Phenotypic And Functional Perturbations Of B Lymphocytes In Chronic HIV-1Infection

Background:Human Immunodeficiency Virus type Ⅰ (HIV-1) infection leads to severe immune dysfunction. In addition to the progressive depletion and dysfunction of CD4+T cells, HIV-1infection also leads to extensive defects in the humoral arm of the immune system. Decreased B cell counts circulating in the peripheral blood and perturbed B-cell subpopulations including over-representation of activated, exhausted and terminally differentiated B cells; over-representation of immature/transitional B cells; and reduced representation of CD27+memory B cells are observed in HIV-1-infected individuals. In addition, functional perturbations of B cells include increased expression of activated, exhausted and apoptotic molecules and decreased expression of anti-apoptotic molecules, as well as polyclonal activation and poor responses to specific or non-HIV antigens. Participants in these studies are from different districts and in different disease progression stages, and the molecular feature of prevalent virus is distinctive. Particularly, after a period of ART, the immunological characterization and mechanism of phenotypic and functional perturbations within B-cell subpopulations, as well as their association with disease progression from chronically HIV-1-infected patients remain to study, in order to establish effective immune intervention strategies to restore perturbed phenotype and functionality of B cells.The participants of our study were former blood donors donated their plasma between1995and1996in China and were infected with HIV from common-source of contaminated blood. Due to the close demography, genetic background and molecular feature of virus, we wish to provide new insights into the studies mentioned above.Methods:Written informed consents were obtained from all participants. The participants were divided into three group:ART-naive HIV-viremic (VI),-ART-treated HIV-aviremic (AV) and HIV-negative (HN). Whole blood samples were collected and then used to enrich peripheral blood mononuclear cell (PBMC) by Ficoll gradient centrifugation.Flow cytometry was employed to determine the differences in the frequencies of B-cell subpopulations:immature B cells (CD19+CD10+CD27-), naive B cells (CD19+CD10-CD21+CD27-), resting memory B cells (CD19+CD10-CD21+CD27+), activated memory B cells (CD19+CD10-CD21-CD27+), tissue-like memory B cells (CD19+CD10-CD21-CD27-) and plasmablast (CD19+CD10’CD21-CD27+CD20-), The expression levels of activated molecule CD38, cell turnover marker Ki-67, apoptosis-associated molecules (CD95, Bcl-2and PD-1) and costimulatory molecules (CD40and CD70) on each B-cell subpopulations were also measured among the three groups to investigate the abnormalities of B-cell phenotype and the restoration of ARTIn addition, functionality study was proceeded on enriched B cells:HIV-1gpl20-specific or influenza HA-specific antibody secreting cells (ASCs) were quantified by Enzyme-linked Immunospot Assay (ELISpot); the levels of cytokines were measure by cytometric beads assay (CBA).Results:HIV-1infection led to decreased counts of CD4+T cells and B cells in peripheral blood (422±33cells/μl vs814±57cells/μl, p<0.01;156±19cells/μl vs216±33cells/μl, p=0.05), and a positive correlation was observed between them in Ⅵ (r=0.376, p=0.037). A slight restoration of CD4+T cells and B cells counts were observed after ART. The frequencies of immature B, naive B, resting memory B cells and plasmablast were significantly decreased, whereas that of tissue-like memory B cells was increased in VI, comparing with HN. However, ART can restore the alterations of naive B cells and tissue-like memory B cells, but resting memory B cells still remained low.During HIV-1infection, significantly high expression of activated molecule CD38was observed on immature B, naive B, resting memory B and tissue-like memory B cells. Meanwhile, the positive correlation of CD38and cell turnover marker Ki-67was observed in VI (r=0.432, p<0.01). In addition, all B-cell subpopulations exhibited increased levels of CD95, and resting memory B cells, activated memory B cells and plasmablast showed quite high levels of CD95expression; whereas, low levels of Bcl-2were on naive B cells, tissue-like memory B cells and plasmablast. Resting memory B cells and plasmablast showed elevated levels of PD-1. And costimulatory molecule CD40expression was downregulated on all B-cell subpopulations, whereas CD70expression was decreased on mature B-cell subpopulations, including naive B cells, resting memory B cells, activated memory B cells and tissue-like memory B cells. However, the abnormalities of activated, apoptotic and costimulatory molecules expression on B cells and B-cell subpopulations in HIV-1infection can be partically restored by successful ART.HIV-1infection resulted in impaired B cell function, as manifested by the lower frequencies of total IgG antibody-secreting cells (ASCs), when compared with HN (3818±931ASCs/106B cells vs10600±2623ASCs/106B cells, p=0.05); in addition, the frequencies of HA-specific ASCs were significantly lower in VI than in HN (22±7ASCs/106B cells vs53±10ASCs/106B cells, p=0.028), however, no elevated frequencies of total IgG and HA-specific ASCs were observed after successful ART. The frequencies of gp120-specific ASCs increased in HIV-1infection, but decreased after initiation of ART (163±58ASCs/106B cells vs30±8ASCs/106B cells, p<0.01). The ability of cytokine secretion of B cells also impaired, for the lower levels of cytokines IL-10, IL-6, IL-8, LT-a and CCL-3in VI.Conclusions:HTV-1chronic infection leads to diminished peripheral B cell counts and perturbations of B-cell subpopulations. In addition, B-cell subpopulations from HTV-viremic individuals exhibit high levels of activation, increased susceptibility to apoptosis and impaired interaction between B and T cells.Defected function of B cells in HIV-1-infected individuals includes decreased HIV or non-HIV antigens specific ASCs and impaired ability to secret cytokines.Initiation of ART could partially restore the perturbed phenotype and functionality of B cells from HIV-1-infected individuals.