Conclusions
We demonstrate that besides their unique immunophenotype and functional differences, BMC-derived HSPCs and BMMSCs have different molecular receptor signaling transcript profiles linked to cell survival, growth, cell differentiation status, growth factor/cytokine production and genes involved in cell migration/trafficking/adhesion that may be critical to maintain their pluripotency, plasticity, and stem cell function.
Methods
Murine HSPCs were enriched from fluorescence-activated cell sorting (FACS)-sorted Lin-c-Kit+Sca-1+ BMCs isolated from the tibia and femoral marrow compartments. Purified BMMSCs (CD73+, CD90+, CD105+, and CD45-, CD34-, CD31-, c-Kit-) with extensive self-renewal potential and multilineage differentiation capacity (into different mesodermal cell lineages including osteocytes, chrondrocytes, adipocytes) were derived from adherent BMC cultures after CD45+ cell depletion. Adherent colony-forming cells were passaged two to three times and FACS analysis was used to assess cell purity and validate cell-specific surface marker phenotype prior to experimentation. Gene transcripts for a number of cell signaling molecules were assessed using a custom quantitative real-time RT-PCR low-density microarray (94 genes; TaqMan® technology).
Results
We identified 16 mRNA transcripts that were specifically expressed in BMC-derived HSPC (including Ptprc, c-Kit, Csf3r, Csf2rb2, Ccr4, Cxcr3 and Tie-1), and 14 transcripts specifically expressed in BMMSCs (including Pdgfra, Ddr2, Ngfr, Mst1r, Fgfr2, Epha3, and Ephb3). We also identified 27 transcripts that were specifically upregulated (≥2-fold expression) in BMMSCs relative to HSPCs (Axl, Bmpr1a, Met, Pdgfrb, Fgfr1, Mertk, Cmkor1, Egfr, Epha7, and Ephb4), and 19 transcripts that were specifically upregulated in HSPCs relative to BMMSCs (Ccr1, Csf1r, Csf2ra, Epor, IL6ra, and IL7r). Eleven transcripts were equally expressed (<2-fold upregulation) in HSPCs and BMMSCs (Flt1, Insr, Kdr, Jak1, Agtrl1, Ccr3, Ednrb, Il3ra, Hoxb4, Tnfrsf1a, and Abcb1b), whilst another seven transcripts (Epha6, Epha8, Musk, Ntrk2, Ros1, Srms, and Tnk1) were not expressed in either cell population. Conclusions: We demonstrate that besides their unique immunophenotype and functional differences, BMC-derived HSPCs and BMMSCs have different molecular receptor signaling transcript profiles linked to cell survival, growth, cell differentiation status, growth factor/cytokine production and genes involved in cell migration/trafficking/adhesion that may be critical to maintain their pluripotency, plasticity, and stem cell function.