Study On Specific MiRNA Markers Related To Immune Reconstitution In HIV-infected Patients

Purpose:Human Immunodeficiency Virus?HIV?infection seriously endangers human health and life.A large number of studies have confirmed that HIV-infected patients with highly active antiretroviral therapy?HAART?as soon as possible have significant significance in reducing HIV-related morbidity and mortality,reconstructing immune function and reducing HIV transmission.After antiviral treatment,the abnormal immune changes caused by HIV can return to normal or near normal levels,that is,immune function reconstruction,includes of the recovery of the number and function of CD4⁺T lymphocytes.However,it is estimated that about 20%of HIV-infected patients have been inhibited by viral replication in their bodies,but the reconstruction of immune function has not reached normal ideal status.This part of patients is called immunological non-responders?INRs?.The risk of morbidity and mortality of INRs was significantly higher than that of patients with immune reconstitution.A variety of factors are known to affect the immune function reconstitution after antiviral therapy,but the mechanism by which viral replication has been inhibited in patients after antiviral therapy but immune function remains unrecoverable is unclear.MicroRNAs?miRNAs?are a class of highly conserved single-stranded small-molecule RNAs that,as important members of non-coding RNAs,play a key role in the development and progression of HIV infection,although they are only18-25 bases..MicroRNAs inhibit or promote HIV infection.Many studies have found different miRNA profiles at different stages of disease progression.However,specific markers for effective early prediction of immune function reconstitution in HIV patients have not been reported.In this study,real-time quantitative PCR was used to detect the expression of miRNA in peripheral blood of patients with different immune function after antiviral therapy,and to screen out miRNAs that can predict the difference in immune function of HIV-infected patients after antiviral therapy.Method:1.Study subjects:The study group included 268 HIV-infected patients receiving HAART,and 5 healthy people.All patients were diagnosed with HIV by immunoblotting.The HIV-infected patients were divided into two groups,the immune non-response group?INR?was 90?the increase CD4⁺T cell count after treatment for one year was less than 20%of the baseline CD4⁺T cells?,and the immune response group?IR?was 178?the increase CD4⁺T cell count after treatment for one year was greater than 30%of the baseline CD4⁺T cells?.2.Experimental procedure:Take 10ml of peripheral frozen blood samples at the time before treatment of HIV-infected patients;total plasma miRNA was extracted;reverse transcription of miRNA was performed;real-time fluorescence quantitative PCR was used to detect miRNA expression levels at various stages.The relative expression level of miRNA was calculated using the formula 2^-??CT.Statistical analysis:In the Statminer of TIBCO Spotfire 7.11,miRNAs with significant differential expression between groups and their fold change were calculated using LIMMA and 2-??CT.SPSS 19.0 for data analysis.Drawing was performed using GraphPad Prism 7.0 drawing software.Data are expressed as?x±s,P<0.05 indicates statistically significant difference.Result:1.444 miRNAs were detected in three stages:discovery stage,training stage and validation stage.Seven miRNAs were obtained,which were significantly different between INR and IR patients,and were up-regulated in INR patients.2.The area under the curve of the 7 miRNA combined diagnosis INR in the ROC analysis was 0.936,the sensitivity and specificity were 0.878 and 0.906.The area under the curve of the 7 miRNA combined diagnosis INR in the ROC analysis of the confirmation stage was 0.865.Sensitivity and specificity were 0.786 and 0.860.3.The expression levels of 7 miRNAs were negatively correlated with CD4+T cell counts after 12 months of treatment?P<0.05?,and negatively correlated with the percentage increase of CD4+T cell counts after treatment?P<0.05?.Conclusion:Seven miRNAs were significantly different in the plasma of INR and IR patients,and the expression level of INR patients was higher than IR patients.The combination of 7 miRNAs can predict the effect of immune reconstitution in HIV-infected patients after HAART treatment in the early stage of infection.