The Study Of Serum MiR-21 Detected By Real Time Fluorescent Quantitative PCR In The Diagnosis Of Hepatic Fibrosis

Background: Hepatic fibrosis is the common pathological process of chronic liver insults,mainly characterized by the excessive deposition of extracellar matrix.This pathologic change can lead to the destruction of hepatic tissue's structure and function.The process of liver fibrosis inevitably leads to cirrhosis.Finally the related serious complications will appear,such as a large number of ascites,esophageal and gastric varices bleeding,hepatic encephalopathy or other systemic dysfunction;at the same time,it may lead to the occurrence of hepatocellular carcinoma(HCC).HCC is the predominant primary liver cancer in many countries and is the third most common cause of cancer-related death in the Asia-Pacific region.As the main risk factor for liver fibrosis and cirrhosis,Chronic hepatitis B(chronic hepatitis,CHB)is prevalent in the world.Even though the vaccine has been widely used,a large number of people are still infected.Chronic HCV infection is now emerging as an important cause of chronic liver diseases.Unlike patients with chronic hepatitis B,those with chronic hepatitis C(CHC)develop HCC in the presence of established cirrhosis in most cases.Because there is no special clinical manifestation in the early stage of liver fibrosis,early diagnosis of liver fibrosis and finding the relatively simple method are of great significance to prevent liver cirrhosis.At present,liver biopsy is still the the "gold standard" to diagnose liver fibrosis and cirrhosis,but this inspection method is invasive,and it may cause serious complications,such as hemorrhage,infection,and so on.The biggest fear among patients thinking of this check is that may cause the death.There exist some of the sampling error check and reading errors,So this inspection is not acceptable for the majority of patients with liver disease.In recent years,the Liver transient elastography(transient elastography TE)has been widely used in clinical.This examination is of good tolerability,non-invasive and reproducible,but the accuracy of the results is also affected by many factors,such as a large amount of ascites,too narrow intercostal space,which may cause the operation failing.There are many factors such as liver inflammation,cholestasis,liver congestion,eating can affect liver stiffness.At present,the four hepatic fibrosis markers including Hyaluronic Acid(HA),Type ?procollagen N peptide(P?NP),Type ?collagen(?-C)and Laminin(LN)can reflect in varying degrees of liver pathological damage to chronic hepatitis,but they are still not suitable as a single independent serological marker for diagnosis of liver fibrosis for the poor sensitivity and specificity.Looking for high sensitivity,good specificity and non traumatic index,is a hotspot in diagnosis of hepatic fibrosis.In resent years,many scholars lead their eyeballs to miRNAs.MiRNAs are endogenous small non-coding RNAs about 22 nt in length that regulate gene expression at the posttranscriptional level and play an important role in cell proliferation,apoptosis and differentiation.And miRNA is involved in the pathological process of a variety of diseases,and it is an important candidate diagnostic marker and potential therapeutic target.It is hypothesized that miRNAs regulate 1/3 genes of human.The research of miRNAs are more common in cancer,but it is still relatively less in the field of organ fibrosis research.At present,it is found that a part of miRNAs can play a regulatory role in the progression of fibrogenesis.A number of studies have indicated that miR-21 level is upregulated during Multiple organs fibrosis.Therefore,the expression level of miR-21 in peripheral blood of patients with liver fibrosis is expected to be used as serological indicators for early detection of liver fibrosis,so that patients with liver fibrosis can get early treatment,then the disease could be well controlled.Objective: We detect the serum miR-21 expression level by fluorescent quantitative PCR.we also investigate whether the expression of miR-21 show some differences in the plasma of patients with hepatic fibrosis compared to cirrhosis?hepatocellular carcinoma and healthy controls.Finally we will evaluate the diagnostic value of serum miR-21 as molecular biomarker in patients with hepatic fibrosis.Methods:1.Patients of Jingzhou central hospital,from January 2016 to October 2016,were diagnosed as Hepatic fibrosis(F),cirrhosis(C)or Hepatomas(H)according to clinical manifestation and relevant inspections.Then 10 patients were selected in each group,and we also choosed 10 healthy people as normal control group.All patients need to sign the informed consent.And we should apply for the approval of the ethics committee of Jingzhou Central Hospital.2.Ward nurses or full-time blood sampling personnel took the intravenous blood of patients after informed consent was obtained.1 sample were collected from each person.The EDTA was used for the detection of the PCR.3.The expression of miR-21 in serum would be detected by fluorescence quantitative PCR.4.The miR-21 relative expression levels in four different groups were statistically analyzed.Then the relative expression of serum miR-21 in patients with hepatic fibrosis were analyzed with other clinical indicators.At last,we confirmed the feasibility of using mi R-21 as a serological marker for the diagnosis of liver fibrosis.Results:1.The expression level of miR-21 in serum had been detected by SYBR Green I real time fluorescent quantitative PCR.The melting curve showed a single peak,which indicated good specificity.2.There were significant difference in the relative value of miR-21 between the four groups including normal control group,liver fibrosis group,cirrhosis group and hepatocellular carcinoma group(p=0.000).3.The relative value of serum miR-21 in liver fibrosis group was higher than that in normal control group,being lower than that in liver cirrhosis group and liver cancer group(p<0.05).4.The expression of miR-21 showed no significant correlation with age,sex,LN,P?NP or ?-C(p>0.05),and it shows positive correlation with HA(p<0.05).Conclusion:1.As the method detecting the expression level of serum miR-21,The SYBR Green I fluorescence quantitative PCR has advantages of being rapid,sensitive and specific.2.The detection of human serum miR-21 is helpful for the early diagnosis of liver fibrosis;serum miR-21 is closely related to the occurrence and development of liver disease,which can provide the molecular basis for the development of liver disease.