Background: These studies were designed to examine the phenotype of circulating S phase lymphocytes in HIV-infected individuals.

Methods: Whole blood from HIV-infected individuals and healthy donors were incubated with bromodeoxyuridine (BrdU) for 1 h ex vivo and subsequently examined by flow cytometry for the expression of CD4, CD8, CD62L, Ki67, and CD45RO. Similar markers were examined on Ki67⁺ cells. The relationship between the expression of Ki67 and BrdU was examined by linear regression and Spearman Rank tests. To examine the potential of BrdU labeled cells to complete mitosis, the BrdU-labeled cells were isolated from whole blood, labeled with tracking dye (PKH26; Sigma), re-suspended in RPMI 1640 medium containing 10% serum and placed into cell culture.

Results: BrdU incorporation and Ki67 expression was well correlated among CD8⁺ cells (R = 0.85) but not CD4⁺ cells (R = 0.27). The cells that spontaneously incorporated BrdU were almost entirely CD45RO⁺CD62L⁺ cells, which is consistent with a “central memory” phenotype. In contrast, cells that expressed Ki67 were less consistently associated with this phenotype and this was especially true for Ki67⁺CD8⁺ cells, which were mostly CD62L negative. Surprisingly, BrdU-labeled cells failed to undergo cell division following isolation, and tended instead to undergo cell death.

Conclusions: 1) The results suggest that spontaneous S phase lymphocytes in HIV-infected patients have previously encountered antigen and are capable of migrating into lymphoid tissues (central memory cells). 2) BrdU incorporation may not reliably identify cells that will complete cell division, and therefore, this population is not likely to contribute to the re-population of peripheral T-cell pools in HIV-infected patients.