Characterization of Triple Negative Clones Isolated from Post-Natal Human Thymus

Abstract

Human triple negative (CD3^- CD4^- CD8^-) thymocytes and double negative (CD4^- CD8^-) thymocytes purified from post-natal thymus were cloned with a feeder cell mixture of irradiated PBL, irradiated JY cells and PHA and expanded with IL-2. The cloning efficiency of triple negative thymocytes was less than 1% and the majority of the clones were triple negative. One out of 11 clones was TCR αβ⁺ CD4⁺. No TCR γδ ⁺ clones were isolated. On the other hand, the cloning efficiency of double negative thymocytes was about 10% and most of the clones isolated were TCR γδ ⁺. We could not find any evidence of in vitro differentiation of triple negative thymocytes into TCR γδ ⁺ cells. Some of the triple negative clones expressed CD16^- brightly and were apparently NK cells. All CD 16-clones isolated from triple negative thymocytes, however, expressed NKH1, which is also an NK cell marker. Cytoplasmic CD3-δ and CD3-ε Ag which have been reported to be expressed in the most immature thymocytes were not detected in any of these clones. Furthermore, the CD 16^- triple negative clones exhibited significant cytolytic activity against K562. Pheno-type of the clones seems to be stable under various conditions in vitro including coculture with human thymic epithelial cells. These data indicate that the CD16^- triple negative clones isolated from triple negative thymocytes are similar to a minor subset of NK cells which is CD16^- NKH-1⁺. It is not clear whether they originated from a distinct subset of mature or immature NK cells resident in the thymus tissue or from common precursors of both T and NK lineage.