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In vivo proliferative and differential properties of murine bone marrow cells separated on the basis of rhodamine-123 retention.

We have studied the in vivo proliferation and differential properties of murine bone marrow cells (BMC) differing in mitochondrial activity. Following centrifugal elutriation, the cells were sorted on the basis of rhodamine-123 (Rh) fluorescence intensity within a predetermined light scatter window. Unfractionated BMC colonized the spleen with a characteristic preponderance of erythrocytic nodules upon infusion into heavily irradiated mice. In contrast, Rh-dull BMC formed few macroscopic spleen colonies up to day 12, but relatively more microscopic colonies when spleens were sectioned, and the majority of the colonies showed megakaryocytic (Meg) and/or granulocytic (G) differentiation. On day 16 many megakaryocytes were found in these spleens, and very large erythroid colonies had developed since day 12. In contrast, Rh-bright BMC were highly enriched for cells forming erythrocytic spleen colonies that disintegrated after day 12, but the percentage of G and Meg colonies was low. Spleens obtained from recipients of Rh-bright cells contained a negligible number of megakaryocytes on day 16. Apparently, the mitochondrial content of cells that form hemopoietic colonies in the spleen related to the onset and duration of their lineage expression, and to the relative sizes of the lineage compartments. Thus, the colony founders of the majority of spleen surface nodules observed on days 8 and 12 are cells with high mitochondrial activity forming transient erythrocytic nodules. Such spleen colonies represent the bulk of the countable nodules that form the basis of the widely applied murine "stem cell" assay.