AbstractDendritic cells (DCs) are specialized antigen-presenting cells. However, DCs exposed to human immunodeficiency virus (HIV) are also able to transmit a vigorous cytopathic infection to CD4+ T lymphocytes, a process that has been frequently related to the ability of DC-SIGN (Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin) to bind HIV-1 envelope glycoproteins. Maturation of DCs can increase the efficiency of HIV transmission through trans-infection. We aimed to comparatively study the effect of maturation in monocyte derived dendritic cells (MDDCs) and blood-derived myeloid DCs during HIV capture process. In vitro capture and transmission of envelope pseudotyped HIV-1 and its homologous replication competent virus to susceptible target cells was assessed by p24gag detection, luciferase activity, and both confocal and electron microscopy. Maturation of MDDCs or myeloid DCs enhanced active capture of HIV in a DC-SIGN and viral envelope glycoprotein independent manner, increasing the lifespan of trapped virus. Moreover, higher viral transmission of mature DCs (mDCs) to CD4+ T lymphocytes was highly dependent on active viral capture, a process mediated through cholesterol-enriched domains. Mature DCs concentrated captured virus in a single large vesicle staining for CD81 and CD63 tetraspanins, while immature DCs lacked these structures, suggesting different intracellular trafficking processes.
Exosomes are secreted cellular vesicles that can be internalized by DCs contributing to antigen specific naive CD4+ T lymphocyte activation. Here, we demonstrate that HIV can exploit this exosome antigen-dissemination pathway intrinsic to mDCs for mediating trans-infection of CD4+ T lymphocytes. Capture of HIV-1, HIV-1 Gag-eGFP viral like particles (VLPs) and exosomes by DCs was upregulated upon maturation, resulting in localization within a CD81+ compartment. Uptake of VLPs or exosomes could be inhibited by a challenge with either particle, suggesting that the expression of common determinant(s) on VLP or exosome surface is necessary for internalization by mDCs. Capture by mDCs was insensitive to proteolysis, but blocked when virus, VLPs, or exosomes were produced from cells treated with sphingolipid biosynthesis inhibitors that modulate the lipid composition of the budding particles. Finally, VLPs and exosomes captured by mDCs were transmitted to CD4+ T lymphocytes in an envelope glycoprotein-independent manner, underscoring a new potential viral dissemination pathway.
Overall, these observations help explaining the greater ability of mDCs transferring HIV to CD4+ T lymphocytes, a process that can potentially contribute to the viral dissemination at lymph nodes in vivo, where viral replication takes place and there is a continuous interaction between susceptible T-cells and mDCs.
|Date of Award||17 Apr 2009|
|Supervisor||Paz Martinez Ramirez (Tutor), Julian Miguel Blanco Arbues (Director) & Javier Martínez Picado (Director)|