Human T helper cell differentiation is regulated by the combined action of cytokines and accessory cell-dependent costimulatory signals

We have developed an in vitro differentiation model for human Th cells to study the role of cytokines and accessory cell-dependent costimulatory signals in this process. Peripheral blood-derived CD4+ "naive" (CD45RA+ RO-) T cells were stimulated in weekly intervals with immobilized anti-CD3 mAb, accessory cells, and exogenous cytokines, and were analyzed for cytokine secretion pattern. With the B cell line JY (B7-1+ B7-2+), as source of accessory cells, we could generate distinct Th subsets. Coculture with the combination of recombinant human (rh) IL-1beta and rhIL-6 gave rise to Th0-like cells, which secreted low levels of IFN-gamma and IL-5. The addition of rhIL-12 led to the generation of Th1-like cells, which secreted high levels of IL-2, IFN-gamma, TNF-alpha, and upon multiple stimulations, significant levels of IL-10. The presence of rhIL-4 induced Th2-like cells that secreted high levels of IL-5 and IL-13, but undetectable levels of IL-4. Only after stimulation with phorbol ester and calcium ionophore could these Th2-like cells be induced to secrete significant levels of IL-4, indicating distinct stimulatory requirements for the induction of IL-5 and IL-13 compared with IL-4. The B7-1-negative monocytic cell line U937 could only provide accessory cell-dependent costimulatory signals for the generation of Th1-like cells, while B7-1-transfected U937 cells acquired the capacity to provide costimulation for the generation of Th2-like cells. These results indicate a differential dependence on CD28-mediated costimulation for the generation of human Th1-like and Th2-like cells.