus and tumor number. The area under curve (AUC) was 0.74.This suggested that LINC RP1130-1 was special and sensitive in the HCC diagnosis and could be a potential marker for early diagnosis.Conclusions: Many studies demonstrated that LncRNA was widely distribute in organism and had involved in the HCC ocurrence and progression. In our study, we found that LINC RP1130-1 was obviously decrease in the HCC cell lines, compared with normal liver cell lines. Accordingly, LINC RP1130-1 was statistically lower expressed in tumor tissues than that in paired normal tissues. Moreover, we revealed that the expression of paired normal tissues was closely correlated with tumor stage, cirrhosis, portal vein tumor thrombus,microvascular tumor thrombus and tumor number. The area under curve (AUC) was 0.74.This suggested that LINC RP1130-1 was special and sensitive in the HCC diagnosis and could be a potential marker for early diagnosis. In general, LncRNA-P7 (LINC RP1130-1)had involved the HCC occurence and disease development, and might be a new potential clinical bio-markers with great prospective.