342 Perforin-Expressing CD4+ T-cells are Found in Oligoclonal Expansions and Decline in the Setting of HAART L. Goodwin*1, S. Bennett1, E. Connick1, B. Kotzin1, ACTG 315/375 Study Team2 1Univ of Colorado Hlth Sci Ctr, Denver and 2Adult AIDS Clin Trials Group, Washington, DC
Background: Perforin-expressing CD4+ T-cells have been reported quite recently in HIV infection. We previously described two HIV-infected individuals with expansions in CD4+ cells that express T-cell receptor variable-beta chains (VB) 2.1 and 6.7 that contained multiple clones, which persisted over 3 years of HAART and were associated with high frequencies of a CD28- phenotype. Here, we investigate the hypothesis that expanded CD4+ VB2.1 and VB6.7 subsets express perforin and evaluate the impact of HAART on perforin-expressing CD4+ cells.
Methods: Three (3) subjects with baseline CD4+ T-cell counts between 100 and 300 cells/mm3 were evaluated. Subjects P490 and P292 were known to harbor clonal expansions in CD4+ VB2.1 and VB6.7 subsets, whereas subject P173 had relatively diverse, unexpanded CD4+ VB2.1 and VB6.7 subsets. Cryopreserved PBMC were stained for CD4, perforin, CD28, VB2.1, and VB6.7 and analyzed by flow cytometry.
Results: The proportion of CD4+ cells that expressed perforin was higher in subjects P490 (71%) and P292 (40%) than in subject P173 (29%). P490 had 69% and 84%, P292 had 53% and 31%, and P173 had 14% and 15% perforin-expressing CD4+ cells in the VB2.1 and VB6.7 subsets, respectively. At baseline, 89%, 94%, and 92% of perforin-expressing CD4+ cells were CD28- in subjects P490, P292, and P173, respectively. After 3 yrs of HAART during which all subjects achieved significant CD4+ T-cell increases and maintained plasma HIV RNA concentration < 100 copies/mL, the proportion of CD4+ T-cells that expressed perforin had declined to 61%, 5%, and 6%, respectively. The percentage of CD28- cells that expressed perforin decreased variably in each subject over 3 years of HAART from 88% to 72% in P490, 50% to 21% in P292, and 84% to 17% in P173.
Conclusions: High percentages of perforin-expressing CD28- CD4+ T-cells were found in whole PBMC of 3 pts. The highest percentages were found in 2 pts known to harbor large oligoclonal expansions, and high percentages were found in those expanded subsets as well. These data suggest that a significant fraction of oligoclonal CD4+ cell expansions express perforin. Perforin-expressing cells declined on HAART in all 3 subjects to some extent, but a high proportion of perforin-expressing cells persisted in one individual even after 3 yrs. These cells likely represent CD4+ T-cells that have differentiated to express cytolytic function and may contribute to the immunopathology of HIV.
