Analysis On The Relationship Between Serum MicroRNA And Occult Hepatitis B Infection

Occult hepatitis B infection（OBI）is one of the symptoms of hepatitis B virus（HBV）infection.It has been defined as the“presence of HBV viral DNA in the liver（with or without detectable HBV DNA in serum）of HBs Ag-negative individuals tested with the currently available serum assays”.According to epidemiological investigation,the prevalence of OBI is about 5%²3%in Chinese"healthy"blood donation population.Therefore,OBI acts as a hidden danger in China because it is a potential source of HBV infection.As the“silence threaten”,OBI is an important problem for the prevention and control of HBV infection.It is very important to understand the epidemiological characteristics of OBI for the further control of HBV transmission.With the development of molecular biology technology,more and more recent studies focused on the relationships between microRNAs and HBV infections.Amongst them,some researches showed that miR-122 plays an important role in the occurrence and development of hepatitis B related cirrhosis/cancer by influencing the HBV replication and the host immune responses.Additionally,some studies indicated that miR-130a could inhibit the HBV replication through regulating HBV associated estrogen receptor alpha in human hepatocellular carcinoma cells.Based on the“135”cohort study in Suzhou Industrial Park,we performed the present study to explore the epidemiological characteristics of OBI,and analyze the expression profiles of serum miR-122,miR-130a and their diagnostic values in OBI.The main results as following:PartⅠThe analysis of epidemiological characteristics of OBIObjective:To investigate the epidemiological characteristics of OBI in Suzhou industrial park.Subjects and Methods:3809 persons（36⁸7 years）were recruited from“135”project in Suzhou Industrial Park.Data on demographic characteristics,history of diseases,life styles were collected.HBsAg and HBsAb were detected by enzyme-linked immunosorbent assay（ELISA）.HBV virus nucleic acid was detected,quantitated and genotyped by TagMan Real-time PCR.Epidata 3.0 software was used for data importing.Chi-square test（χ²）was used to examine differences in the categorical data distribution.The continuous variables were described with mean and standard deviation（mean±SD）and tested difference in groups using two-sample t test or ANOVA when they followed normal distributions.The median or interquartile range was calculated for continuous variables with abnormal distribution.All p values were based on a 2-sided test and a significance level of 0.05.Results:1.Among 3809 subjects,60 persons were diagnosed with OBI,including 36males（0.95%）and 24 females（0.63%）.Compare to the normal group and HBs Ag positive group,the OBI patients were significantly older（P=0.0212 and P=0.0007）.Additionally,the level of ALT in OBI group was significantly higher than those in normal group.2.The prevalence of OBI was 1.18%in females.Compared to those in normal or HBsAg positive group,the HBsAb positive rate was significantly higher in female OBI patients.Additionally,the levels of ALT and AST were significantly increased.In males,the prevalence of OBI was 2.02%.Compared to those in normal or HBsAg positive group,BMI index and the HBsAb positive rate were significantly higher while the ALT level was significantly lower in male OBI patients.3.Among 219 isolated HBV in HBsAb positive groups,108 HBV strains were clustered into genotype B（49.32%）,and 111 HBV were determined as genotype C（50.68%）.Totally,29 virus strains genotyped among 60 OBI patients.2 strains were clustered into genotype B（7.42%）.23 strains belong to genotype C（85.29%）and 2cases were co-infected with genotype B and C HBV.However,the remaining 33HBV strains could not be successfully genotyped with RT-PCR.Conclusions:In the population of Suzhou industrial park,the prevalence OBI was 1.58%.The predominant virus circulated in OBI was genotype C HBV.Compared to those in HBsAg positive group and normal group,seroprevalence of HBsAb and the level of AST were higher in OBI patients.PartⅡAnalysis on the relationship between serum miR-122 and miR-130a and Occult hepatitis B infectionObjective:To analyze the expression profiles of serum miR-122 and miR-130a in OBI and explore their diagnostic values in OBI diagnosis.Subjects and Methods:110 persons were randomly selected from“135”project in Suzhou Industrial Park.Amongst them,28 patients with OBI,27 cases with chronic asymptomatic HBV carriers（ASC）,28 cases of patients with chronic hepatitis B（CHB）and 27 cases were healthy controls（HC）.Data were analyzed using SAS 9.4 software.Comparisons of differences in the categorical data between groups were performed usingχ² test or Fisher’s exact test,multivariate analyses using logistic regression.The one-way ANOVA test was used to analyze continuous variables where appropriate,multivariate analyses using covariance analyze.All statistical tests were two-tailed,and a probability level of P<0.05 was considered as statistically significantly.Results:1.The expression level of serum miR-122 was gradually increased from OBI group to HC,ASC and CHB groups（F=21.58,P<0.0001）.However,the expression level of serum miR-130a and the value of miR-130a/mi R-122 in OBI group were significantly higher than those in other three groups.2.There was a significantly correlation between serum miR-122 level and HBV DNA level（P<0.001,r=0.73）.However,the serum miR-130a level was not correlated with HBV DNA levels.3.MiR-130a alone or in combination with AST and ALT could separate OBI from HC,OBI and CHB with≥0.61 of the area under the curve（AUC）.MiR-122 or in combination with AST and ALT could differently diagnose OBI from HC,ASC and CHB with≥0.75 of AUC.The ratio of mi R-130a to miR-122 alone or combined with AST and ALT could separate OBI from HC,ASC and CHB with≥0.86.Moreover,miR-130a and miR-122 or in combination with AST and ALT could differential diagnose OBI from HC,ASC and CHB with≥0.87.Conclusions:1.The expression level of miR-122 in OBI patients was significantly higher than those in HC,ASC and CHB patients.However,the expression level of mi R-130a was significantly lower in OBI patients compared to those in HC,ASC and CHB patients.2.MiR-130a combined with miR-122 could successfully separate OBI from CH,ASC and CHB.MiR-130a and miR-122 combined with AST,ALT could significantly diagnose OBI from HC,ASC and CHB.