Characterization of the requirements for human T cell mitogenesis by using suboptimal concentrations of phytohemagglutinin

To characterize the requirements for T cell proliferation, we have studied the response of purified populations of human T cells to varying concentrations of the mitogen phytohemagglutinin (PHA). Concentrations of PHA which induce optimal proliferative responses induce increases in cytosolic free calcium ([Ca²⁺](i)), expression of interleukin 2 (IL 2) receptors, and production of IL 2. As the concentration of PHA is decreased, each of these processes decreases in parallel. At suboptimal concentrations of PHA, the addition of exogenous IL 2 reconstitutes both the proliferative response and the expression of the IL 2 receptor, as measured by immunofluorescence with antibodies directed against the TAC/IL 2 receptor molecule, but without reconstituting the increase in [Ca²⁺](i). Therefore, the concentration dependence of responses to PHA appears to be secondary to an absence of IL 2 production due to a failure to induce an increase in [Ca²⁺](i). The addition of the calcium ionophores A23187 and ionomycin or of accessory cells to low concentrations of PHA induces increases in [Ca²⁺](i) and subsequent proliferative responses, suggesting that the two events are linked. The proliferative response can be inhibited by antibodies directed towards IL 2 or the IL 2 receptor, indicating that the proliferative response was at least partially dependent on the production and action of IL 2. This suggests that, although increases in [Ca²⁺](i) are an integral event in the induction of proliferation by PHA, the increase in [Ca²⁺](i) is required for the production but not the action of IL 2. In addition, low concentrations of PHA deliver an additional signal to cells, independent of an increase in [Ca²⁺](i), which induces IL 2 receptor expression and allows a proliferative response in the presence of exogenous IL 2.