AEGiS-13IAC: Kinetics of HIV reactivation from the state of latency. Efficacy of different immune stimuli in triggering HIV replication.

13th International AIDS Conference

Durban, South Africa - July 9-July 14, 2000

Kinetics of HIV reactivation from the state of latency. Efficacy of different immune stimuli in triggering HIV replication.

Int Conf AIDS 2000 Jul 9-14; 13:(abstract no. MoOrA172)

Alcami J, Pedraza M, Bermejo M, Rullas J
J. Alcami, Unidad de Inmunopatologia del SIDA, Instituto de Salud Carlos III, Spain

Background and objectives: We have developed an efficient transfection system of resting peripheral blood lymphocytes (PBL) that allow us to reactivate a full-length transfected provirus using different stimuli. In this system we have addressed the following questions: 1. The kinetics of HIV replication from the state of proviral latency in T lymphocytes. 2. The impact of different pharmacokinetic parameters on the efficacy of PI. 3. The efficacy of different immune stimuli to trigger HIV replication.

METHODS: PBL were transfected with a whole provirus (NL4.3). In some experiments a NL4.3 construct containing a luciferase reporter gene was used. After transfection cells were activated with PMA (25 ng/ml), PHA, Interleukin-2 (80 ng/ml) or anti-CD3 antibodies. Ritonavir was added at different times after transfection and cell activation at doses ranging from 1 m M to 0.12 m M. Viral transcription and production of viral particles were assessed by quantifying luciferase activity and levels of p24 HIV antigen in culture supernatants.

RESULTS: Full transcription of HIV proteins from proviral latency was achieved in two hours and viral particles were detected in culture supernatants from 6 to 8 hours after T cell activation. Ritonavir was useful blocking viral replication when added before four hours following T cell transfection and activation but when added at latest times viral escape occurs. In contrast with high viral replication induced by PHA or CD3, IL2 stimulation resulted in bare replication levels in culture in both HIV-transfected cells and lymphocytes from infected patients. Lack of HIV replication was not due to weak proliferation of IL-2 activated lymphocytes as demonstrated by thymidine uptake and cell progression throughout G2 and M phases of the cell cycle. In agreement with these results NF-kB, a critical factor in HIV reactivation, was not induced by IL2 treatment. An increase in HIV DNA levels was observed in lymphocytes of HIV patients cultured in the presence of IL2 as compared to unstimulated cells.

Keywords: AEGIS, HIV, Virus Replication, HIV Infections, HIV Seropositivity, Interleukin-2, HIV Long Terminal Repeat, Proviruses, Kinetics, Antigens, CD3, T-Lymphocytes, Phytohemagglutinins, HIV Core Protein p24, Transfection, Ritonavir, Transcription, Genetic, Human, pharmacokinetics, virology, genetics, immunology
000709
MoOrA172