Hsa＿circ＿0006091 As A Molecular Marker In Hepatocellular Carcinoma And Exploration Of Its Mechanism

Research background: The incidence and mortality of liver cancer have been rising slowly in recent years,and hepatocellular carcinoma(HCC)is a type of liver cancer,which accounts for the vast majority of all types.The key to determining whether patients with liver cancer can undergo surgery,their survival time after surgery,and their quality of life lies in early diagnosis.Early diagnosis requires specific and sensitive markers.At present,the most commonly used clinical diagnostic marker for HCC is alpha-fetoprotein(AFP),but its sensitivity and specificity are not ideal.Therefore,we are committed to selecting new HCC biomarkers.As a new type of non-codingRNA,circularRNA(circRNA)has been one of the focuses of scientific research in recent years,and it will also become the target of our research.Purpose: In humans,circRNA is considered to be differentially expressed in various tissues,including liver cancer tissues,and is a stable and widespread non-codingRNA molecule.In the existing research,we aim to study circ-RNA as a new candidate biomarker for hepatocellular carcinoma(HCC),and to make a preliminary exploration of its mechanism of action.Methods: Through high-throughput sequencing and data analysis of three groups of HCC and adjacent tissues,the differentially expressed circRNAs in liver cancer and adjacent healthy tissues were identified.Reverse transcription polymerase chain reaction(RT-PCR)and quantitative polymerase chain reaction(QPCR)were used to evaluate the elevated levels of circRNA in HCC.The results show that there are significant differences in the expression of hsa＿circ＿0006091 in HCC.Then we obtained its target gene through biological analysis,and conducted preliminary verification of its target gene.The regulator of G protein signal 12(RGS12)showed a higher expression level in HCC tissues.Receiver operating characteristics(ROC) curve analysis was performed on hsa＿circ＿0006091 and RGS12,and joint analysis was performed.Finally,we use cell scratch experiment,cell proliferation experiment(CCK8 method)and apoptosis detection experiment to explore the effect of knocking down the expression of hsa＿circ＿0006091 on the migration ability,proliferation and apoptosis of HCC cells.Results:(1): The three groups of HCC and adjacent normal tissues were sequenced by high-throughput sequencing technology,and the result data was analyzed and screened.Finally,4 up-regulated and 19 down-regulated differential circRNAs were obtained.(2)Validation of HCC and adjacent tissues by q PCR confirmed that the expression level of hsa＿circ＿0006091 in liver cancer tissues was higher than that in adjacent tissues.(3)By predicting the Circ-0006091-miRNA-mRNA network pathway and downstream genes,it is concluded that his target genes may be QKI,CTNNA1,GSK-3b and RGS12,and then the q PCR verification shows that RGS12 is highly expressed in liver cancer tissues.(4)Simultaneously verify the serum of HCC patients before and after surgery and normal humans.The same results show that hsa＿circ＿0006091 and RGS12 are highly expressed in HCC patients before operation,and low in serum after operation and in normal people.Then analyze them separately by ROC curve.The results showed that the combined analysis of hsa＿circ＿0006091 and RGS12 yielded an ROC value of 0.92,which has high sensitivity and specificity.(5)Validation of various cell lines of HCC shows that hsa＿circ＿0006091 is highly expressed in various cell lines,among which SMMC-7721 cell line has the highest expression abundance.And use small interferingRNA(siRNA)transfection to down-regulate the expression of hsa＿circ＿0006091 in SMMC-7721 cells.(6)Down-regulating the expression of hsa＿circ＿0006091 has no significant difference in the effects of HCC cell proliferation and apoptosis,but it significantly delays the migration of HCC cells.Conclusion: The existing research results show that the expression levels of hsa＿circ＿0006091 and RGS12 in liver cancer tissues are elevated.In addition,has＿circ＿0006091 can be used as a new candidate biomarker for liver cancer,which can improve the diagnosis strategy,prediction and follow-up of liver cancer patients.The combined diagnosis of the two has diagnostic significance and can be used as a molecular marker for the diagnosis of liver cancer.At the same time,hsa＿circ＿0006091 may participate in and promote the occurrence and development of HCC by enhancing the migration ability of tumor cells.