After adding the enhanced chemiluminescent substrate the relative light units emitted were determined in a microplate luminometer
After adding the enhanced chemiluminescent substrate the relative light units emitted were determined in a microplate luminometer. in immunocompetent mice, but not in the immunodeficient mice. Thus, neither cells expressing Env after adenoviral gene transfer nor VLPs induce a T LDN-192960 cell impartial primary anti-Env antibody response. However, secondary B cell responses to Env, but not to Gag, were observed in immunodeficient mice after transfer of primed B cells and boosting with VLPs or adenoviral vectors expressing Gag and Env. This T cell impartial secondary antibody response to Env was reduced after stimulation with VLPs altered to contain monomeric LDN-192960 membrane bound gp130 surface subunit of Env and undetectable after injection of soluble gp130. Conclusions Membrane-bound trimeric Env seems to be responsible for the maintenance of high levels of anti-Env antibodies during progression to AIDS. This T cell impartial secondary antibody response may prevent T cell-dependent affinity maturation and thus contribute to viral immune escape by favoring persistence of non-protective antibodies. Keywords: SIV, HIV, Adenoviral vectors, T-independent antibody response, VLP Background Although HIV contamination induces a vigorous antibody response to Gag and Env proteins, LDN-192960 the induced antibodies do not prevent progression to AIDS. The induction of neutralizing antibodies seems to be too slow and inefficient to keep pace with the rapidly mutating HIV [1,2]. Whether other antibody-mediated antiviral effector mechanisms, such as complement activation and antibody-dependent cytotoxicity, slow down the progression of the disease is unknown [3-5]. The major LDN-192960 targets for antibody-mediated inhibition of HIV are the gp120 surface protein (SU), the gp41 transmembrane protein (TM), and possibly the gp160 Env precursor protein. However, the magnitude of the antibody response to Env does not correlate with the slower progression of disease [6-8]. While high levels of Env antibodies are maintained throughout contamination, the decline of Gag antibodies with progressing HIV contamination is an indicator for a poor prognosis [6-11]. This correlation is unlikely to reflect a direct antiviral activity of Gag antibodies, since Gag proteins are either located inside an infected cell or inside the computer virus particle with a lipid membrane blocking the accessibility of Gag proteins by antibodies. Macaques chronically infected with simian immunodeficiency computer virus also maintained high levels of anti-Env antibodies, whereas anti-Gag antibodies declined with progression to AIDS [12,13]. It was, therefore, suggested that this decline of Gag antibodies with progression to AIDS is due to the loss LW-1 antibody of CD4+ T cell help, while a T cell impartial antibody response to Env allows persistence of Env antibodies [6]. A differential regulation of Gag and Env antibody responses was also observed during natural non-progressive contamination of African green monkeys. While anti-Gag antibody responses were weak or not observed at all, the anti-Env antibody responses were as strong as observed in HIV contamination [14]. Since the limitation of immune activation has been proposed to be a key determinant of non-pathogenic immunodeficiency computer virus infections [15], the paucity of the Gag-specific antibodies might be due to a limited T helper cell activation. A T cell-independent Env antibody response might then explain the high levels of Env antibodies observed. The molecular mechanisms LDN-192960 mediating the differential requirement of Gag and Env antibodies for T cell help have not been unraveled. Presentation of trimeric Env in a repetitive manner on the surface of computer virus particles or infected cells might allow cross-linking of Env-specific B cell receptors providing the first signal during B cell activation. However, a T cell impartial antiviral antibody response seems to depend on the precise arrangement of the viral surface protein around the viral particle. Contamination with vesicular stomatitis computer virus, which forms viral particles with densely packed G protein spikes, induces T cell impartial antibody responses. In contrast, antibody responses to contamination with lymphocytic choriomeningitis computer virus, the virions of which contain less densely packed spikes, are T cell dependent [16]. In addition to the particulate nature of the HIV virion, gp120 was found to have direct B cell stimulatory activity [17]. Thus, gp120 SU might provide both cross-linking of surface BCR (signal 1) and an innate stimulus (signal 2). Theoretically, these two signals alone could be sufficient to trigger differentiation of na?ve Env-specific B cells into Ig secreting plasma cells [18]. A T cell impartial Env antibody response might also play an important role in viral immune escape: the affinity and avidity of T cell impartial Env antibodies might be too low for efficient neutralization. Thus, a better understanding of the differential regulation of Gag and Env-specific B cell responses might provide further insights in the pathogenesis of HIV and guideline novel strategies in HIV vaccine development. In the present report, we therefore provide.