Francis C. Monette

Myeloid stem cell kinetics during erythropoietic stress.

Summary. The committed myeloid stem cell compartment, monitored by the agar colony technique, has been assayed in the bone marrow, spleen and blood of mice during erythropoietic stress. When red cell production appeared maximal, the size of the committed myeloid stem cell comparment was reduced in the bone marrow and there was usually a reciprocal rise in the spleen with varying numbers of these cells in the circulation. The role of stem cell competition and migration in the development of these changes is discussed.

HEMOPOIETIC STEM CELL PROLIFERATION AND MIGRATION FOLLOWING BORDETELLA PERTUSSIS VACCINE

The ability of a single injection of killed, intact bacteria to effect an increase in the proliferative rate of hemopoietic stem cells was studied. The total numbers of colony forming units in bone marrow, spleen and peripheral blood as well as the proportion of CFU in cycle was assessed. Splenic CFU were observed to rise exponentially due initially to in situ proliferation and later to proliferation in bone marrow with migration via the blood to the spleen. The results are discussed in the light of current concepts of stem cell regulation.

Studies on the Regulation of Granulopoiesis

Tritiated thymidine autoradiography was used to study the control of granulocyte production and release using the irradiated leg shielded mouse as an experimental model. Neutropenia resulted in a shortening of 24–36 hr in the mean transit time through the non‐proliferating granulocyte compartment. There was little difference between neutropenic and control animals in labelling indices of the cells in the proliferative granulocyte compartments, which suggests that the granulocytic hyperplasia observed in the neutropenic mice was predominantly due to differentiation of morphologically unrecognizable precursor cells rather than increased proliferation of morphologically recognizable cells. The persistence of a labelling index of approximately 60% in the myeloblast‐promyelocyte compartment 24 hr after injection suggests that these precursor cells were rapidly proliferating.

Hydroxyurea-Induced Erythroid Differentiation

Summary Hydroxyurea, a cytotoxic agent which destroys cells in DNA synthesis, has been shown to evoke the differentiation of a small number of hemopoietic precursor cells in the erythroid series of erythropoietically suppressed hypertransfused mice. This effect does not appear to be mediated by erythropoietin (EP) since the simultaneous injection of anti-EP did not alter this response.

Autoradiographic Studies of Human Lymphocytes Cultured in Vivo

Summary Human lymphocytes were cultured in diffusion chambers implanted into the peritoneal cavities of rats. In cultures treated with PHA, the pattern of morphologic transformation and initiation of DNA synthesis paralleled closely similar studies made in vitro. Immunologic stimulation of the human cells by the heterologous host was presumably minimal or absent since control cultures (without PHA) showed no significant degree of blastogenesis when compared with PHA-treated cultures. Minimum DNA synthesis time determined for the PHA-stimulated lymphocytes was 9–10 hr. Values found for the duration of Tc (16–18 hr) and minimum TG2 (2 hr) were shorter than those reported for similar studies in vitro. The in vivo culture method using Millipore chambers appears to offer a more physiologic method for studying lymphocytes.

Studies on the Regulation of Diffusion Chamber Granulopoiesis

We have evaluated the role of colony-stimulating activity (CSA) and stem cells in diffusion chamber (DC) myelopoiesis. CF1 host mice were implanted with DC with 0.5 x 106 normal marrow cells and then injected for 3–4 days intravenously with 5 μg Salmonella typhosa endotoxin or saline. There were no significant increases in DC myelopoiesis, but significant increases in host animal total tibial proliferative granulocytes on day 4 and suggestive increases in nonproliferative granulocytes on day 7 of DC culture. Further studies utilizing progressively increasing doses of intravenous endotoxin in irradiated or normal host mice implanted with DC with normal marrow showed: (1) a lack of correlation of serum CSA or inhibitor level with DC myelopoiesis; (2) elevations of CSA levels within DC relative to serum levels in all groups; and (3) a general inverse correlation of DC myelopoiesis with marrow cellularity.

Utilization of Mouse Stem Cell-Depleted Marrow in the Study of Diffusion Chamber Myelopoiesis

Abstract Mouse brain antisera with anti-CFU-S activity was utilized to evaluate the role of the pluripotent stem cell (CFU-S) in a diffusion chamber (DC) myelopoiesis. Antisera incubation with marrow resulted in a 93 ± 2% reduction in detectable CFU-S in the standard spleen colony assay and a variable reduction in the granulocyte-monocyte progenitor cell (CFU-C). Marrow virtually devoid of detectable CFU-S and containing variable CFU-C relative to control demonstrated equal or greater numbers of CFU-S, CFU-C, and differentiated progeny at varying time intervals after DC implantation. These results may indicate that neither the CFU-S nor the CFU-C is the progenitor cell for DC growth. Alternatively, CFU-S antibody may not kill CFU-S in vitro or in the in vivo DC system but may opsonize the cell prior to eventual reticuloendothelial destruction. The latter concept is supported by reversal of antisera effect on CFU-S recovery after in vitro exposure of antisera-treated cells to pronase.