Comparative analysis of total and integrated HIV-1 DNA in peripheral CD4 lymphocytes and monocytes after long treatment with HAART.
OBJECTIVE
To determine the level of total and integrated HIV-1 DNA load in CD4 lymphocytes and monocytes of patients undergoing HAART treatment for at least 2 years.


METHODS
CD4 lymphocytes were isolated by subjecting monocyte-depleted blood samples to immune-purging carried out with M-450 Dynabeads. Monocytes were separated by blood through a combined procedure of cell adherence to dishes and complement induced immune lysis with anti-CD3 Mab. HIV DNA in CD4 lymphocytes and monocytes was quantified by polymerase chain reaction (PCR) based limit dilution assay with two PCR protocols, specific for total (LTR PCR) and integrated (Alu PCR) forms of HIV DNA. The replication competence of the provirus harboured in monocyte-depleted peripheral blood mononuclear cells (PBMC) and adherent monocytes was assayed by measuring HIV-1 p24 antigen produced by in-vitro cultures established with these cells.


RESULTS
The CD4 lymphocytes of all patients contained a consistent number of HIV DNA copies. Most patients were also positive for HIV DNA in monocytes. The Alu PCR analysis detected, integrated provirus in CD4 lymphocytes of 9 patients and in the monocytes of only three. Four patients had replication-competence virus in their PBL. The monocytes of all patients did not produce virus in vitro.


CONCLUSIONS
The HIV infection of CD4 lymphocytes and monocytes is maintained even after HAART related, apparent, and durable suppression of viral replication. We suggest that the viral persistent infection of monocytes may play a role in maintaining the residual HIV activity found in patients undergoing HAART.
Calcaterra
S
S
Laboratory of Virology, L. Spallanzani Hospital IRCCS, Rome, Italy.
Cappiello
G
G
Di Caro
A
A
Garbuglia
A R
AR
Benedetto
A
A
D023241:Antiretroviral Therapy, Highly Active; D015496:CD4-Positive T-Lymphocytes / Q000821:virology*; D004279:DNA, Viral / Q000032:analysis*; D015658:HIV Infections / Q000097:blood* / Q000188:drug therapy / Q000821:virology*; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D009000:Monocytes / Q000821:virology*; D016133:Polymerase Chain Reaction; D012720:Severity of Illness Index; D013997:Time Factors; D016896:Treatment Outcome
D004279:DNA, Viral