Hematopoietic cells in cultures of the murine embryonic aorta-gonad-mesonephros region are induced by c-Myb

Definitive hematopoiesis begins in the para-aortic, splanchnopleural (P-Sp) and aorta-gonad-mesonephros (AGM) regions of mouse embryos and then switch es to the fetal liver [1-3]. Gene-targeted mice lacking the c-Myb transcrip tion factor have severe hematopoietic defects in the fetal liver [4], The r ole of c-Myb, if any, in P-Sp/AGM hematopoiesis has not been examined, howe ver, Recently, we reported that oncostatin M can effectively expand both he matopoietic and endothelial-like cells from in vitro cultures of the AGM re gion [5]. Using this cell culture system, we examined the involvement of c- Myb in definitive hematopoiesis in the P-Sp and AGM regions. When primary c ultures from the P-Sp or AGM regions of wildtype mouse embryos were probed with an anti-c-Myb antibody, hematopoietic cells but not endothelial like c ells showed positive staining. In contrast, in the P-Sp/AGM culture from c- myb(-/-) embryos, no hematopoietic cells were generated and endothelial lik e cells predominated, indicating that the impairment of hematopoiesis in th e liver of c-myb(-/-) embryos is actually preceded by a defect in P-Sp/AGM hematopoiesis. Hematogenic precursor cells were, however, still present in an inert but competent form among the endothelial-like, adherent cell popul ation of c-myb(-/-) P-Sp/AGM cultures. When infected with a retrovirus carr ying c-myb cDNA, these cultures gave rise to a significant number of hemato poietic cells. The rescued cells, unlike wild-type hematopoietic cells, wer e negative for c-Kit (a marker of hematopoietic progenitors), but did expre ss other hematopoietic cell surface markers such as Mac-1, Gr-1 (myeloid ma rkers), CD19, B220, Thy-1.2 (lymphoid markers), and Ter119 (an erythroid ma rker). Thus, c-Myb plays a role in the generation of hematopoietic cells in the embryonic P-Sp and AGM regions.