Background: Loss of CD4⁺ T-cells is the main feature of HIV infection. Impaired thymopoiesis is believed to play a role in this loss, especially in children. We have previously reported that the CD4C/HIV Nef transgenic (Tg) mice, which express HIV Nef under the control of human CD4 regulatory elements, develop an AIDS-like disease with phenotypes similar to pediatric AIDS, a loss of CD4⁺ T-cells in peripheral lymphoid organs, and thymic atrophy. The purpose of our current study was to describe in greater detail the nature of this thymic phenotype.

Methods: In order to study thymic development in these Tg mice, we have utilized multi-color FACS analysis on thymocytes as well as peripheral lymphoid organs isolated from Tg and non-Tg control mice. To characterize selection in the thymus, we have bred the CD4C/HIV Tg mice with the AD10 TCR Tg mice. In order to determine whether stromal cells can impair T-cell development, we performed fetal liver transplantation.

Results: We observed that 6–8 week-old Tg mice have smaller thymuses with fewer total thymocytes. FACS analysis revealed a downregulation of CD4 cell surface expression, decrease percentage of CD4⁺ single positive (SP, CD4⁺CD8^-), while CD4⁺CD8⁺ (DP) thymocyte percentage was not significantly different from that of non-Tg mice. More importantly, there was a dramatic loss of TCR^Hi CD4⁺ SP cells. These data suggested impaired DP to CD4 SP transition. In order to further characterize this defect, we analyzed expression of maturational markers (TCR, CD5, CD69, HSA). We found modulated expression of these markers in distinct thymic populations. The double (CD4C/HIV x AD10 TCR) Tg mice also exhibited impairment of CD4⁺ SP cell generation. Fetal liver transplantation ruled out significant contribution of thymic epithelium to this defect. The Vb repertoire assessed by FACS in cells from peripheral lymphoid organs was similar to non-Tg controls, with an absence of Vb3⁺ CD4⁺ T-cells, suggesting that superantigen induced negative selection was intact in these mice.

Conclusions: Overall our data suggest that Nef impairs DP to CD4⁺ SP transition in the thymus. This thymic maturation defect is likely to play an important role in the lymphopenia seen in these mice and may also represent an important defect in HIV-1 infected humans.