title: The microenvironment of AFT024 cells maintains primitive human hematopoiesis by counteracting contact mediated inhibition of proliferation.
abstract: We have previously shown that maintenance of primitive human hematopoietic stem cells is poor when cultured in contact with marrow stromal feeders. However, when separated from stromal contact, human progenitors can be maintained because adhesion mediated proliferation inhibition does not occur. In this study we demonstrate how the murine fetal liver cell line, AFT024, supports primitive human hematopoiesis better in contact cultures compared to primary feeders. We evaluated if better progenitor maintenance in contact with AFT024 cells can be explained by decreased adhesion itself or decreased adhesion mediated inhibition of proliferation. We show that primitive human hematopoietic cells adhered equally well to AFT024 and primary feeders, such as M2-10B4. Further, contact with metabolically inactive AFT024 cells prevented cell cycle progression and decreased maintenance of primitive progenitors to the same extent as contact with M2-10B4 feeders. However, contact with viable AFT024 feeders did not inhibit proliferation, suggesting that AFT024-factors counteract contact mediated inhibition of proliferation. Cytokine production by M2-10B4 and AFT024 cells was similar. Large-size O-sulfated heparan sulfate glycosaminoglycans, known to be important for hematopoietic support, were found only in AFT024-matrix. We hypothesize that these factors may explain, in part, our observations. Finally, we show that more than 100% of primitive myeloid progenitors could be maintained for at least five weeks when cultured in contact with AFT024 feeders in the presence of Interleukin-3 and Macrophage Inflammatory Protein-1alpha. In conclusion, AFT024 cells produce factor(s), that counteract contact induced growth inhibition of primitive human hematopoietic progenitors, leading to expansion of these cells in contact with the microenvironment.
authors.affiliation: Department of Medicine, Division of Hematology and Stem Cell Institute, University of Minnesota, Minneapolis, MN, USA. michael_punzel@med.uni-heidelberg.de
authors.forename: Michael
authors.initials: M
authors.lastname: Punzel
authors.forename: Pankaj
authors.initials: P
authors.lastname: Gupta
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Verfaillie
mesh_terms: D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D018929:Cell Culture Techniques; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D054407:Chemokine CCL4; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D006410:Hematopoiesis / Q000502:physiology*; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000502:physiology; D006497:Heparan Sulfate / Q000494:pharmacology; D006801:Humans; D008099:Liver / Q000166:cytology*; D019402:Macrophage Inflammatory Proteins / Q000378:metabolism; D051379:Mice; D011509:Proteoglycans / Q000494:pharmacology; D017154:Stromal Cells
chemical_list: D054407:Chemokine CCL4; D016207:Cytokines; D006497:Heparan Sulfate; D019402:Macrophage Inflammatory Proteins; D011509:Proteoglycans