Two approaches have been used to address this issue. (1) Healthy and HIV-infected patients receive up to 5 days of continuous infusions with 6,6-2H2-glucose, a nonradioactive, stable isotope of glucose that is safe to administer. The deuterium is incorporated into DNA via metabolism of glucose to ribose and incorporation into nucleotides. The rate of incorporation can be measured in subpopulations of cells to determine the rate of replication of those cells, and the rate of loss of the incorporated deuterium can be used to examine the turnover rate of the replicated cells. (2) Administering bromodeoxyuridine (BrDU; 200 mg/m2), an analogue of thymidine, to HIV-infected patients. BrDU is incorporated into DNA and incorporation can be measured using an anti-BrDU monoclonal antibody. By FACS analysis, both surface markers and BrDU can be measured. Thus, FACS analysis can be used to directly measure subpopulations of cells that have replicated. To date, approximately 110 patients have been enrolled in these studies. Techniques for measuring incorporation have been developed and validated for both methods. Studies with BrDU have identified two populations of proliferating cells, one with a rapid turnover, and the second with a slow turnover. The size of the rapidly proliferating pool, but not the slowly proliferating pool, is directly related to the log viral load, suggesting that HIV drives cells to enter the rapidly proliferating pool. Activated central memory cells are the primary population of cells that are proliferating for both CD4 and CD8 cells. We have also examined lymphocyte kinetics in patients receiving IL-2, and have found that intermittent IL-2 therapy expands both CD4 and CD8 cells; CD4 cells then have a very long survival in responding patients. Naive and central memory CD4 cells appear to be preferentially expanded. The increased CD4 cell survival is due to prolongation of survival of individual subpopulation of cells as well as a relative increase in longer-lived naive cells. There is a direct correlation between average survival of CD4 cells and the CD4 cell increase seen following IL-2 therapy. Of note, the BrdU studies are no longer enrolling patients based on a DSMB recommendations, but analysis of the data that have been obtained are ongoing. These two approaches should provide information about lymphocyte kinetics that will have relevance to better understanding HIV infection and other disease states.
