Visualizing memory phenotype development after in vitro stimulation of CD4(+) T cells

Stimulation of naive CD4 cells by specific antigen results in proliferation and changes in cell surface marker expression as the cells differentiate into effector and memory cells. Several of the marker changes (e.g., differences in CD45RB, CD44, and L-selectin levels) appear to be relatively stable and permit the identification of memory T cells. In this study, we examined the acquisition of memory markers after the initial stimulation of naive T cells. CD4(+) T cells from DO11.10 TCR transgenic mice were labeled with the fluorescent dye carboxyfluorescein diacetate succinimidyl ester (CFSE) and were stimulated with specific antigen (OVA323-339). Specific activation was observed, as CFSE-associated fluorescence was reduced twofold with each division of DO11.10 clonotype-bearing cells. Phenotypic changes could also be observed as the cells differentiated into effector/memory cells. However, individual surface markers exhibited a varied relationship to cell division. Although changes in some markers (L-selectin) occurred independently of cell division, changes in other markers were either strictly related to cell division (CD45RB) or were a prerequisite to cell division (CD4, CD44).