we have investigated molecular and functional properties in early B-lineage cells from Pax-5 deficient animals crossed to a B-lineage restricted reporter mouse. Gene expression analysis of ex vivo isolated progenitor cells revealed that Pax-5 deficiency has a minor impact on Bcell specification.By comparison of gene expression patterns in ex vivo isolated Pax-5 and Ebf-1 deficient progenitors, it was possible to identify a set of B-cell restricted genes dependent of Ebf-1 but not Pax-5, supporting the idea that B-cell specification and commitment is controlled by distinct regulatory networks.
Single-cell analysis of early B-lymphocyte development suggests independent regulation of lineage specification and commitment in vivo.
Specimen part
View SamplesLoss of one allele of Ebf1 impairs pre-B cell (B220+CD19+CD43low/negIgM-) expansion. In order to better understand the underlying cause of the reduced pre-B cell compartment in Ebf1+/- mice, we sorted pro-B (B220+CD19+CD43highIgM- ) as well as pre-B cells from Wt and Ebf1 heterozygote mutant mice and performed Affymetrix based microarray gene expression analysis.
Early B-cell factor 1 regulates the expansion of B-cell progenitors in a dose-dependent manner.
Specimen part
View SamplesTo identify the true molecular features of the Ebf2+ cells, we performed microarray analysis of freshly sorted CD45-TER119-Ebf2+ and Ebf2- cells. This allowed for the detection of 1968 genes that were 2-fold differentially expressed in Ebf2+ and Ebf2- cells. Among these, 1075 genes were upregulated and 893 genes including Ebf2, were downregulated in the Ebf2- as compared to the Ebf2+ cells. These include Nov, Fmod, Ndn, Dcn, Ctgf, Angiopoietin like-1(Angptl1), Fn1 and Jag1, some of which has been reported to be expressed in culture-selected MSCs. Furthermore, consistent with antigen expression analysis by FACS, the Ebf2+ cells highly expressed transcripts of Pdgfra, Pdgfrb, Sca1/Ly6a, Thy1 and Itga7 and Itgav, that have been suggested to be linked to MSCs. Nestin was mainly expressed in the Ebf2+ cells whereas it was hardly detectable in the Ebf2- cells. Altogether, molecularly, the Ebf2+ cells displayed features of a MSC.
Molecular characterization of prospectively isolated multipotent mesenchymal progenitors provides new insight into the cellular identity of mesenchymal stem cells in mouse bone marrow.
Specimen part
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