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Ruffin N, Lantto R, Pensieroso S, Sammicheli S, Hejdeman B, Rethi B, Chiodi F (Karolinska Institutet, Stockholm; and South Hospital, Stockholm, Sweden). Immune activation and increased IL-21R expression are associated with the loss of memory B cells during HIV-1 infection. J Intern Med 2012; 272: 492–503. Objectives. Microbial translocation and chronic immune activation were previously shown to be associated with impairment of T cell functions and disease progression during infection with human immunodeficiency virus type-1 (HIV-1); however, their impact on B cell function and number remains unknown. By measuring markers of immune activation and molecules involved in apoptosis regulation, we have evaluated the association between microbial translocation and loss of memory B cells in HIV-1-infected patients. Methods. Markers of activation [the interleukin-21 receptor (IL-21R) and CD38] and apoptosis (Bim, Bcl-2 and annexin V) were measured in B cell subpopulations by multicolour flow cytometry. Levels of soluble CD14 (sCD14) and lipopolysaccharide (LPS), measures of microbial translocation, were determined in plasma. Purified B cells were also exposed in vitro to Toll-like receptor (TLR) ligands. Results. IL-21R expression was higher in cells from HIV-1-infected patients, compared with control subjects, with the highest levels in nontreated patients. An inverse correlation was observed between IL-21R expression and percentages of circulating resting memory (RM) B cells. IL-21R-positive memory B cells were also more susceptible to spontaneous apoptosis and displayed lower levels of Bcl-2. It is interesting that the levels of sCD14, which are increased during HIV-1 infection, were correlated with decreased percentages of RM B cells and high IL-21R expression. In the plasma of HIV-1-infected individuals, a correlation was found between sCD14 and LPS levels. TLR activation of B cells in vitro resulted in IL-21R up-regulation. Conclusions. Microbial translocation and the associated immune activation during HIV-1 infection may lead to high expression levels of the IL-21R activation marker in RM B cells, a feature associated with increased apoptosis and a reduced number of these cells in the circulation.

Objective To characterize the level of immature-transitional B-cells in blood during pediatric HIV-1 infection in relation to active or suppressed viremia. We also aimed at characterizing the level of expression of CXCR4, CXCR5 and CCR7 on immature-transitional B-cells, as these receptors are important mediators for homing of B-cells. Design Forty-eight HIV-1 vertically infected children (33 viral controllers and 15 viremic patients) and 33 age-matched healthy controls were enrolled in a cross-sectional study. Methods We measured the levels of peripheral immature-transitional B-cells in all groups in relation to switched memory B-cells by flow cytometry. In parallel we evaluated CXCR4, CXCR5 and CCR7 expression on immature-transitional B-cells and measured plasma levels of CXCL12, BAFF and interleukin-7 by ELISA. Results We observed a lack of physiological age-related decline of immature-transitional B-cells in viremic children in parallel to a decreased level of switched memory B-cells. Interestingly, immature-transitional B-cells from viremic children presented with high levels of CXCR4. On the contrary, the level of CXCL12, the natural ligand for CXCR4, was lowest in the HIV-1 infected group, as compared with controls. Conclusion Control of HIV-1 viremia through antiretroviral treatment appears to be crucial in decreasing the expansion and alteration of immature-transitional B-cells.

Summary Although HIV-1 infection does not directly target B cells, B-cell numbers are reduced and their function is impaired during HIV infection. Antibody titres against antigens previously encountered through vaccination or natural infection are low in patients with HIV. Intrinsic B-cell defects might be involved in the impairment of humoral immunity during early HIV infection. Abnormal T-cell activation and the altered expression of molecules involved in the B-cell homing process cause dysfunctional interaction between T and B cells in the germinal centres of lymphoid tissues, which might impair B-cell responses during HIV infection. Class-switch recombination is also impaired in individuals with HIV. Protective immune responses against T-cell-dependent antigens, including influenza antigens, rely on the production of neutralising antibodies. Impaired B-cell responses during HIV infection could therefore hamper the effectiveness of vaccinations against seasonal influenza or the new pandemic influenza A H1N1 vaccines in individuals with HIV. By maintaining B-cell responses, highly active antiretroviral therapy might improve the efficacy of influenza vaccines in individuals with HIV.

HIV-1 infection induces a progressive disruption of the B cell compartment impairing long-term immune responses to routine immunizations. Depletion of specific memory B cell pools occurs during the 1st stages of the infection and cannot be reestablished by antiretroviral treatment. We reasoned that an early control of viral replication through treatment could preserve the normal development of the memory B cell compartment and responses to routine childhood vaccines. Accordingly, we evaluated the effects of different highly-active antiretroviral therapy (HAART) schedules in 70 HIV-1 vertically-infected pediatric subjects by B cell phenotypic analyses, antigen-specific B cell enzyme-linked immunosorbent spot (ELISpot) and ELISA for common vaccination and HIV-1 antigens. Initiation of HAART within the 1st year of life permits the normal development and maintenance of the memory B cell compartment. On the contrary, memory B cells from patients treated later in time are remarkably reduced and their function is compromised regardless of viral control. A cause for concern is that both late-treated HIV-1 controllers and noncontrollers loose protective antibody titers against common vaccination antigens. Timing of HAART initiation is the major factor predicting the longevity of B cell responses in vaccinated HIV-1-infected children.

CD70 molecules expressed by activated T cells provide potent B cell stimulatory signals. We hypothesized that an altered CD70 expression might contribute to B cell abnormalities during HIV-1 infection.CD70 expression and the functional and migratory properties of the CD4CD70 T lymphocytes were analyzed in HIV-1-infected patients and in humanized mice. Correlations were tested between CD70 expression and features of B-cell activation, apoptosis sensitivity and functional exhaustion.CD4CD70 T cells were analyzed in cohorts of CD4 T-cell lymphopenic, viremic or nonlymphopenic, nonviremic HIV-1-infected patients and in noninfected individuals. CD70 upregulation was also followed in HIV-1-infected humanized mice. CD38, CD95, LAIR1 and PD-1 expressions were monitored on B-cell subpopulations, Ki67 was assessed to estimate B-cell proliferation and antibody levels were measured in plasma.Blood CD4CD70 T-cell frequencies increased in response to CD4 T-cell depletion or high viremia levels as a possible consequence of increased activation and proliferation in this subset. CD4CD70 T cells produced T-helper 1-type cytokines and expressed chemokine receptors mobilizing toward sites of inflammation but not to lymphoid follicles. High CD70 expression was observed in HIV-1-infected humanized mice at extrafollicular sites (peritoneum, bone-marrow). CD4CD70 T-cell frequencies correlated with the expression of the activation marker CD38 and the death receptor CD95 on various memory B-cell subsets, with B-cell proliferation and with plasma IgG levels.CD4CD70 T cells may contribute to B cell hyperactivation and accelerated memory B-cell turnover during HIV-1 infection.

To evaluate the effects of switching from successful long-term protease inhibitor (PI)-based HAART to a three nucleoside reverse transcriptase inhibitor PI-sparing regimen, viral load quantification, HIV-specific lymphoproliferative assay and T-cell receptor (TCR) spectratyping were performed during 96 weeks of simplification follow-up in 19 HIV-infected children. Our data showed that simplification of therapeutic strategies acts positively on immune competence in HIV paediatric patients. Our children maintained viral suppression, increased lymphoproliferative responses and normalized TCRBV repertoire on the CD8 subset.

Introduction During HIV-1 infection the B-cell compartment undergoes profound changes towards terminal differentiation, which are only partially restored by antiretroviral therapy (cART). Materials and Methods To investigate the impact of infection as early as during primary HIV-1 infection (PHI) we assessed distribution of B-cell subsets in 19 PHI and 25 chronic HIV-1-infected (CHI) individuals before and during 48 weeks of cART as compared to healthy controls (n = 23). We also analysed Immunoglobulin-expression of memory B-cell subsets to identify alterations in Immunoglobulin-maturation. Results Determination of B-cell subsets at baseline showed that total and Naive B-cells were decreased whereas Activated Memory (AM), Tissue-like Memory (TLM) B-cells and Plasma cells were increased in both PHI and CHI patients. After 4 weeks of cART total B-cells increased, while AM, TLM B-cells and Plasma cells decreased, although without reaching normal levels in either group of individuals. This trend was maintained until week 48, though only total B-cells normalized in both PHI and CHI. Resting Memory (RM) B-cells were preserved since baseline. This subset remained stable in CHI, while was expanded by an early initiation of cART during PHI. Untreated CHI patients showed IgM-overexpression at the expenses of switched (IgM-IgD-) phenotypes of the memory subsets. Interestingly, in PHI patients a significant alteration of Immunoglobulin-expression was evident at BL in TLM cells, and after 4 weeks, despite treatment, in AM and RM subsets. After 48 weeks of therapy, Immunoglobulin-expression of AM and RM almost normalized, but remained perturbed in TLM cells in both groups. Conclusions In conclusion, aberrant activated and exhausted B-cell phenotypes rose already during PHI, while most of the alterations in Ig-expression seen in CHI appeared later, despite 4 weeks of effective cART. After 48 weeks of cART B-cell subsets distribution improved although without full normalization, while Immunoglobulin-expression normalized among AM and RM, remaining perturbed in TLM B-cells of PHI and CHI.

We evaluated the dynamics of innate and adaptive immunity in patients treated with combined antiretroviral therapy (cART) during primary human immunodeficiency virus infection (PHI), enrolled in a prospective randomized trial (MAIN, EUDRACT 2008-007004-29). After 48 weeks of cART, we documented a reduction in activated B cells and CD8 + T cells. Natural killer cell and dendritic cell frequencies were measured and a decrease in CD16 + CD56 dim with a reciprocal rise in CD56 high natural killer cells and an increase in myeloid and plasmacytoid dendritic cells were recorded. In conclusion, 48 weeks of cART during PHI showed significant benefits for both innate and adaptive immunity.

Introduction Understanding the mechanisms by which some individuals are able to naturally control HIV-1 infection is an important goal of AIDS research. We here describe the case of an HIV-1+ woman, CASE1, who has spontaneously controlled her viremia for the last 14 of her 20 years of infection. Methods CASE1 has been clinically monitored since 1993. Detailed immunological, virological and histological analyses were performed on samples obtained between 2009 and 2011. Results As for other Elite Controllers, CASE1 is characterized by low to undetectable levels of plasma HIV-1 RNA, peripheral blood mononuclear cell (PBMC) associated HIV-1 DNA and reduced in vitro susceptibility of target cells to HIV-1 infection. Furthermore, a slow rate of virus evolution was demonstrated in spite the lack of assumption of any antiretroviral agent. CASE1 failed to transmit HIV-1 to either her sexual male partner or to her child born by vaginal delivery. Normal values and ratios of T and B cells were observed, along with normal histology of the intestinal mucosa. Attempts to isolate HIV-1 from her PBMC and gut-derived cells were unsuccessful, despite expression of normal cell surface levels of CD4, CCRC5 and CXCR4. CASE1 did not produce detectable anti-HIV neutralizing antibodies in her serum or genital mucosal fluid although she displayed potent T cell responses against HIV-1 Gag and Nef. CASE1 also possessed multiple genetic polymorphisms, including HLA alleles (B*14, B*57, C*06 and C*08.02) and HLA-C single nucleotide polymorphisms (SNPs, rs9264942 C/C and rs67384697 del/del), that have been previously individually associated with spontaneous control of plasma viremia, maintenance of high CD4+ T cell counts and delayed disease progression. Conclusions CASE1 has controlled her HIV-1 viremia below the limit of detection in the absence of antiretroviral therapy for more than 14 years and has not shown any sign of immunologic deterioration or disease progression. Co-expression of multiple protective HLA alleles, HLA-C SNPs and strong T cell responses against HIV-1 proteins are the most likely explanation of this very benign case of spontaneous control of HIV-1 disease progression. Electronic supplementary material The online version of this article (doi:10.1186/s12967-014-0335-6) contains supplementary material, which is available to authorized users.

The relevance of CD4+T-cells, viral load and age in the immunological response to influenza infection and vaccination in HIV-1 infected individuals has previously been pointed out. Our study aimed at assessing, in the setting of 2009 A(H1N1)pdm09 influenza vaccination, whether quantification of activation-induced deaminase (AID) expression in blood B-cells may provide additional indications for predicting antibody response to vaccination in HIV-1 infected patients with similar CD4+T-cell counts and age. Forty-seven healthy controls, 37 ART-treated and 17 treatment-naïve HIV-1 infected patients were enrolled in the study. Blood was collected prior to A(H1N1)pdm09 vaccination and at 1, 3 and 6 months after vaccination. Antibody titers to A(H1N1)pdm09 vaccine were measured by hemagglutination inhibition (HI) assay while the mRNA expression levels of AID were measured by quantitative real time PCR. Upon B-cell activation in vitro, AID increase correlated to antibody response to the A(H1N1)pdm09 vaccine at 1 month after vaccination in all individuals. In addition, the maximum expression levels of AID were significantly higher in those individuals who still carried protective levels of A(H1N1)pdm09 antibodies after 6 months from vaccination. No correlation was found between CD4+T-cell counts or age at vaccination or HIV-1 viral load and levels of A(H1N1)pdm09 antibodies. Assessing AID expression before vaccination may be an additional useful tool for defining a vaccination strategy in immune-compromised individuals at risk of immunization failure.

Objectives: During HIV-1 infection, the development, phenotype, and functionality of B cells are impaired. Transitional B cells and aberrant B-cell populations arise in blood, whereas a declined percentage of resting memory B cells is detected. Our study aimed at pinpointing the demographic, immunological, and viral factors driving these pathological findings, and the role of antiretroviral therapy in reverting these alterations. Design: B-cell phenotype and correlating factors were evaluated. Methods: Variations in B-cell subsets were evaluated by flow cytometry in HIV-1-infected individuals naive to therapy, elite controllers, and patients treated with antiretroviral drugs (virological control or failure). Multivariable analysis was performed to identify variables independently associated with the B-cell alterations. Results: Significant differences were observed among patients' groups in relation to all B-cell subsets. Resting memory B cells were preserved in patients naive to therapy and elite controllers, but reduced in treated patients. Individuals naive to therapy and experiencing multidrug failure, as well as elite controllers, had significantly higher levels of activated memory B cells compared to healthy controls. In the multivariate analysis, plasma viral load and nadir CD4+ T cells independently correlated with major B-cell alterations. Coinfection with hepatitis C but not hepatitis B virus also showed an impact on specific B-cell subsets. Successful protracted antiretroviral treatment led to normalization of all B-cell subsets with exception of resting memory B cells. Conclusion: Our results indicate that viremia and nadir CD4+ T cells are important prognostic markers of B-cell perturbations and provide evidence that resting memory B-cell depletion during chronic infection is not reverted upon successful antiretroviral therapy. © 2013 Wolters Kluwer Health | Lippincott Williams & Wilkins.

Objective Decreased memory B-cell maintenance during HIV-1 infection has been associated with the viraemia-induced accumulation of activated memory B cells, sensitive to Fas-mediated apoptosis. We aimed at clarifying whether other B-cell subsets might also be affected by an increased Fas expression in HIV-1-infected patients, and we studied the possible contribution of viraemia, lymphopenia or T-cell activation in Fas upregulation on B cells. We analysed whether Fas upregulation might have collaborative effects with the dysregulation of other B-cell modulatory molecules, leukocyte-associated immunoglobulin-like receptor 1 (LAIR1) and programmed cell death protein 1 (PD-1), on B-cell homeostasis. Design Fas, LAIR1 and PD-1 were analysed on B-cell subpopulations in HIV-1-infected patients who were treatment naive, nonlymphopenic; antiretroviral therapy (ART)-treated, nonlymphopenic; or ART-treated, lymphopenic or in noninfected controls. Methods Flow cytometry was used to study B-cell subsets and Milliplex for serum cytokines. Results Fas expression increased on all B-cell subpopulations of viraemic or lymphopenic individuals. The decreased ratio of resting memory B cells and their increased Fas expression were not normalized by ART. Cytokines associated with T-cell activation might influence Fas expression on the naive and transitional B cells. LAIR1 expression decreased in all HIV-1-infected patients, but only on memory B cells, whereas PD-1 increased on resting memory B cells in viraemic patients. Conclusion Fas is regulated by the concerted action of viraemia, lymphopenia and T-cell activation during HIV-1 infection, and Fas expression is altered on all peripheral B-cell subsets. Resting memory B-cell homeostasis shows the highest sensitivity to HIV-1-induced perturbations.

Antiviral T-cell immune responses appear to be crucial to control HIV replication. Infants treated before the third month of life with highly active antiretroviral treatment (HAART) did not develop a persistent HIV-specific immune response. We evaluated how delayed initiation of HAART after 3 months of age influences the development of HIV-1-specific T-cell responses during long-term follow-up in 9 HIV-1 vertically infected infants. These data suggest that a longer antigenic stimulation, due to a larger window for therapeutic intervention with HAART, is associated with the establishment of a persistent specific HIV immune response resulting in a long-term viral control of vertically infected infants.

OBJECTIVE To assess the virological, immunological and metabolic effects of switching from an efficacious first-line protease inhibitor (PI)-based HAART to a simplified triple nucleoside reverse transcriptase inhibitor (NRTI) regimen in children vertically infected with HIV. DESIGN Prospective, open-label, before-after study of 20 vertically infected children with at least 12 consecutive months of undetectable viral load under a PI-based HAART and no previous history of NRTI treatment. METHODS At study entry, HAART was shifted to a triple-NRTI combination. RESULTS The children were aged 2 to 18 years (median, 7.9) and were followed for 96 weeks. All were receiving a PI-based regimen for an average duration of 4 years before enrollment. At study entry, 12 patients (60%) switched to abacavir, 5 (25%) to lamivudine; 2 (10%) to zidovudine and 2 to didanosine (10%). All but one patient maintained plasma HIV RNA < 50 copies/ml during the entire follow-up. No immunological failure was observed at week 96. A trend of normalization (P < 0.001) of T cell receptor Vbeta families of the CD8 cell subset was detected in 19/20 (95%), with an increased HIV-specific CD8 T cell response (P < 0.01) in 17/20 (85%). Dyslipidaemia significantly improved during the follow up (P < 0.001). No new cases of lipodystrophy were detected. CONCLUSIONS Switching to triple-NRTI regimens in selected HIV-infected children with an extremely low likelihood of harbouring nucleoside-associated mutations maintains viral suppression and immunological function, improving metabolic abnormalities and the effort to take medication for up to 96 weeks.