Detection Of The Expression Level Of MiR-122in Liver Diseases Serum By Taqman Probe FQ-PCR And Its Clinical Significance

ObjectiveEstablishing Taqman probe real-time fluorescence quantitative PCR（FQ-PCR）to detectthe level of miR-122expression in the serum,selecting the optimal concentration of TaqmanDNA polymerase and doing the feasibility study of using U6snRNA as the reference，andapplying the method to detect the expression level of serum miR-122in patients with liverdiseases，and discussing its clinical significance.Methods1. Establishing Taqman probe FQ-PCR to detect the expression level of serum miR-122,selecting the optimal concentration of Taqman DNA polymerase and doing the feasibilitystudy of using U6snRNA as the reference.2. Design the stem-loop RT primer, PCR primer and Taqman probe of miR-122andU6snRNA,by using Taqman probe FQ-PCR to detect the expression level of miR-122in theserum from27cases of hepatocellular carcinoma,15cases of hepatitis B,15cases ofhepatitis C,15cases healthy control，15cases of postoperative hepatocellular carcinoma and15reoccurred cases of postoperative hepatocellular carcinoma, using U6snRNA as theinternal control.3. Study the relationship between serum miR-122and liver disease markers(HBV DNA，HCVRNA， ALT, HBsAg, HBeAg, HCV-Ab and AFP).Results1. The most suitable concentration of Taqman DNA polymerase is0.0625U/μl and U6snRNAcan be used as the reference serum miR-122.2. The serum miR-122expression level in hepatocellular carcinoma, hepatitis B, hepatitis Cand recurrence patients are higher than that in healthy control and postoperativehepatocellular carcinoma（P<0.05）.Meanwhile, the level of serum miR-122increases in thehepatitis C than that of hepatocellular carcinoma，hepatitis B and recurrence patients（P<0.05）, but there is no any difference among hepatocellular carcinoma，hepatitis B andrecurrence patients of miR-122expression level in serum（P=0.12）, the serum level of miR-122is lower in postoperative hepatocellular carcinoma patients compare with that ofhepatocellular carcinoma and recurrence patients（P<0.05）.3. In hepatitis B virus surface antigen（HBsAg）and/or hepatitis B virus e antigen（HBeAg）positive patients the serum level of miR-122is higher than negative ones（P<0.05）, and thelevel of serum miR-122raises in hepatitis C antibody（HCV-Ab） positive patientscomparing with negative ones（P<0.05）. The serum level of alanine transaminase（ALT）and the serum miR-122expression are positive correlation（r=0.34，P<0.05）. The miR-122expression level in serum alpha fetal protein （AFP）≥400ug/L patients is higher than thatin the patients of serum AFP <400ug/L（P<0.05）.ConclusionsThe Taqman probe FQ-PCR can apply to detect the serum miR-122expression level. Andthe serum miR-122might be used as a new biomarker of liver diseases, especially in earlydiagnosis of primary hepatocellular carcinoma, curative effect of surgical operation and theprognosis.