Distinct patterns of Bcl-2 expression occur in R5- and X4-tropic HIV-1-producing lymphoid tissue cells infected ex vivo

Most HIV-1 replication occurs in secondary lymphoid tissues in T cells within B cell follicles. Mechanisms underlying the accumulation of HIV-1-producing cells at these sites are not understood. Antiapoptotic proteins such as Bcl-2 could promote follicular CD4⁺ T cell survival, contributing to sustained virus production. Tonsils obtained from subjects without known HIV infection were disaggregated and analyzed for Bcl-2 expression in follicular (CXCR5⁺) and extrafollicular (CXCR5^-) CD3⁺CD4⁺ cells by flow cytometry. Additional tonsil cells were cultured with phytohemagglutinin (PHA) and interleukin-2 (IL-2) for 2 days, infected with either CCR5(R5) or CXCR4-tropic (X4) GFP reporter viruses, and analyzed for Bcl-2 expression. In freshly disaggregated CD3⁺CD4⁺ tonsil cells, mean florescence intensity (MFI) for Bcl-2 was higher in CXCR5⁺ (median, 292) compared to CXCR5^- cells (median, 194; p=0.001). Following in vitro stimulation with PHA and IL-2, Bcl-2 MFI was higher in both CXCR5⁺ cells (median, 757; p=0.03) and CXCR5^- cells (median, 884; p=0.002) in uninfected cultures compared to freshly isolated tonsil cells. Bcl-2 MFI was higher in GFP⁺CD3⁺CD8^- R5-producing cells (median, 554) than in X4-producing cells (median, 393; p=0.02). Bcl-2 MFI was higher in R5-producing CXCR5⁺ cells (median, 840) compared to all other subsets including R5-producing CXCR5^- cells (median, 524; p=0.04), X4-producing CXCR5⁺ cells (median, 401; p=0.02), and X4-producing CXCR5^- cells (median, 332; p=0.008). Bcl-2 expression is elevated in R5 HIV-1-producing CXCR5⁺ T cells in vitro, which may contribute to propagation of R5 virus in B cell follicles in vivo.