whole_WoodsGregoryMark1984_thesis.pdf (8.36 MB)
Studies on human T lymphocyte colonies
thesisposted on 2023-05-27, 12:14 authored by Woods, Gregory Mark
This thesis examined the ability of human T lymphocytes to form colonies in semisolid agar. A two step procedure requiring a continued supply of PHA, a source of SRBC during the agar culture and a high agar concentration to restrict the freely mobile proliferating cells was developed for the culturing of human lymphocytes in semisolid agar. The technique was later modified by replacing PHA with an IL2-containing lymphokine during the agar phase. The T cell nature of the colonies was confirmed by their ability to form E-rosettes. An absence of colonies from E-rosette depleted cells compared to E-rosette enriched cells strengthened this finding and indicated that the colony forming cells were T cells. The requirement for SRBC was investigated and it was found that red cells from a variety of mammalian sources supported colony formation through the elaboration of soluble factors which diffused through the agar and did not require cell-cell contact for their expression. Experiments relating to the cellular interactions involved in colony formation emphasised that cooperation between at least three cell populations was essential. The interaction of these cells could be described by two reactions. The first reaction required an adherent cell (probably monocyte) population which, together with PHA, provided the necessary signals for T cell activation. The activated T cells interacted during the second or proliferative reaction. In response to PHA, accessory T cells released IL2 which permitted colony forming cells to proliferate. Thus PHA-induced T colony formation required the cooperation of adherent cells for the activation reaction (T ->T') and IL2-producing T cells for the proliferative reaction (T' -> nT'). The microfilament/Microtubule network and the redistribution of receptor :membrane complexes appeared to be an important feature of T cell function with regard to both mitogen and IL2 binding. Utilising the colony forming ability of T lymphocytes studies on radiation induced mitotic death were carried out. Colony formation was sensitive to low doses of irradiation although PHA offered a limited degree of protection when cells were irradiated prior to, or in the early stages of activation. This protection could be explained by activities of repair enzymes which coincided with early cellular activation events. The human T lymphocyte colony assay was found to be a valuable in vitro model for studying the steps involved in T lymphocyte activation and proliferation.
Rights statementCopyright 1984 the Author - The University is continuing to endeavour to trace the copyright owner(s) and in the meantime this item has been reproduced here in good faith. We would be pleased to hear from the copyright owner(s). Thesis (Ph.D) - University of Tasmania, 1984. Bibliography: leaves 142-152