Signal requirements for the in vitro differentiation of cytotoxic T lymphocytes (CTL): distinct soluble mediators promote preactivation of CTL‐precursors, clonal growth and differentiation into cytotoxic effector cells

Abstract

We describe here that the requirement of accessory cells for the polyclonal activation of high-density (resting) murine T lymphocytes can be bypassed by soluble mediators present in culture supernatants of concanavalin A (Con A)-activated murine spleen cells. Induction of responsiveness is confined to Lyt-2+ T cells; GK 1.5+ T helper cells require signals provided by accessory cells. Using this system the lymphokine signal requirements for the polyclonal activation of Lyt-2+ cytotoxic T lymphocyte (CTL) precursors could be defined. We show that Con A fails to trigger in Lyt-2+ responder T cells the expression of interleukin 2 (IL 2) receptors and assume that this explains why recombinant (rec) DNA-derived IL 2 fails to induce proliferative responses. Complementation of rec IL 2 with an IL 2 receptor-inducing factor (RIF) induces proliferative responses. RIF alone triggers IL 2 receptor expression in 10-12% of Lyt-2+ T cells exposed to Con A. This lymphokine appears to be distinct from colony-stimulating factor 1, IFN-gamma and IL 1. Resting Lyt-2+ T cells cultured in limiting numbers in the presence of Con A, RIF plus rec IL 2 do proliferate, yet exhibit no cytolytic activity. Differentiation into CTL can be brought about by addition of cytotoxic T cell differentiation factor (CTDF). We conclude that the polyclonal activation pathway of CTL from resting CTL precursors can be subdivided into three stages: preactivation, clonal growth and CTL differentiation. Each of these stages appears to be controlled by a distinct lymphokine, RIF, IL 2 and CTDF, respectively.

Interleukin 5, a T-cell-derived B-cell differentiation factor also induces cytotoxic T lymphocytes.

“…Early studies by Raulet and Bevan (24) and other investigators strongly suggested that a non-IL-2 lymphokine was required for the generation of CTL (25)(26)(27)(28)(29). With our approach, using PNA+ thymocytes, the generation of CTL against TNP-modified syngeneic spleen cells in the presence of low concentrations of IL-2 (10 units/ml) requires the presence of the other factor, KHF (12)(13)(14).…”

“…Wagner and colleagues (28) reported that the receptor-inducing factor that induces IL-2 receptors is required for induction of CTL from resting Lyt-2+ splenic T cells. Falk et al (29), using PNA-thymocytes as responders and glutaraldehyde-fixed TNP-modified spleen cells as stimulators, also found it necessary to add T-cell cytotoxic inducing factor type 1 (TCF1) other than IL-2 whether these lymphokines are the same or distinct molecules from the TRF and KHF described here.…”

Developmental control of lymphokine gene expression in fetal thymocytes during T-cell ontogeny.

“…Studies seeking to examine this issue by measuring the production and use of growth factors by thymocytes have depended on the stimulation of these cells by phorbol esters and ionophores outside the thymic environment (1)(2)(3)(4)(5). These results may not be representative ofevents inside the intact organ for several reasons.…”

Differentiation of human mature thymocytes: Existence of a T3+4−8− intermediate stage

“…In this regard, relevant differences in the effector populations tested could account for the discrepancies, which might rather respond to a yet ill-defined heterogeneities within early thymocyte compartment ([ 181, M. L. Toribio et al, submitted). Thus, mouse effector cells were either nonfunctional double-positive "cortical" cells [ 161 or receptor less and still double-negative precursors [35], while human thymocytes bore functional T3 complexes [lo].…”