(F) Neutralization activity (AUC) with VCs split into LTNP and EC subgroups. FCGR activation may provide a useful method for screening and distinguishing protective anti-HIV IgG responses in HIV-infected patients and in DBCO-NHS ester 2 monitoring HIV vaccination regimens. Introduction HIV-1 viremic controller (VC) patients represent a unique population of individuals who naturally suppress HIV replication in the absence of highly active antiretroviral therapy (HAART) (1, 2). These VC patients can be divided into 2 subgroups made up of long-term nonprogressors (LTNPs) and elite controllers (ECs). Both LTNPs and ECs maintain normal DBCO-NHS ester 2 CD4 levels, but ECs inhibit computer virus replication below detectable levels (50 copies/ml), while LTNPs maintain potent suppression below 1,000 viral copies/ml of blood. Several studies have reported strong Ab-dependent cellular cytotoxicity (ADCC) and Ab-dependent cell-mediated viral inhibition (ADCVI) responses in both subgroups of VCs (3C7). These reports suggest that, in addition to associations with favorable HLA, which can promote strong cytotoxic T cell responses (8, 9), Ab-mediated effector cell responses (Ab-MERs) may also contribute to the HIV controller phenotype. In the RV144 HIV vaccine trial, a small reduction in HIV acquisition correlated with the induction of non-neutralizing IgG Ab responses that mediate ADCC after controlling for IgA responses (10, 11). Given this common feature, further characterization of IgG responses from VCs may yield valuable insights into the mechanisms that underlie potent antiviral Fc effector responses and identify Ab profiles that may be desired to emulate in HIV vaccine strategies. Most Ab-MERs are initiated by engagement of Fc- receptor (FCGR) activation on the surface of innate immune effector cells (e.g., NK cells, macrophages, monocytes, and DCs) (12, 13). FCGRs are a diverse family of receptors that orchestrate an array of cellular effector functions, such as ADCC, Ab-dependent cellular phagocytosis (ADCP), and effector cytokine production. Two activating FCGRs that are particularly important to the induction of ADCC, ADCP, and cytokine production are FCGR2A and FCGR3A. These low-affinity FCGRs are activated by receptor cross-linking induced by binding of IgG-antigen immune complexes. Recently, Ackerman et al. exhibited that HIV controller IgG displayed an enhanced breadth of ADCC, ADCP, and NK activation and cytokine production; however, no single Ab-MER was observed to be significantly higher as compared with noncontrollers (14). This result contrasts with other studies that have observed enhanced ADCC/ADCVI activity in controller patients (3C7). Given that FCGR signaling modulates the efficiency of Fc-induced effector responses, assessing the IgG Fc activation profiles associated with controller status may yield insights as to why some VCs display an enhanced breadth of Fc effector responses (14). IgG Fc effector responses against HIV-1 are dependent on the DBCO-NHS ester 2 acknowledgement of the HIV-1 envelope (Env) glycoproteins (gp), since gp120 and gp41 are the only viral antigens uncovered on the surface of virions and HIV-infected cells. The form (e.g., computer virus, cell-surface, or free protein) in which these antigens are offered to antibodies can also greatly influence their ability to activate Fc-mediated responses. Many studies examining Fc effector responses utilized peptides, monomeric soluble recombinant gp120, or recombinant gp140 trimers (rgp120; rgp140) (3, 6, 14C25), but these antigens may not reflect the epitopes that are uncovered around the conformationally flexible and highly glycosylated native structure of the HIV-1 Env trimer expressed on the surface of HIV-infected T cells (26C29). Previous studies demonstrate that this antiviral factor tetherin enhances the presentation of viral antigens to the innate immune system through Ab-dependent mechanisms (30C33). Building on these observations, we have developed a tetherin-expressing, cell-based system to measure HIV-specific Ab binding, FCGR signaling, and Ab-MERs using a single platform expressing functional infectious antigens. This system characterizes Abs that identify epitopes that are uncovered on cell-associated and computer virus incorporated (CANVI) forms of HIV Env. Herein, we utilize this system in conjuction with neutralization assays to examine the levels of HIV-specific binding, FCGR activation, and neutralization mediated by IgG from VCs versus HIV+ chronically infected patients (CIPs) on Rabbit Polyclonal to ABCF1 HAART. Results A cell-based Ab binding assay to characterize HIV-specific Abs. Expression of the anti-viral factor tetherin on the surface of HIV-infected T lymphocytes allows for the enhanced detection of FCGR signaling and Fc-mediated effector functions (30). To exploit the potential of tetherin to increase the detection of cell-surface viral antigens, we developed an HIV-infected, cell-based system that allows for the characterization of.
