Observations of Residual Differentiation Potential during Lineage Commitment

Abstract

We have recently described a subset of the multipotent progenitor pool that contains a common lymphoid progenitor. This subset of cells is lineage negative and expresses c-kit and Sca-1, but lacks expression of Thy 1.1 (Thy neg ). Based on the observation that lethally irradiated mice transplanted with these cells die from anemia unless supported with competitor marrow, we hypothesized that these progenitors lacked erythroid potential. We analyzed the erythroid potential of these cells by transplanting them into mice allelic at the hemoglobin locus and compared their erythroid potential with the Thy-1.1 low (Thy low ) subset that contains hematopoietic stem cells. We also performed CFU-C assays in methylcellulose containing recombinant cytokines and determined erythroid contribution to colonies using in situ benzidine staining. Donor-derived hemoglobin was observed following transplant of Thy neg cells, even though 19 of 20 of these animals died from anemia. In contrast, recipients of Thy low cells showed complete donor-derived engraftment 30 days following transplant. While approximately 60% of day 4 colonies derived from Thy neg cells expressed hemoglobin, by day 11 less than 5% were hemoglobinized. In contrast, greater than 70% of the Thy low subset contained hemoglobinized cells at the end of the observation period. A similar transient appearance of myeloid progeny was also observed in colonies derived from c-kit low Thy neg lymphoid progenitor cells. We conclude that these studies demonstrate commitment to the lymphoid lineage at the Thy low -to-Thy neg interface, and that the loss of erythroid and myeloid potential is gradual rather than abrupt. Hemoglobinized colonies may be undergoing apoptosis because of down-regulation of GATA-1 or because of a death signal from surrounding nonerythrocytic cells.

L-selectin defines a bone marrow analog to the thymic early T-lineage progenitor

“…on May 10, 2018. by guest www.bloodjournal.org From performance due to the higher sensitivity of in vitro assays for detecting progenitors with low proliferation potentials. 31,32 …”

Gradients of Antigen Expression and Developmental Potential in Hematopoiesis

“…Colony growth and differentiation were stimulated with recombinant cytokines, including steel factor (100 ng/mL), G‐CSF, Flt3L, IL‐3, IL‐6, IL‐7 (all at 10 ng/mL), and erythropoietin (4 U/mL). After the time intervals indicated in the figures, quadruplicate plates were stained using benzidine hydrochloride and scored for total colonies and for colonies containing hemoglobinized erythroid lineage cells colonies based on benzidine staining 14 . Results are reported as mean number of colonies per 100 cells plated, and error bars represent standard error values.…”

“…After the time intervals indicated in the figures, quadruplicate plates were stained using benzidine hydrochloride and scored for total colonies and for colonies containing hemoglobinized erythroid lineage cells colonies based on benzidine staining. 14…”