Lymphoid differentiation of the hematopoietic stem cell that reconstitutes total erythropoiesis of a genetically anemic W/Wv mouse

Abstract

We investigated whether the stem cell that reconstitutes total erythropoiesis of a WBB6F1-W/Wv mouse differentiates into lymphoid lineage. The electrophoretic pattern of hemoglobin was used as a marker of the reconstitution; 3-phosphoglycerate kinase (PGK), an X chromosome- linked enzyme was used as a tool for estimating clonality. We injected 10(5) bone marrow cells of 5-FU treated C57BL/6-Pgk-1b/Pgk-1a female mice, in which each stem cell had either A-type PGK or B-type PGK due to random inactivation of one of two X chromosomes, into genetically anemic (WB x C57BL/6)F1-W/Wv (hereafter WBB6F1-W/Wv) mice that contained only B-type PGK. The recipient WBB6F1-W/Wv mice, in which erythropoiesis was reconstituted with donor cells for a long term, were killed and the PGK patterns of bone marrows, thymus, lymph nodes, and Peyer's patches were examined. A considerable amount of A-type PGK was detected in the lymphoid organs of the WBB6F1-W/Wv mice in which erythrocytes showed only A-type PGK when killed. In contrast, A-type PGK was scarcely detectable in the lymphoid organs of the WBB6F1-W/Wv mice in which erythrocytes showed only B-type PGK when killed. The present results suggest that the hematopoietic stem cells estimated by the erythropoiesis reconstituting assay differentiate into lymphoid lineage and that the long-term erythropoiesis reconstitution assay is useful for detecting the true primitive hematopoietic stem cells.

High c-Kit expression identifies hematopoietic stem cells with impaired self-renewal and megakaryocytic bias

“…Although various strategies to identify long-term HSCs based on surface antigen expression have been described, almost all immunophenotypic definitions of HSCs rely on expression of c-Kit, which is a functional signaling receptor critical for maintaining HSC self-renewal and niche interactions (McCulloch et al, 1964; Maloney et al, 1978; Nakano et al, 1989; Wang and Bunting, 2008). Remarkably, despite the fact that the most highly purified HSCs exhibit a log-fold variation in expression of this receptor that is known to be critical for HSC function, relatively little effort has been devoted to investigating whether HSCs with varying c-Kit levels may represent functionally distinct HSC subtypes.…”

In vitro proliferation of primitive hemopoietic stem cells supported by stromal cells: evidence for the presence of a mechanism(s) other than that involving c-kit receptor and its ligand.

“…It is well established that a single stem cell can give rise to cells of all lymphohemopoietic lineages (2)(3)(4). Such stem cells have been operationally defined and assayed by their capacity for long-term repopulation of lymphohemopoietic cells after injection of them into either lethally irradiated or genetically anemic WBB6F1-W/W ~ mice (5)(6)(7)(8). These stem cells are physically separable from the majority of the cells that form macroscopic colonies in the spleens of lethally irradiated mice (spleen colony-forming units [CFU-S] 1) (9-11).…”

“…The hemopoietic cells from individual dishes were intravenously injected into each WBB6F1-W/W ~ mouse. At 8,12,16,20, and 24 wk after the injection, peripheral blood of the mice was obtained by retroorbital puncture under anesthesia. Electrophoretic pattern of hemoglobin was determined after modification with cystamine according to the method described by Whitney (33).…”

Characterization of spleen colonies derived from mice with mutations at the w locus

“…This confirmed the necessity of lysing red blood cells prior to tissue evaluation to avoid contamination. Thymus was used as a negative control because lymphocyte replacement occurs more slowly than erythrocyte replacement (Harrison and Astle, 1976;Nakano et al, 1989a). Thymus contained donor cells in only three of the 49 mice tested, confirming minimal contribution from donor cells.…”