| Part1Expression and significance of miR-183in hepatocellular carcinomaMicroRNAs (miRNAs) are small endogenous, noncoding RNAs, which are deregulated in tumor tissues, their deregulation acts as oncogenes or oncosuppressors in cancer onset and progression. Many studies have shown that microRNAs (miRNAs) are deregulated in HCC, the expression of miRNAs is associated with clinicopathologic factors, and may be new marker for diagnosis and prognosis.Objective:In our previous study, we found some miRNAs were revealed to be deregulated in hepatocellular carcinoma (HCC), including miR-183. However, the expression of miR-183in the progression of benign liver diseases to HCC and its correlation with clinicopathologic factors remain undefined. This study aimed to investigate miR-183expression and it correlation with clinicopathologic factors. And for further study of serum/plasma miRNA as biomarker of liver disease, the method for detection of serum/plasma miRNA had been optimizedMethods:MiR-183expression was measured in normal controls tissues (NC)(n=21), chronic viral hepatitis B or C (CH) tissues (n=10), liver cirrhosis (LC) tissues (n=18), HCC tissues (n=92) and adjacent non-tumor tissues (NT)(n=92) by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The expression of miR-183was calculated using2-ΔΔACT method.Results:The expression levels of miR-183were1.254(0.415-2.592),2.825(1.964～4.670),2.249(0.868～4.821),4.121(1.609～10.890) and9.015(2.687～28.786) in NL, CH, LC, NT and HCC, respectively. MiR-183expression was significantly higher in HCC than NC, LC, CH and NL (P=0.001, P<0.001, P=0.011, P<0.001respectively), no significant differences were found among CH, LC and NT. The up-regulated miR-183in HCC was correlated with TNM stage (P=0.042) and cirrhosis (P=0.025). There was no correlation between miR-183levels and other clinicopathological factors, such as patient age, sex, hepatitis B or C virus status, Child-Pugh score, AFP levels, tumor size, tumor number, vein invasion, tumor grade. Kaplan-Meier survival analysis showed that miR-183expression was not associated with survival of HCC patients. Multivariate analysis revealed that growth pattern and vein invasion were independent prognosis factors for HCC, but not miR-183. However, Receiver operating characteristic (ROC) curve analysis suggested that miR-183yielded an area under the curve (AUC) of0.808with59.8%sensitivity and91.8%specificity in discriminating HCC from benign liver diseases (CH and LC) or NC.Conclusions:The up-regulated miR-183may associate with onset and progression of HCC, but not patient survival. Further research is needed to determine the potential of miR-183as biomarker for HCC.Part2Study on detection of serum/plasma miRNA of liver diseases MiRNAs are present in serum and plasma in a stable form that is protected from low/high PH, extend storage, freeze-thaw cycles, and so on. MiRNAs are differential expressed in different diseases. Base on these characteristics, serum/plasma miRNAs may be new biomarker for tumor.Objective:For further study of serum/plasma miRNA as biomarker of liver disease, the method for detection of serum/plasma miRNA had been optimizedMethods:The miRNA extracted from normal controls serum/plasma (n=21), chronic viral hepatitis B or C (CH) serum/plasma (n=19), liver cirrhosis (LC) serum/plasma (n=12) and HCC serum/plasma (n=19) by miRcute miRNA isolation kit; The miRNA extracted from HCC serum/plasma (n=19) by miRNeasy serum/plasma kit. The extractions of serum/plasma were measured by spectrophotometer. U6and miR-183expression were measured by qRT-PCR. The expression of miR-183was calculated using2-ΔΔCT method.Results:MiRNAs extracted by miRNeasy serum/plasma kit (A260,260/280, 260/230, U6and miR-183) were better than miRcute miRNA isolation kit (P<0.05, resPectively). All Ct values were lower than35. The quantity of U6in plasma was more than serum both NC and LC (P=0.000, P=0.011). There was no difference between plasma and serum in HV (P=0.312); In HCC, the quantity in serum was more than plasma (.P=0.022); The expression levels of serum miR-183were2.06(0.271-4.429),4.427(1.580-5.372),5.203(0.751-18.472) and2.534(0.332-10.885) in NC, HV, LC and HCC, no difference was found among groups (P>0.05, respectively). The expression levels of plasma miR-183were1.738(0.455-3.722),8.202(4.138-20.591),1.241(0.577-8.073) and33.081(24.606-86.759) in NC HV LC and HCC, MiR-183expression was significantly higher in HCC than NC, HV and LC CP=0.000, P=0.008, P=0.000). Plasma miR-183discriminated HCC from NC, HV and LC, with AUC of ROC of0.837(95%CI,0.713-0.961), at the cutoff value of3.955, the sensitivity was78.9, and the specificity was81.0%. Serum miR-183discriminated HCC from NC, HV and LC, with AUC of0.662(95%CI:0.488-0.835), at the cutoff value of4.272, the sensitivity and specificity were57.9%and76.2%, respectively.Conclusions:MiRNeasy serum/plasma kit is better than miRcute miRNA isolation kit, which used to extract serum/plasma miRNA, but both of them can be used for miRNA extraction. Plasma samples were more suitable for the research of miRNA as diagnosis marker of liver HCC. |
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