210 Peripheral blood mononuclear cells activated by HIV-VLPs in correlation with HIV-1 seropositivity status

Abstract

We have recently shown that HIV-1 Pr55gag Virus-Like Particles (HIVVLPs), produced in a baculovirus expression system and presenting a gp120 molecule from an Ugandan HIV-1 isolate of the clade A (HIV-VLPAs), induce maturation and activation of monocyte-derived dendritic cells (MDDCs) with a production of Th1- and Th2-specific cytokines. Furthermore, HIV-VLP-loaded MDDCs are able to induce a primary and secondary response in autologous human CD4+ T cells, in an ex vivo immunization assay. Fresh human peripheral blood mononuclear cells were isolated by Ficoll-Hypaque density gradient centrifugation and plated in 6-well plates at a concentration of 1 × 107/well. PBMCs were pulsed with 6_g/mL of HIV-VLPs. After 16 hrs, the cells were harvested, washed, and stained for phenotypic analysis by flow cytometry. The cellular supernatants were collected for quantification of cytokine production by ELISA. In the present study we show that similar data can be obtained directly on fresh peripheral blood mononuclear cells (PBMCs) and the HIV-1 seropositivity status, with either low or high viremia, does not significantly impair the immune activation status and the responsiveness of circulating monocyte CD14+ cell populations to an immunogenic stimulus. The established Th2 polarization in both HIV seropositive groups is efficiently boosted by HIV-VLP induction and does not switch into a Th1 pattern, strongly suggesting that specific Th1 adjuvants would be required for a therapeutic effectiveness in HIV-1 infected subjects. These results indicate the possibility of screening PBMCs for donor susceptibility to an immunogen treatment, which would greatly simplify the identification of “responsive” vaccinees as well as the understanding of eventual failures in individuals enrolled in clinical trials.