Studies on the thymus and ontogeny of lymphocyte heterogeneity in the clawed toad Xenopus laevis (Daudin)

Abstract

(1) Functionally mature lymphocytes can be demonstrated in vitro by their proliferation (measured by scintillation counting) in response to mitogens. Studies in Chapter 2 examine the responsiveness of adult Xenopus lymphocytes from both the thymus and the spleen to B and T cell mitogens. Optimum mitogen doses and levels of foetal calf serum supplementation are established. Good stimulation indices are found when splenocytes are stimulated with both PHA and Con A. Thymocytes also respond well to these T cell mitogens, but a small response to B cell mitogens (E. coli LPS and PPD) is also found. (2) The ontogeny of lymphocyte responsiveness to T and B cell mitogens is examined in Chapter 3. Very little stimulation is found in the larval thymus, but shortly after the end of metamorphosis a thymocyte response begins to emerge and by 6 months of age good stimulation is found with both T and B cell mitogens. While good levels of thymocyte stimulation with the T cell mitogens are maintained in later life, the response to the B cell mitogen LPS declines to only a low level by 12 months of age. Mitogenic responses emerge in the spleen after the end of metamorphosisin a similar manner to thymocytes and good responses were found in all the subsequent ages tested. The transient response of thymic cells to LPS, which is maximal at 6 months of age, was confirmed by auto radiography and was shown to be dependent on a population of nylon-wool adherent lymphocytes. (3) The larval thymus plays a critical role in the establishment of T cell functions, and Chapter 4 looks at the effect of thymectomy at different stages of larval development on the subsequent alloimmune response, mixed lymphocyte reactivity and T cell mitogen responsiveness. The results reveal that (a) skin allograft rejection displayed by 7-day thymectomized animals is effected in the absence of spleen and blood lymphocytes reactive to T cell mitogens, (b) T-mitogen responsive cells and MLR T cells cannot easily be separated into distinct populations in terms of their degree of thymus dependency. (4) Chapter 5 looks at the ontogeny of both T helper cell and B cell function by examining the in vivo antigen-binding reactivity of splenocytes to a T-dependent (TNP-SRBC) and a T-independent (TNP- LPS) antigen. Good levels of TOP binding cells can be induced in the spleen when larvae are only 3 weeks old following injection of TNP-SRBC (after low dose priming with SRBC) or TNP-LPS. The level of antigen binding in both cases appears to peak at metamorphosis. However, when cellular antibody production was measured, no plaque forming cells could be induced in the spleen until after the end of metamorphosis. (5) Chapter 6 presents (a) a brief histological study of the thymus in late larval life and at metamorphosis. This work characterises the dramatic depletion of thymocyte numbers that occurs at the end of metamorphosis and the renewed lymphopoiesis immediately afterwards, and (b) preliminary results on the effect of 7-day organ culture on thymus histogenesis.