Characterization and purification of a primitive hematopoietic cell type in adult mouse marrow capable of lymphomyeloid differentiation in long-term marrow "switch" cultures (original) (raw)
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Immunological Reviews, 2002
The earliest progenitors of lymphocytes are extremely rare and typically present among very complex populations of hematopoietic cells. Additionally, it is difficult to know how cells with any given set of characteristics are developmentally related to stem cells and maturing lymphoid precursors. However, it is now possible to divide bone marrow into progressively smaller fractions and exploit well defined culture systems to determine which ones contain cells that can turn into lymphocytes. Analysis of steroid hormone sensitive cells and use of two-step cultures is providing additional information about the most likely differentiation pathways for B and NK lineage lymphocytes. A newly identified category early lymphoid progenitors (ELP) can now be sorted to high purity from RAG1/GFP knock in mice. Furthermore, the same experimental model makes it possible to image lymphoid progenitors in fetal and adult hematopoietic tissues.
Vox Sanguinis, 1979
Myeloid and lymphoid stem cell colony formation (GM-CFU and L-CFU) was studied in patients with lymphoproliferative diseases, aplastic anemia and other hematological abnormalities. Most patients with acute lymphatic leukemia had low number of L-CFU with decreased or normal GM-CFU, while in Hodgkin's disease and chronic lymphatic leukemia L-CFU growth was very poor with only minor abnormalities of GM-CFU formation. Aplastic anemia was characterized by a decreased CM-CFU and normal L-CFU. Coculture studies suggested that a diminished colony formation may be linked to circulating lymphocytes that inhibit L-CFU as well as the reduction in number of precursor cells. 10 Shen, J.; Wilson, F.; Shifrine, M., and Gershwin, M. E.: Select growth of human T lymphocytes in single phase semisolid culture. J. h imun. f19:
Rapid, B Lymphoid-Restricted Engraftment Mediated by a Primitive Bone Marrow Subpopulation
The Journal of Immunology, 2000
Utilizing multiparameter flow cytometry, we have defined a subset of bone marrow cells containing lymphoid-restricted differentiation potential after i.v. transplantation. Bone marrow cells characterized by expression of the Sca-1 and c-kit Ags and lacking Ags of differentiating lineages were segregated into subsets based on allele-specific Thy-1.1 Ag expression. Although hematopoietic stem cells were recovered in the Thy-1.1low subset as previously described, the Thy-1.1neg subset consisted of progenitor cells that preferentially reconstituted the B lymphocyte lineage after i.v. transplantation. Recipients of Thy-1.1neg cells did not survive beyond 30 days, presumably due to the failure of erythroid and platelet lineages to recover after transplants. Thy-1.1neg cells predominantly reconstituted the bone marrow and peripheral blood of lethally irradiated recipients with B lineage cells within 2 weeks, although a low frequency of myeloid lineage cells was also detected. In contrast, ...
Experimental Hematology, 2000
Objective. In previous work, we showed that CD34 ϩ bone marrow cells can be successfully expanded along the myeloid pathway in stroma-and serum-free conditions in the presence of SCF ϩ IL-3 ϩ IL-6 ϩ Flt3-l ϩ G-CSF ϩ MGDF. Due to the lack of phenotypically detectable lymphoid cells, it was necessary to address the question of the lymphoid potential of the expanded populations under these conditions. Materials and Methods. The present report describes a long-term culture system that supports human Band NK-cell differentiation from the day 14 fraction without further selection of the more primitive cells. In NK proliferation assays, the cells were maintained over stroma cells in the presence of IL-2 for 4-5 weeks. NK initiating cells (NK-IC) were determined by a limiting dilution assay. In B-cell cultures, the expanded cells were maintained over MS5 in the presence of Flt3-l for 4-8 weeks. Results. NK cells rose from 0.2% Ϯ 0.04% at culture initiation to 71% Ϯ 6% at week 5. These cells displayed cytolytic activity. NK-IC evaluation showed a mean 18-fold expansion in the day 14 expanded fraction as compared to the initial day 0 fraction. Similarly, CD19 ϩ cells rose from 0.1% at culture initiation to 30% Ϯ 1% at week 6. Cells produced under these B-LTC conditions were CD34 Ϫ CD19 ϩ CD10 ϩ. We also demonstrated that the CD34 ϩ /Lin Ϫ sorted cells from the day 14 fraction gave rise to NK and B cells. Conclusion. This culture system permits the revelation of a population that, although poorly represented in terms of phenotypically detectable cells, nevertheless retains high levels of lymphoid NK and B potential after 14 days expansion. Such data suggest the persistence, or expansion, of lymphoid progenitors and, hence, the multipotentiality of the expanded progenitor/stem cells.
Clonal proliferation of murine lymphohemopoietic progenitors in culture
Proceedings of the National Academy of Sciences, 1992
We have used a two-step clonal culture system to unequivocally demonstrate that individual primitive lymphohemopoietic progenitor cells have the capacity for differentiation along either the myeloid or the B-lymphoid lineage. Highly enriched murine marrow cells were plated individually in culture by micromanipulation in the presence of pokeweed mitogen-stimulated spleen cell conditioned medium, erythro
Characterization of engraftable hematopoietic stem cells in murine long-term bone marrow cultures
Experimental Hematology, 2001
Objective. Long-term bone marrow cultures (LTBMC) are a potential source of hematopoietic stem cells (HSC) for transplantation. Previous reports indicate that feeding LTBMCs induces hematopoietic progenitor cycling, and other studies link HSC cycle phase with engraftability. Our study was initiated to further characterize LTBMC engraftability and determine if a cycle phase-related engraftment defect affects HSC from LTBMCs. Materials and Methods. Competitive repopulation of lethally irradiated BALB/c females was used to examine engraftability of LTBMCs under "fed" or "unfed" conditions at 3 to 5 weeks culture. Tritiated thymidine suicide was used to determine the cycle status of HPP-CFC and CFU-S from LTBMCs. Results. Total cell number in LTBMCs decreases from input. Quantitatively, both fed and unfed 3-, 4-, or 5-week cultures compete strongly with fresh marrow for 2 and 8 weeks, but not 6 months, after transplantation. Short-term engraftable HSCs expand between 3 and 5 weeks of culture. Clonal assays indicate no peak in S-phase of CFU-S at 24 and 48 hours after feeding, and fluctuation in both content and cycle status of HPP-CFC after feeding. Conclusions. Our LTBMCs engraft in all conditions, and the level of engraftment capability does not correlate with cell-cycle phase of CFU-S or HPP-CFC, or with time from feeding. Although the total cell number decreases from input, the proportion of short-and intermediateterm engrafting HSC in whole LTBMCs approximates that of fresh marrow and expands from 3 to 5 weeks in culture, whereas long-term engraftable HSCs are decreased in culture.
Blood, 1984
T cell differentiation in human marrow was studied in Dexter type long- term bone marrow cultures. In these cultures, T lymphocyte colony- forming units (TL-CFU), E rosette-forming cells (E+), and T3+, T4+, and T8+ cells (assayed by indirect immunofluorescence) were found to be present for at least 7 weeks. It was investigated whether the existence of T cells in long-term culture resulted from the persistence of inoculated T lymphocytes or from the production by immature progenitors. No significant numbers of E+, T3+, T4+, or T8+ cells were detected in cultures that were established from E+ lymphocyte-depleted bone marrow, indicating little or no production of T lymphocytes from E- negative precursors. On the other hand, bone marrow cells purged of E+ lymphocytes did not contain TL-CFU, but appeared to regain high numbers of TL-CFU during Dexter culture; this suggested that an earlier step in T cell differentiation may take place in this culture system. The generation of TL-CFU in t...
African Journal of Pharmacy and Pharmacology, 2021
Initial myeloid and lymphoid differentiation of hematopoietic and non-hematopoietic stem/progenitor cells by appropriate ex vivo-and in vitro-incubation was evaluated. So, hematopoietic stem/progenitor cells from mouse spleen were ex vivo-incubated in the presence of different interleukins, as well as of various combinations of them. Non-hematopoietic mouse embryonic stem cells (mESCs) were in vitroincubated in the presence of GM-CSF and malignant antigens of human cervical carcinoma cells HeLa. Analogically, mouse embryonic fibroblasts from line 3T3 were pre-incubated in the presence of malignant antigens from mouse myeloma cells. Possibilities for derivation of myeloid and lymphoid cells by hematopoietic and non-hematopoietic cellular progenitors were proposed. The literature ability for production of immune molecules, including membrane glycoprotein receptors and antibodies/immunoglobulins by initial myeloid and lymphoid cells, derived from hematopoietic, as well as by non-hematopoietic cellular progenitors was accepted when appropriate factors are available. Because the produced antibodies are out of the germinative centers of the specialized lymphoid tissues and organs, control of their function is very important, for escape of chronic inflammatory processes. Realistically, this control proved the role of small ions and molecules, by direct and/or indirect influence on different intra-and extra-cellular inter-molecular interactions in various cascade regulatory pathways.