Resistance and susceptibility to cyclosporin A as CD3-4-8- human thymocytes differentiate in vitro

Human T-cell development appears to be relatively resistant to cyclosporin A (CsA). Children exposed to CsA in utero as part of kidney transplant maintenance have few abnormalities. The objective of the study described here was to analyse the effects of CsA on the development in vitro of human multinegative (MN) (CD3-4-8-) thymocytes as a model system for thymic progenitor development in vivo. MN thymocytes, prepared by depletion methods, differentiated in vitro to acquire CD3 and undergo transitions in CD45 isoform expression analogous to those postulated to occur in vivo. In this work MN thymocytes were cultured with IL-2 and on thymic epithelial cells (TEC) with or without IL-2, either in the presence or absence of CsA. For many thymocyte preparations, differentiation in the presence of CsA resulted in almost complete inhibition of the acquisition of CD3 and of the low Mr isoform CD45R0. Expression of CD45RA and of total CD45 were reduced but not eliminated and the density of CD29 was unaffected. For others, neither CD3 nor CD45 expression was affected, but selective inhibition of TCR delta expression TCR delta expression occurred. At all doses of CsA (0.1-100 micrograms/ml), MN thymocytes continued to cycle indicating a CsA-resistant generative compartment. Treatment of peripheral blood T cells with CsA had no effect on surface expression of CD3 or CD45 isoforms but did reduce the amount of de novo-synthesized CD45R0 mRNA. Culture of MN thymocytes on TEC rendered them virtually resistant to the negative effects of CsA. CD3 acquisition was unhindered and total CD45 remained high, but the transition from CD45RA to CD45R0 appeared to be delayed. In the absence of TEC, expression of both TCR alpha beta and of TCR delta was inhibited, but on TEC, TCR delta was actually up-regulated in some conditions. The effects of CsA on human thymocyte development appeared to be modulated by the physiological state of the donor and the growth conditions to which the cells were subjected. Conditions which most closely approximated those manifest in vivo rendered thymocytes most resistant to the negative effects of CsA. The amount of CsA required to affect differentiation in vitro was significantly higher than could be attained in vivo suggesting that the immunomodulatory effects of CsA in the maintenance of organ transplants may derive from an as yet uncharacterized mechanism.