Modeling latent infection of Human Immunodeficiency Virus in vitro.

摘要

Quiescent T lymphocytes containing latent Human Immunodeficiency Virus (HIV) provide a long-lived viral reservoir. This reservoir may be the source of active infection that is re-initiated following the cessation of anti-retroviral therapy. Therefore it is important to understand the mechanisms involved in latent infection to develop new strategies to eliminate the latent HIV reservoir. It has been shown that latently infected quiescent lymphocytes can be generated during thymopoiesis in vivo, in the SCID-hu mouse system. However, there is still a pressing need for an in vitro model of HIV latency in primary human cells. This dissertation presents a novel in vitro model that recapitulates key aspects of dormant HIV infection. Human CD4+CD8+ thymocytes generate a stable infection with an Enhanced Green Fluorescent Protein (EGFP) - Luciferase fusion protein containing reporter virus in the absence of viral gene expression. T cell activation induces a >200 fold induction of reporter activity. The induced reporter activity originates from a fully reverse transcribed and integrated genome. It is demonstrated that this model can be useful to study LTR regulation, as previously characterized NF-kappaB response element mutations decrease activation of viral gene expression. The model has also been adapted to micro-well format to allow high-throughput screening for activators or inhibitors of latent HIV. This model can therefore be used to study intricate molecular aspects of latent HIV infection in primary cells.