| Backgroud: Hepatocellular carcinoma(HCC) is one of the most common malignancy in digestive system and the third leading cause of cancer mortality worldwide. Early diagnosis of HCC is associated with favorable prognosis. Unfortunately, HCC is often asymptotic, patients diagnosed at the advanced stage when the prospects for cure are dismal. Traditional serological markers of HCC is based on alpha-fetoprotein(AFP), but the sensitivity and specificity of it have not been ideal. Therefore, searching new and noninvasive biomarkers in serum will have a significant impact on patient diagnosis and therapy. A large number of research have found that mi RNAs play an important role in tumor, it specially expressed in various tissues and could be a new potential biomarker, and exhibit tissue and cell specificity, making mi RNAs be the potential biomarker. Mi RNAs are a group of small nocoding RNAs, which become a hot spot because of numorous target genes and complicated regulation mechanism in recent research. short, nocoding RNAs with numorous target genes and complicated regulation mechanism, making them being a hot area of research recently. Studies have showed that several signaling pathway were activated abnormally in HCC, such as Notch, Wnt and Hedgehog(Hh)signaling pathway. Hedgehog is a highly conserved cell signaling pathway that plays an important roles in embryogenesis and oncogenesis. Gli1 is nuclear transcription factor of Hh pathway, and it directly regulate downstream target genes expression. Numerous documents have found that mi RNA could targetedly suppressed Hh pathway, and play an important role in oncogenesis and tumor development. Among all the HCC related mi RNAs, mi R-202 of which targeted regulate Gli1 in HCC tissues was reported only once.), and so far there is no reaserch to report mi R-202 level in HCC serum and its connection with clinicopathologic characteristic of HCC, and its interassociation with Hh pathway in HCC oncogenesis and develoment. This study aim to investgate the expression and clinical significance of mi R-202 in HCC, and the regulation mechanism of mi R-202 together with Hh pathway in HCC. Objective: To detect mi R-202 expression in HCC cells, FFPE tissues and serum,and analyse the relationship between clinicopathological parameters and the expression level of mi R-202 in HCC, whether mi R-202 can be a potential specific diagnostic marker for HCC, and discuss the link between Hh pathway and mi R-202 in HCC, provide the basis for investigating the molecular mechanism of HCC for further study. Methods: Real-time PCR( RT-PCR) was used to detect the expression of mi R-202 in 3 HCC cell lines(Huh7, CMCC7721, Hep G2), HCC Formalin Fixed Paraffin Embedded(FFPE) tissues and HCC serum, respectively, and investigate the relationship between clinicopathological parameters and the expression level of mi R-202 in HCC. We used ROC curves to compare the ability of serum mi R-202 to discriminate between patients with HCC and controls with that of circulating AFP, discuss whether mi R-202 can be a potential diagnostic marker for HCC. U6 was used as internal control, LO2 cell, normal hetatic FFPE tissue and healthy volunteers’ serum were used as control, respectively. In addition, to detect Gli1 expression in HCC FFPE tissue, and discuss the link between changes of GLI1 gene expression and mi R-202 in HCC. Three HCC cell lines and LO2 cell was presented by the Key Laboratory of Clinical Laboratory Diagnostics of Chongqing Medical University. 70 cases of primary HCC patients included in this study were selected from the Affiliated Hospital of Si Chuan Medicine University from July, 2013 to September, 2014. Results: 1.The mi R-202 level in 3 HCC cell lines were all significantly lower than in LO2 cell line(P<0.05),and the RQ of mi R-202 was lowest in Hep G2 cell line(0.0394±0.0141); mi R-202 expression in HCC FFPE tissueand serum were both significantly lower than in healthy control(P<0.05); The mi R-202 expression in HCC FFPE tissue did not correlate with any clinicopathological parameters such as sex, age, tumor location, tumor maximal diameter, clinical TNM stage, metastasis or not, the level of serum AFP, et al(P>0.05), and that in HCC serum was relevant to tumor nodes only; The RQ of mi R-202 in HCC serum(0.33±0.30) were significantly lower than in HCC FFPE tissue(0.51±0.26)(P<0.05). 3. Combined detection of mi R-202 and tumor marker AFP could get the higher sensitivity(81.8%) and specificity(93.0%) in the diagnosis of HCC, the AUC is 0.942, which is higher than that of single gene with specificity, Youden index, and positive likelihood/negative likelihood ration. 4. The expression of Gli1 m RNA in HCC FFPE tissues(3.45±2.07) was significantly higher than normal hepatic FFPE tissues(1.34±0.58)(P<0.05), Gli1 expression in HCC FFPE tissues was correlated with the expression of mi R-202 in HCC FFPE tissue(r=0.583, P=0.005). Conclusion: 1. mi R-202 may play an important role in the tumorigenesis and development of HCC, the dysregulated expression of mi R-202 in HCC may occur in early stage of tumorigenesis and was stable quietly in HCC.2. Combined detection of mi R-202 and tumor marker AFP could get the higher sensitivity and specificity than that of single AFP, indicating that combined diagnosis of mi R-202 and AFP were superior to that of single indicator. 3. Gli1, the nuclear transcription factor of hedgehog signal pathway, might play a certain role in HCC, and Gli1 expression was correlated with mi R-202 expression in HCC. |
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