Long-term maintenance of hematopoietic stem cells does not require contact with embryo-derived stromal cells in cocultures.
We recently established that two midgestation-derived stromal clones--UG26-1B6, urogenital ridge-derived, and EL08-1D2, embryonic liver-derived--support the maintenance of murine adult bone marrow and human cord blood hematopoietic repopulating stem cells (HSCs). In this study, we investigate whether direct HSC-stroma contact is required for this stem cell maintenance. Adult bone marrow ckit+ Ly-6C- side population (K6-SP) cells and stromal cells were cocultured under contact or noncontact conditions. These experiments showed that HSCs were maintained for at least 4 weeks in culture and that direct contact between HSCs and stromal cells was not required. To find out which factors might be involved in HSC maintenance, we compared the gene expression profile of EL08-1D2 and UG26-1B6 with four HSC-nonsupportive clones. We found that EL08-1D2 and UG26-1B6 both expressed 21 genes at a higher level, including the putative secreted factors fibroblast growth factor-7, insulin-like growth factor-binding proteins 3 and 4, pleiotrophin, pentaxin-related, and thrombospondin 2, whereas 11 genes, including GPX-3 and HSP27, were expressed at a lower level. In summary, we show for the first time long-term maintenance of adult bone marrow HSCs in stroma noncontact cultures and identify some secreted molecules that may be involved in this support.
Oostendorp
Robert A J
RA
Department of Cell Biology and Genetics, Erasmus University, rotterdam, Netherlands. oostendorp@lrz.tum.de
Robin
Catherine
C
Steinhoff
Christine
C
Marz
Stefanie
S
Bräuer
Rosalinde
R
Nuber
Ulrike A
UA
Dzierzak
Elaine A
EA
Peschel
Christian
C
D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D002097:C-Reactive Protein / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D002478:Cells, Cultured; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D004622:Embryo, Mammalian / Q000166:cytology*; D004797:Enzyme-Linked Immunosorbent Assay; D005312:Fetal Blood / Q000166:cytology; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000378:metabolism; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000196:embryology; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D018832:Molecular Chaperones; D009363:Neoplasm Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology; D017154:Stromal Cells / Q000166:cytology*; D019699:Thrombospondins / Q000378:metabolism; D013997:Time Factors; D000099153:Pentraxins; D000099262:Thrombospondin 2
D002097:C-Reactive Protein; D002352:Carrier Proteins; D016207:Cytokines; D017931:DNA Primers; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D005979:Glutathione Peroxidase; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018972:Insulin-Like Growth Factor Binding Protein 3; D018974:Insulin-Like Growth Factor Binding Protein 4; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D012313:RNA; D012333:RNA, Messenger; D019699:Thrombospondins; D000099153:Pentraxins; D000099262:Thrombospondin 2; C496453:FGF7 protein, human; C496454:Fgf7 protein, mouse; C057725:HSPB1 protein, human; C066845:pleiotrophin; C486484:GPX3 protein, human