Characterization of prothymocytes with cloning capacity in human bone marrow

The identity of human bone marrow (BM)-derived T cell precursors with colony forming capacity has led to controversy because of contamination with mature clonogenic T cells. We achieved 2 Log elimination of mature T cells from BM using a cocktail of monoclonal antibodies: CD2, CD3, CD4, CD6, and CD8 followed by two successive baby rabbit C' treatment. T cell depleted BM can generate colonies of CD2+, CD3+, Ti+, mostly CD4+, in the presence of PHA, rIL2, and a prothymocyte differentiating activity derived from phytohemagglutinin (PHA) induced mononuclear cells. These precursors could be enriched three- to sixfold by percoll gradient centrifugation and then significantly bypass the number of contaminant mature T cells as shown by limiting dilution analysis. Colony generation by marrow precursors was inhibited by the addition of autologous T cells. This inhibition was mostly caused by the T8+ subset. CFU-TL growth was dramatically inhibited by eliminating CD7+ cells suggesting their positivity for this surface marker. These precursors needed major histocompatibility complex (MHC) II-positive cells for optimal growth but lack DR themselves.