Relationship Between CD4/SD8 Ratio And ART Adverse Outcomes In HIV-1 Infected Patients And The Molecular Mechanism Of Plasmacytoid Dendritic Cells Depletion

Objective CD4/CD8 ratio is independently associated with mortality and morbidity of immune deficiency and metabolic syndrome in HIV-1 infected patients,and is considered as an emerging biomarker of HIV-related diseases.This study was designed to evaluate the role of CD4/CD8 ratio in adverse outcomes in HIV-1 infected patients with antiretroviral therapy(ART)from the perspectives of macro survival outcome and micro peripheral blood plasmacytoid dendritic cells(p DCs)depletion.Furthermore,the molecular mechanism of pyroptosis mediated by HMGB1/Caspase-1/GSDMD signal axis in the p DCs depletion in HIV-1 infected patients was explored.This study provides new research ideas for the use of CD4/CD8 ratio as an important index to evaluate the adverse outcome of ART in HIV-1 infected patients,and also for the immune reconstitution of these patients.Methods Firstly,a retrospective cohort study was conducted based on the ART data of HIV-1 infected patients in Guangxi from December 2003 to October 2020,and the all-cause mortality of the patients was analyzed by grouping according to the recovery of CD4/CD8 ratio.Kaplan-Meier was used to evaluate the cumulative risk of all-cause death during ART.Cox regression model was used to explore the influencing factors of all-cause death in patients.Propensity score matching was then used for further verification.Also,the changes of CD4/CD8 ratio during ART were visualized analyzed.Secondly,flow cytometry and some other experiments were used to analyze the differences of p DCs frequency and pyroptosis level between HIV-1 infected patients and healthy donors.Bivariate correlation analysis was used to analyze the relationship between p DCs frequency and CD4/CD8 in HIV-1 infected patients.Furthermore,according to flow cytometry,RT-PCR,ELISA,WB and other experiments,in vitro differentiation of THP1-DCs cell model was used to detect the pyroptosis,HMGB1 release and caspase-1/GSDMD signal axis activation of HIV-1 infected/un-infected THP1-DCs.Results(1)The cumulative risk of all-cause death was higher in patients whose CD4/CD8 ratio didn’t recovery during ART than in patients with CD4/CD8 ratio recovery(Log rank:P<0.0001;total study population:a HR=2.72,95%CI:2.55-2.90;PSM population:a HR=2.83,95%CI:2.60-3.08).(2)The risk of all-cause death was lower(a HR=0.41,95%CI:0.38-0.44)in PIs-based group than that the NNRTIs-based group.(3)The risk of all-cause death in patients with ART immunological failure was lower in the PIs-based group than the NNRTIs-based group(total population:a HR=0.32,95%CI:0.29-0.36;PSM population:a HR=0.40,95%CI:0.32-0.48).However,CD4/CD8ratio recovery reduced the risk of all-cause death regardless of which ART regimen was used(Log rank:P<0.0001,total study population:a HR=2.74,95%CI:2.44-3.09;PSM population:a HR=2.80,95%CI:2.43-3.23).(4)With the prolongation of ART time,the median CD4~+T cell counts of the patients increased to normal,while CD8~+T cell counts remained high,resulting in the slowly recovery of CD4/CD8 ratio.CD8~+T cell counts decreased and CD4/CD8ratio recovered more significantly in NNRTIs-based group than in PIs-based group.(5)p DCs frequency in PBMCs of HIV-1 infected patients was significantly lower than that of healthy donors(P=0.001),and there was a positive correlation between p DCs frequency and CD4/CD8 ratio(r=0.688,P=0.001).(6)HIV-1 activated p DCs to secrete IFN-αand up-regulated p DCs pyroptosis level and also the m RNA level of Caspase-1、GSDMD、IL-1βand HMGB1.(7)HIV-1 infection promoted THP1-DCs to release HMGB1,and activated the Caspase-1/GSDMD signaling axis.Meanwhile,the expression of IL-1βand IL-18 were increased.(8)After the addition of HMGB1 inhibitor Glycyrrhizin,pyroptosis of THP1-DCs was reduced,the m RNA and/or protein expression levels of Caspase-1,GSDMD,IL-1βand IL-18 were decreased.Conclusions The recovery of CD4/CD8 ratio was correlated with the reduction of all-cause mortality risk in HIV-1 ART patients.The change of CD4/CD8 ratio was mainly affected by CD8~+T cell counts,and NNRTIs-based regimen had a better effect on reducing CD8~+T cell counts level and recovering CD4/CD8 ratio than PIs-based regimen.However,the recovery of CD4/CD8ratio significantly reduced the risk of the all-cause mortality risk regardless of ART regimen.Moreover,there was a positive correlation between p DCs frequency and CD4/CD8 ratio in HIV-1 ART patients,which further indicated that CD4/CD8 ratio could be used as an important indicator to evaluate the adverse outcomes from the perspective of microscopic of natural immune cells.HIV-1 infection induces pyroptosis of p DCs by promoting the secretion of HMGB1,which activating the Caspase-1/GSDMD signaling axis,and releasing inflammatory factors such as IL-1βand IL-18.This may be an important reason of p DCs depletion in peripheral blood of HIV-1 infected patients.