Monocyte subsets and HIV reservoirs in patients on HAART

P. Ellery*, C. Sonza*, A. Candlein*^, G. Paukovics*, A. Solomom#, S. Lewin#, S. Crowe*#. Burnet Institute, Melbourne (Australia); #Alfred Hospital Melbourne (Australia); ^University of Minnesota Medical School (USA)

Background: A minor subset of monocytes (5-15%), characterized phenotypically as CD14loCD16hi, reportedly has higher CCR5 expression than the majority of monocytes (85-95%) which are CD14hi and express little or no CD16. We and others have shown that infectious HIV-1 can be recovered from peripheral blood monocytes of HIV-infected individuals on HAART with prolonged viral suppression. Our recent data (below) suggest that it is the CD14loCD16hi monocytes that selectively harbour HIV-1 in these individuals. In support of this, our preliminary data (below) also suggest that CD14loCD16hi monocytes are more permissive to HIV infection in vitro than the majority of monocytes. Material and Methods: CD14lo/CD16hi and CD14hi/CD16- monocytes were enriched from peripheral blood collected from HIV+ patients on HAART (n=10) using either magnetic bead technology or high speed flow cytometric sorting (MoFlo). Monocytes were also isolated from HIV-1 seronegative buffy packs by high speed flow cytometric sorting (MoFlo). Cells from each subset were pulsed for 2 hrs with  DNase-treated HIV-1Ba-L at a MOI of 0.1-1 infectious particle per cell. Excess virus was removed by washing and cells were cultured for 48 hours. Results: HIV-1 was identified by PCR (gag primers) in monocytes from 9 of 10 pts; HIV-1 was preferentially detected in CD14loCD16hi monocytes in 7 of the 9 donors. Ability to detect HIV DNA did not correlate with VL, CD4 or therapy regimen. In the in vitro-infected monocytes, significantly higher levels of HIV-1 specific DNA were detected in the CD14lo/CD16hi monocytes in 5 of 6 experiments as detected by either real-time PCR or semi-quantitative PCR. Our preliminary data also confirm that the CD14loCD16hi subset has higher expression of CCR5 than CD14hi CD16- monocytes. Conclusions: Our data suggest that a minor subset of monocytes (CD14loCD16hi) is more susceptible to HIV-1 in vitro and in vivo than the majority of monocytes. This  heightened susceptibility to HIV infection is possibly due to higher CCR5 expression. These cells are likely to contribute to HIV persistence that prevents eradication of HIV-1 by current HAART regimens.