Subpopulations of CD4-CD8-murine thymocytes: differences in proliferation rate in vivo and proliferative responses in vitro

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Abstract
Cell sorting and cytotoxic depletion procedures were used to subdivide the population of CD4‐ CD8‐(“double‐negative”) thymocytes from adult CBA mice on the basis of expression of Ly‐1, HSA (the “heat‐stable antigen” M1/69 or B2A2), Pgp‐1 glycoprotein, Thy‐1, MEL‐14 and the PC61 antigenic determinant on the IL2 receptor (IL2R). The level of dividing cells within these subsets was assessed by brief in vivo administration of [3H]t‐hymidine, followed by radioautography, or by flow cytometric cell cycle analysis after DNA staining. The capacity of the subsets to proliferate in culture, in response to stimulation with concanavalin A (Con A), or with phorbol myristate acetate (PMA) and the calcium ionophore ionomycin, was assessed in high cloning efficiency single‐cell culture systems. In general, the proliferative response in culture was inversely related to the rate of cell division in vivo. Response of the double‐negative subsets to Con A correlated with expression of the T cell antigen receptor complex; although a high cloning efficiency was obtained from the receptorpositive fractions, very few of the clones were cytotoxic. In particular, a major Ly‐1+' HSA‐ Pgp‐1+ double‐negative subset, as well as minor Ly‐1‐ HSA‐ Pgp‐1+ subsets, contained very few cells in cycle in vivo, but showed a high cloning efficiency in both culture systems. Conversely, the other major double‐negative subset, Ly‐1‐ HSA+ Pgp‐1‐, included most of the cells in cycle, but showed a reduced cloning efficiency in response to PMA and ionomycin and failed to respond to Con A. The dividing cells within the Ly‐1‐ HAS+' Pgp‐1‐ group were strongly enriched in the IL2R‐ rather than in the IL2R+ subset, suggesting IL2 was not the growth factor maintaining their proliferation in vivo.

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