Excess BAFF May Impact HIV-1-Specific Antibodies and May Promote Polyclonal Responses Including Those from First-Line Marginal Zone B-Cell Populations.

We have previously shown that blood levels of B-cell Activating Factor (BAFF) rise relatively to disease progression status in the context of HIV-1 infection. Excess BAFF was concomitant with hyperglobulinemia and the deregulation of blood B-cell populations, notably with increased frequencies of a population sharing characteristics of transitional immature and marginal zone (MZ) B-cells, which we defined as marginal zone precursor-like" (MZp). In HIV-uninfected individuals, MZp present a B-cell regulatory (Breg) profile and function, which are lost in classic-progressors. Moreover, RNASeq analyses of blood MZp from classic-progressors depict a hyperactive state and signs of exhaustion, as well as an interferon signature similar to that observed in autoimmune disorders such as Systemic Lupus Erythematosus (SLE) and Sjögren Syndrome (SS), in which excess BAFF and deregulated MZ populations have also been documented. Based on the above, we hypothesize that excess BAFF may preclude the generation of HIV-1-specific IgG responses and drive polyclonal responses, including those from MZ populations, endowed with polyreactivity/autoreactivity. As such, we show that the quantity of HIV-1-specific IgG varies with disease progression status. In vitro, excess BAFF promotes polyclonal IgM and IgG responses, including those from MZp. RNASeq analyses reveal that blood MZp from classic-progressors are prone to Ig production and preferentially make usage of IGHV genes associated with some HIV broadly neutralizing antibodies (bNAbs), but also with autoantibodies, and whose impact in the battle against HIV-1 has yet to be determined.

The HIV-1 gp120 CD4-bound conformation is preferentially targeted by antibody-dependent cellular cytotoxicity-mediating antibodies in sera from HIV-1-infected individuals.

UNLABELLED: Recent studies have linked antibody Fc-mediated effector functions with protection or control of human immunodeficiency type 1 (HIV-1) and simian immunodeficiency (SIV) infections. Interestingly, the presence of antibodies with potent antibody-dependent cellular cytotoxicity (ADCC) activity in the Thai RV144 vaccine trial was suggested to correlate with decreased HIV-1 acquisition risk. These antibodies recently were found to recognize HIV envelope (Env) epitopes exposed upon Env-CD4 interaction. CD4 downregulation by Nef and Vpu, as well as Vpu-mediated BST-2 antagonism, were reported to modulate exposure of those CD4-induced HIV-1 Env epitopes and were proposed to play a role in reducing the susceptibility of infected cells to ADCC mediated by this class of antibodies. Here, we report the high prevalence of antibodies recognizing CD4-induced HIV-1 Env epitopes in sera from HIV-1-infected individuals, which correlated with their ability to mediate ADCC responses against HIV-1-infected cells, exposing these Env epitopes at the cell surface. Furthermore, our results indicate that Env variable regions V1, V2, V3, and V5 do not represent a major determinant for ADCC responses mediated by sera from HIV-1-infected individuals. Altogether, these findings suggest that HIV-1 tightly controls the exposure of certain Env epitopes at the surface of infected cells in order to prevent elimination by Fc-effector functions. IMPORTANCE: Here, we identified a particular conformation of HIV-1 Env that is specifically targeted by ADCC-mediating antibodies present in sera from HIV-1-infected individuals. This observation suggests that HIV-1 developed sophisticated mechanisms to minimize the exposure of these epitopes at the surface of infected cells.

Short communication: Anti-HIV-1 envelope immunoglobulin Gs in blood and cervicovaginal samples of Beninese commercial sex workers.

Characterization of the immune correlates of protection against HIV infection is crucial for the development of preventive strategies. This study examined HIV-1 envelope (Env) glycoproteins, specifically immunoglobulin G (IgG), in systemic and mucosal compartments of female Beninese commercial sex workers (CSWs). Samples of 23 HIV-1-positive and 20 highly exposed HIV-1-seronegative (HESN) CSWs were studied. HIV-1 Env-specific IgG detection in sera and cervicovaginal lavages (CVLs) from the study population was done by cell-based ELISA. The HIV neutralizing activity was evaluated with a neutralization assay. The HIV-1-specific antibody-dependent cellular cytotoxicity (ADCC) response of the cohort was measured with a FACS-based assay evaluating the ADCC-mediated elimination of gp120-coated target cells. No anti-HIV-1 Env-specific IgG neutralizing or ADCC activities were detected in samples from HESN CSWs. Samples from HIV-1-infected CSWs presented ADCC activity in both sera and CVLs. Anti-Env IgG from sera and CVLs from HIV-1-infected CSWs preferentially recognized Env in its CD4-bound conformation. HIV-1-infected CSWs have ADCC-mediating IgG that preferentially recognizes Env in its CD4-bound conformation at the mucosal site.

Conformational evaluation of HIV-1 trimeric envelope glycoproteins using a cell-based ELISA assay.

HIV-1 envelope glycoproteins (Env) mediate viral entry into target cells and are essential to the infectious cycle. Understanding how those glycoproteins are able to fuel the fusion process through their conformational changes could lead to the design of better, more effective immunogens for vaccine strategies. Here we describe a cell-based ELISA assay that allows studying the recognition of trimeric HIV-1 Env by monoclonal antibodies. Following expression of HIV-1 trimeric Env at the surface of transfected cells, conformation specific anti-Env antibodies are incubated with the cells. A horseradish peroxidase-conjugated secondary antibody and a simple chemiluminescence reaction are then used to detect bound antibodies. This system is highly flexible and can detect Env conformational changes induced by soluble CD4 or cellular proteins. It requires minimal amount of material and no highly-specialized equipment or know-how. Thus, this technique can be established for medium to high throughput screening of antigens and antibodies, such as newly-isolated antibodies.

Flow cytometry-based assay to study HIV-1 gp120 specific antibody-dependent cellular cytotoxicity responses.

Increased attention on the role of Fc-mediated effector functions against HIV-1 has led to renewed interest into the role that antibody-dependent cellular cytotoxicity (ADCC) could play in controlling viral transmission and/or the rate of disease progression. While (51)Chromium release assays have traditionally been used to study ADCC responses against HIV-1, a number of alternative flow-cytometry-based assays were recently developed. In this study, an alternative flow-cytometry-based assay was established to allow non-radioactive measurement of ADCC-mediated elimination of HIV-1 gp120 envelope glycoprotein (Env)-coated target cells. This assay relies on staining target and effector cells with different dyes, which allows precise gating and permits the calculation of the number of surviving target cells by normalization to flow-cytometry particles. By using small concentrations of recombinant gp120 Env, suitable targets cells that recapitulate the ADCC response mediated against HIV-1-infected cells were generated. Finally, this method was applied successfully to screen human sera for ADCC activity directed against HIV-1 gp120 Env.

The cytokines cardiotrophin-like cytokine/cytokine-like factor-1 (CLC/CLF) and ciliary neurotrophic factor (CNTF) differ in their receptor specificities.

Ciliary neurotrophic factor (CNTF) and cardiotrophin-like cytokine (CLC) are two cytokines with neurotrophic and immunomodulatory activities. CNTF is a cytoplasmic factor believed to be released upon cellular damage, while CLC requires interaction with a soluble cytokine receptor, cytokine-like factor 1 (CLF), to be efficiently secreted. Both cytokines activate a receptor complex comprising the cytokine binding CNTF receptor α (CNTFRα) and two signaling chains namely, leukemia inhibitory factor receptor ß (LIFRß) and gp130. Human CNTF can recruit and activate an alternative receptor in which CNTFRα is substituted by IL-6Rα. As both CNTF and CLC have immune-regulatory activities in mice, we compared their ability to recruit mouse receptors comprising both gp130 and LIFRß signaling chains and either IL-6Rα or IL-11Rα which, unlike CNTFRα, are expressed by immune cells. Our results indicate that 1) mouse CNTF, like its human homologue, can activate cells expressing gp130/LIFRß with either CNTFRα or IL-6Rα and, 2) CLC/CLF is more restricted in its specificity in that it activates only the tripartite CNTFR. Several gp130 signaling cytokines influence T helper cell differentiation. We therefore investigated the effect of CNTF on CD4 T cell cytokine production. We observed that CNTF increased the number of IFN-γ producing CD4 T cells. As IFN-γ is considered a mediator of the therapeutic effect of IFN-ß in multiple sclerosis, induction of IFN-γ by CNTF may contribute to the beneficial immunomodulatory effect of CNTF in mouse multiple sclerosis models. Together, our results indicate that CNTF activates the same tripartite receptors in mouse and human cells and further validate rodent models for pre-clinical investigation of CNTF and CNTF derivatives. Furthermore, CNTF and CLC/CLF differ in their receptor specificities. The receptor α chain involved in the immunomodulatory effects of CLC/CLF remains to be identified.