Characterization of prethymic progenitors within the chicken embryo.

The thymic primordium in both birds and mammals is first colonized by cells emerging from the intra-embryonic mesenchyme but the nature of these precursors is poorly understood. We demonstrate here an early embryonic day 7 prethymic population with T lymphoid potential. Our work is a phenotypic analysis of, to date, the earliest embryonic prethymic progenitors arising in the avian para-aortic area during ontogeny. The phenotype of these cells, expressing the cell surface molecules alpha2beta1 integrin, c-kit, thrombomucin/MEP21, HEMCAM and chL12, reflects functional properties required for cell adhesion, migration and growth factor responsiveness. Importantly, the presence of these antigens was found to correlate with the recolonization of the recipient thymus following intrathymic cell transfers. These intra-embryonic cells were also found to express the Ikaros transcription factor, the molecular function of which is considered to be prerequisite for embryonic lymphoid development.

Antigenic phenotype of early intra-embryonic lymphoid progenitors in the chicken.

The stem cells for the definitive haematopoiesis are derived from intra-embryonic sources originally described in an avian model and later also in mammals. However, the molecular make-up of the early embryonic haematopoietic progenitors is not yet clearly defined. We have recently characterized the phenotype of prethymic intra-embryonic progenitors capable of thymic colonization. Here we studied the ontogeny of cell-surface antigens HEMCAM, alpha2beta1 integrin, thrombomucin, chL12 and c-kit and their co-expression on prethymic T-cell progenitors. The early intra-embryonic expression of avian B-cell antigen chB6 was also demonstrated on cells derived from the intra-embryonic areas. We suggest that in the chicken, embryonic B-cell progenitors segregate earlier than T-cell progenitors in the differentiation of multipotent haematopoietic stem cells to committed progenitors.

Expression of chL12 surface antigen is associated with cell survival in the avian bursa of Fabricius.

During B-cell development in the avian bursa of Fabricius most of the developing B cells die by apoptosis and only a minority survive to emigrate into the periphery. Recently, it has been shown that when developing bursal cells become mature and ready to migrate they start to express chL12 antigen. The expression of this cell-surface molecule was found to be associated with the survival of the bursal cells both after in vitro culture and after in vivo cyclophosphamide (CY) treatment. The frequency of early apoptotic cells in freshly isolated bursal cells was found to be high. The high susceptibility of these cells to apoptosis is in line with the finding of low bcl-2 mRNA expression. We conclude that expression of avian chL12 antigen is associated with the survival of bursal cells.

Intraembryonic haemopoietic cells and early T cell development.

T cell precursors in the chick embryo have been localized into the intraembryonic mesenchyme (IEM) and into the para-aortic region before the first wave of the thymic colonization on embryonic day (ED) 6,5-8. The cell surface markers of avian prethymic stem cells are not known. It is also not known whether these precursor cells are already committed to the T cell lineage before their thymic colonization. In 7-day-old chick embryos Ov+ cells were found in the para-aortic region. Also the endothelial cells of the embryonic dorsal aorta were positively stained. Ov antigen might represent a very primitive marker for precursor cells having the potentiality to differentiate both to haemopoietic and endothelial cells. Scattered CD45+ cells were observed in the same para-aortic area as in many haemopoietic areas in the loose embryonic mesenchymal tissues. CD8 alpha (MoAb 3-298) expressing haemopoietic cells were detected before thymic colonization on ED6. In flow cytometric analysis of IEM precursors Ov, CD45 and CD8 alpha expressing cells seemed to form distinct subsets suggesting heterogeneity of these haemopoietic cells.

Intra-embryonic haemopoietic cells and early MHC expression.

In the avian embryo the haemopoietic stem cells originate from the intra-embryonic area near dorsal aorta. The surface-marker expression of haemopoietic stem cells and their potential to produce different haemopoietic cells are still largely unknown. The surface antigen expression and particularly the MHC antigen expression on intra-embryonic haemopoietic cells was studied. Expression of B-F antigens, homologous to mammalian MHC class-I antigens, was found already on embryonic day (ED) 5. The first B-L antigens, analogous to mammalian MHC class-II antigens, were detected also from ED5 onwards. The appearance of surface antigens defined by MoAbs T10A6 and 3-298 during embryogenesis also was studied. The antigen defined with T10A6 was detected from ED4 onwards on endothelial cells but not on haemopoietic cells in the para-aortic region. The first 3-298+ haemopoietic cells were found on ED6, whereas endothelial cells were negative. These findings imply that some surface markers are shared with haemopoietic and endothelial cells indicating either a common embryonic origin or the importance of these molecules in embryonic stem-cell homing.