Roles of dosage, pharmacokinetics, and cellular sensitivity to damage in the selective toxicity of cyclophosphamide towards B and T cells in development

Abstract

Cyclophosphamide (CP) is a known immunomodulating agent. When presented to either late stage chick embryos (e.g. 18 days of incubation (DI)) or neonatal chicks, CP induces selective B cell damage resulting in humoral immunosuppression in chickens. The present study was undertaken in order to provide further insights into CP's selective immunotoxic effects. We investigated the influences of age, CP-dose, and drug distribution on CP-induced cytotoxicity in the B and T cell compartments of the developing chick. In this test system, differential immunotoxicity was strongly dosage-dependent; at all ages tested, B cell depletion predominated at low CP dosages (50 and 100 mg/kg) whereas equally extensive lymphocyte toxicity was observed in both thymus and bursa at 200 mg/kg drug levels. Furthermore, while immunotoxicity profiles were similar at the two later developmental timepoints (18 DI and 1 day posthatch), significantly higher CP dosages were required to induce lymphoid damage at 12 DI. Pharmacokinetic studies with radiolabeled CP revealed that approximately two-fold higher levels of CP and its metabolites are taken up in bursal tissue as compared to the thymus. Experiments concerning the possible inherent differences in susceptibility to CP-induced mitotic inhibition and cell killing mediating selective toxicity towards B cells versus T cells showed that B cell mitosis was inhibited at CP dosages as low as 5 mg/kg. No such inhibition of T cell mitosis was observed at this same low dosage. However, mitosis was completely arrested in both B and T cells at 50 mg/kg CP. Observations of cellularity in immune organs shortly after CP exposure revealed that the bursa is at least ten times more sensitive than the thymus to CP-induced mitotic inhibition and killing of resident lymphocytes. These studies reveal that multiple factors are involved in modulating CP selective immunotoxicity in the developing embryo. These include the dosage level, preferential distribution of the drug to the bursa, and a much greater sensitivity of B cells to CP-mediated mitotic inhibition and cell killing.