Endocytosis of HIV-1 activates plasmacytoid dendritic cells via Toll-like receptor-viral RNA interactions.

HIV-1 directly activates human plasmacytoid DCs (pDCs) by upregulating the expression of costimulatory and MHC molecules and maturation markers, increasing T cell stimulatory activity, and inducing the production of type I interferons and TNF-alpha. A consequence of this activation is the bystander maturation of myeloid DCs and overall enhancement of antigen-presenting function. However, little is known about the mechanism(s) of pDC activation by HIV-1. Here we demonstrate by in vitro studies that IFN-alpha production by pDC in response to HIV-1 requires at least 2 interactions between the cell and virus. Initially, envelope-CD4 interactions mediate endocytosis of HIV-1, as demonstrated through the use of inhibitors of binding, fusion, endocytosis, and endosomal acidification. Subsequently, endosomally delivered viral nucleic acids, particularly RNA, stimulate pDCs through TLRs, as activation is reproduced with purified genomic RNA but not viral RNA packaging-deficient HIV-1 and blocked with different inhibitory TLR ligands. Finally, by using genetic complementation, we show that TLR7 is the likely primary target. Viral RNA rather than DNA in early retrotranscripts appears to be the active factor in HIV-1 that induces IFN-alpha secretion by pDCs. Since the decline in pDCs in chronic HIV-1 infection is associated with high viral loads and opportunistic infections, exploiting this natural adjuvant activity of HIV-1 RNA might be useful in the development of vaccines for the prevention of AIDS.

Broad vaccine coverage predicted for a bivalent recombinant factor H binding protein based vaccine to prevent serogroup B meningococcal disease.

Factor H binding proteins (fHBP), are bacterial surface proteins currently undergoing human clinical trials as candidate serogroup B Neisseria meningitidis (MnB) vaccines. fHBP protein sequences segregate into two distinct subfamilies, designated A and B. Here, we report the specificity and vaccine potential of mono- or bivalent fHBP-containing vaccines. A bivalent fHBP vaccine composed of a member of each subfamily elicited substantially broader bactericidal activity against MnB strains expressing heterologous fHBP than did either of the monovalent vaccines. Bivalent rabbit immune sera tested in serum bactericidal antibody assays (SBAs) against a diverse panel of MnB clinical isolates killed 87 of the 100 isolates. Bivalent human immune sera killed 36 of 45 MnB isolates tested in SBAs. Factors such as fHBP protein variant, PorA subtype, or MLST were not predictive of whether the MnB strain could be killed by rabbit or human immune sera. Instead, the best predictor for killing in the SBA was the level of in vitro surface expression of fHBP. The bivalent fHBP vaccine candidate induced immune sera that killed MnB isolates representing the major MLST complexes, prevalent PorA subtypes, and fHBP variants that span the breadth of the fHBP phylogenetic tree. Importantly, epidemiologically prevalent fHBP variants from both subfamilies were killed.

Detection of LP2086 on the cell surface of Neisseria meningitidis and its accessibility in the presence of serogroup B capsular polysaccharide.

The outer membrane protein LP2086, a human factor H binding protein, is undergoing clinical trials as a vaccine against invasive serogroup B meningococcal (MnB) disease. As LP2086 is a surface protein, expression of capsular polysaccharide could potentially limit accessibility of anti-LP2086 antibodies to LP2086 expressed on the surface of bacteria. To determine whether variability in expression levels of the serogroup B capsule (Cap B) might interfere with accessibility of anti-LP2086 antibody binding to LP2086, we evaluated the ability of anti-Cap B and anti-LP2086 antibodies to bind to the surface of 1263 invasive clinical MnB strains by flow cytometry. One of the anti-LP2086 monoclonal antibodies used recognizes virtually all LP2086 sequence variants. Our results show no correlation between the amount of Cap B expressed and the binding of anti-LP2086 antibodies. Furthermore, the susceptibility of MnB bacteria to lysis by anti-LP2086 immune sera was independent of the level of Cap B expressed. The data presented in this paper demonstrates that Cap B does not interfere with the binding of antibodies to LP2086 expressed on the outer membrane of MnB clinical isolates.

Activation of influenza virus-specific CD4+ and CD8+ T cells: a new role for plasmacytoid dendritic cells in adaptive immunity.

Plasmacytoid dendritic cells (pDCs) contribute to innate antiviral immune responses by producing type I interferons (IFNs) upon exposure to enveloped viruses. However, their role in adaptive immune responses, such as the initiation of antiviral T-cell responses, is not known. In this study, we examined interactions between blood pDCs and influenza virus with special attention to the capacity of pDCs to activate influenza-specific T cells. pDCs were compared with CD11c(+) DCs, the most potent antigen-presenting cells (APCs), for their capacity to activate T-cell responses. We found that like CD11c(+) DCs, pDCs mature following exposure to influenza virus, express CCR7, and produce proinflammatory chemokines, but differ in that they produce type I IFN and are resistant to the cytopathic effect of the infection. After influenza virus exposure, both DC types exhibited an equivalent efficiency to expand anti-influenza virus cytotoxic T lymphocytes (CTLs) and T helper 1 (TH1) CD4(+) T cells. Our results pinpoint a new role of pDCs in the induction of antiviral T-cell responses and suggest that these DCs play a prominent role in the adaptive immune response against viruses.

Human immunodeficiency virus type 1 activates plasmacytoid dendritic cells and concomitantly induces the bystander maturation of myeloid dendritic cells.

In this study, we analyzed the phenotypic and physiological consequences of the interaction of plasmacytoid dendritic cells (pDCs) with human immunodeficiency virus type 1 (HIV-1). pDCs are one cellular target of HIV-1 and respond to the virus by producing alpha/beta interferon (IFN-alpha/beta) and chemokines. The outcome of this interaction, notably on the function of bystander myeloid DC (CD11c+ DCs), remains unclear. We therefore evaluated the effects of HIV-1 exposure on these two DC subsets under various conditions. Blood-purified pDCs and CD11c+ DCs were exposed in vitro to HIV-1, after which maturation markers, cytokine production, migratory capacity, and CD4 T-cell stimulatory capacity were analyzed. pDCs exposed to different strains of infectious or even chemically inactivated, nonreplicating HIV-1 strongly upregulated the expression of maturation markers, such as CD83 and functional CCR7, analogous to exposure to R-848, a synthetic agonist of toll-like receptor-7 and -8. In addition, HIV-1-activated pDCs produced cytokines (IFN-alpha and tumor necrosis factor alpha), migrated in response to CCL19 and, in coculture, matured CD11c+ DCs, which are not directly activated by HIV. pDCs also acquired the ability to stimulate naïve CD4+ T cells, albeit less efficiently than CD11c+ DCs. This HIV-1-induced maturation of both DC subsets may explain their disappearance from the blood of patients with high viral loads and may have important consequences on HIV-1 cellular transmission and HIV-1-specific T-cell responses.