Sustained in vitro trigger of self-renewal divisions in Hoxb4hiPbx1(10) hematopoietic stem cells.
Factors that trigger and sustain self-renewal divisions in tissue stem cells remain poorly characterized. By modulating the levels of Hoxb4 and its co-factor Pbxl in primary hematopoietic cells (Hoxb4hiPbxl(10) cells), we report an in vitro expansion of mouse hematopoietic stem cells (HSCs) by 105-fold over 2 weeks, with subsequent preservation of HSC properties. Clonal analyses of the hematopoietic system in recipients of expanded HSCs indicate that up to 70% of Hoxb4hiPbxl(10) stem cells present at initiation of culture underwent self-renewal in vitro. In this setting, Hoxb4 and its co-factor did not promote an increase in DNA synthesis, or a decrease in doubling time of Scal+Lin- cells when compared to controls. Q-PCR analyses further revealed a downregulation of Cdknlb (p27Kipl) and Mxdl (MadI) transcript levels in Hoxb4hiPbxl(l0) primitive cells, accompanied by a more subtle increase in c-myc and reduction in Ccnd3 (Cyclin D3). We thus put forward this strategy as an efficient in vitro HSC expansion tool, enabling a further step into the avenue of self-renewal molecular effectors.
Cellot
Sonia
S
Laboratory of Molecular Genetics of Stem Cells, Institute for Research in Immunology and Cancer (IRIC), Montreal, QC, Canada.
Krosl
Jana
J
Chagraoui
Jalila
J
Meloche
Sylvain
S
Humphries
R Keith
RK
Sauvageau
Guy
G
D000818:Animals; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D056743:Cyclin D3; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics; D016213:Cyclins / Q000235:genetics; D015536:Down-Regulation / Q000235:genetics; D020869:Gene Expression Profiling; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000502:physiology*; D018398:Homeodomain Proteins / Q000502:physiology*; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D000075062:Pre-B-Cell Leukemia Transcription Factor 1; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000235:genetics
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; C534697:Ccnd3 protein, mouse; C493961:Cdkn1b protein, mouse; D056743:Cyclin D3; D016213:Cyclins; D018398:Homeodomain Proteins; C513262:Hoxb4 protein, mouse; C499479:Mad protein, mouse; C481797:Pbx1 protein, mouse; D000075062:Pre-B-Cell Leukemia Transcription Factor 1; D016271:Proto-Oncogene Proteins c-myc; D012097:Repressor Proteins; D014157:Transcription Factors; D050760:Cyclin-Dependent Kinase Inhibitor p27