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CD34+ cells represent highly functional endothelial progenitor cells in murine bone marrow.

Yang J, Ii M, Kamei N, Alev C, Kwon SM, Kawamoto A, Akimaru H, Masuda H, Sawa Y, Asahara T - PLoS ONE (2011)

Bottom Line: Although CD34(+) cells showed the lowest EPC colony forming activity, CD34(+) cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1.Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34(+) cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34(+) cells.Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34(+) cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others.

View Article: PubMed Central - PubMed

Affiliation: Group of Vascular Regeneration Research, Institute of Biomedical Research and Innovation/RIKEN Center for Developmental Biology, Kobe, Japan.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) were shown to have angiogenic potential contributing to neovascularization. However, a clear definition of mouse EPCs by cell surface markers still remains elusive. We hypothesized that CD34 could be used for identification and isolation of functional EPCs from mouse bone marrow.

Methodology/principal findings: CD34(+) cells, c-Kit(+)/Sca-1(+)/Lin(-) (KSL) cells, c-Kit(+)/Lin(-) (KL) cells and Sca-1(+)/Lin(-) (SL) cells were isolated from mouse bone marrow mononuclear cells (BMMNCs) using fluorescent activated cell sorting. EPC colony forming capacity and differentiation capacity into endothelial lineage were examined in the cells. Although CD34(+) cells showed the lowest EPC colony forming activity, CD34(+) cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1. Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34(+) cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34(+) cells. Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34(+) cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others.

Conclusion: These findings suggest that mouse CD34(+) cells may represent a functional EPC population in bone marrow, which could benefit the investigation of therapeutic EPC biology.

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Stem/progenitor differentiation hierarchy of KSL, KL, SL and CD34+ cells.In EPC-CFA, small EPC colonies are capable of differentiating into large colonies and large EPC colonies can further differentiate into adherent, more vasculogenic non-colony forming EPCs in culture. Small colonies reveal a predominant potential for proliferation, while large colonies demonstrate a predominantly vasculogenic potential including cell adhesion and tube-like structure formation in vitro (unpublished data, lower panel). Based on the EPC colony numbers of KSL, KL, SL and CD34+ cells, the conceivable extent of homing capacity (Y-axis) and differentiation status (X-axis) in each cell type is indicated in the upper panel.
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pone-0020219-g011: Stem/progenitor differentiation hierarchy of KSL, KL, SL and CD34+ cells.In EPC-CFA, small EPC colonies are capable of differentiating into large colonies and large EPC colonies can further differentiate into adherent, more vasculogenic non-colony forming EPCs in culture. Small colonies reveal a predominant potential for proliferation, while large colonies demonstrate a predominantly vasculogenic potential including cell adhesion and tube-like structure formation in vitro (unpublished data, lower panel). Based on the EPC colony numbers of KSL, KL, SL and CD34+ cells, the conceivable extent of homing capacity (Y-axis) and differentiation status (X-axis) in each cell type is indicated in the upper panel.

Mentions: In terms of EPC colony forming activity, KSL cells formed a substantially greater number of small (primitive) EPC colonies and large (definitive) EPC colonies, especially small EPC colony forming activity of KSL cells was robust compared to the others, which is consistent with our previous data [16]. Both KL cells and SL cells showed a less level of small/large colony forming activity, while CD34+ cells demonstrated a further low activity. In the cell culture assay and in vivo assays, CD34+ cells had the highest activity. It seems to be controversial that CD34+ cells are the most functional cells, but they can only form a few colonies. In fact, it is not surprising and very reasonable. Because CD34+ cells count 12.23±0.02% in BMMNCs, while KL cells count 3.737±0.004%, SL cells count 1.327±0.001%, KSL count 0.597±0.001%, respectively, which indicates CD34+ cells are partially differentiated cells. In EPC differentiation hierarchy, it was confirmed that small EPC colonies are capable of differentiation into large EPC colonies and that large colonies can further differentiate into adherent, more vasculogenic EPCs in culture (unpublished data by Masuda et al) (Figure 11). In addition, small EPC colonies have a predominant stem cell-like potential for proliferation and large EPC colonies possess a predominantly vasculogenic potential including cell adhesion, tube formation activity in vitro and contribution to de novo blood vessel formation in ischemic tissue (Kwon et al [16], unpublished data by Masuda et al and Tsukada et al and Figure S2) (Figure 11). Taken the EPC differentiation and in vivo data together, our results suggest that KSL cells are most immature, while CD34+ cells are most differentiating population in mouse EPCs that we studied (Figure 11). Therefore we hypothesize the possible differentiation hierarchy of KSL, KL, SL and CD34+ cells (Figure 11).


CD34+ cells represent highly functional endothelial progenitor cells in murine bone marrow.

Yang J, Ii M, Kamei N, Alev C, Kwon SM, Kawamoto A, Akimaru H, Masuda H, Sawa Y, Asahara T - PLoS ONE (2011)

Stem/progenitor differentiation hierarchy of KSL, KL, SL and CD34+ cells.In EPC-CFA, small EPC colonies are capable of differentiating into large colonies and large EPC colonies can further differentiate into adherent, more vasculogenic non-colony forming EPCs in culture. Small colonies reveal a predominant potential for proliferation, while large colonies demonstrate a predominantly vasculogenic potential including cell adhesion and tube-like structure formation in vitro (unpublished data, lower panel). Based on the EPC colony numbers of KSL, KL, SL and CD34+ cells, the conceivable extent of homing capacity (Y-axis) and differentiation status (X-axis) in each cell type is indicated in the upper panel.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3105013&req=5

pone-0020219-g011: Stem/progenitor differentiation hierarchy of KSL, KL, SL and CD34+ cells.In EPC-CFA, small EPC colonies are capable of differentiating into large colonies and large EPC colonies can further differentiate into adherent, more vasculogenic non-colony forming EPCs in culture. Small colonies reveal a predominant potential for proliferation, while large colonies demonstrate a predominantly vasculogenic potential including cell adhesion and tube-like structure formation in vitro (unpublished data, lower panel). Based on the EPC colony numbers of KSL, KL, SL and CD34+ cells, the conceivable extent of homing capacity (Y-axis) and differentiation status (X-axis) in each cell type is indicated in the upper panel.
Mentions: In terms of EPC colony forming activity, KSL cells formed a substantially greater number of small (primitive) EPC colonies and large (definitive) EPC colonies, especially small EPC colony forming activity of KSL cells was robust compared to the others, which is consistent with our previous data [16]. Both KL cells and SL cells showed a less level of small/large colony forming activity, while CD34+ cells demonstrated a further low activity. In the cell culture assay and in vivo assays, CD34+ cells had the highest activity. It seems to be controversial that CD34+ cells are the most functional cells, but they can only form a few colonies. In fact, it is not surprising and very reasonable. Because CD34+ cells count 12.23±0.02% in BMMNCs, while KL cells count 3.737±0.004%, SL cells count 1.327±0.001%, KSL count 0.597±0.001%, respectively, which indicates CD34+ cells are partially differentiated cells. In EPC differentiation hierarchy, it was confirmed that small EPC colonies are capable of differentiation into large EPC colonies and that large colonies can further differentiate into adherent, more vasculogenic EPCs in culture (unpublished data by Masuda et al) (Figure 11). In addition, small EPC colonies have a predominant stem cell-like potential for proliferation and large EPC colonies possess a predominantly vasculogenic potential including cell adhesion, tube formation activity in vitro and contribution to de novo blood vessel formation in ischemic tissue (Kwon et al [16], unpublished data by Masuda et al and Tsukada et al and Figure S2) (Figure 11). Taken the EPC differentiation and in vivo data together, our results suggest that KSL cells are most immature, while CD34+ cells are most differentiating population in mouse EPCs that we studied (Figure 11). Therefore we hypothesize the possible differentiation hierarchy of KSL, KL, SL and CD34+ cells (Figure 11).

Bottom Line: Although CD34(+) cells showed the lowest EPC colony forming activity, CD34(+) cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1.Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34(+) cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34(+) cells.Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34(+) cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others.

View Article: PubMed Central - PubMed

Affiliation: Group of Vascular Regeneration Research, Institute of Biomedical Research and Innovation/RIKEN Center for Developmental Biology, Kobe, Japan.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) were shown to have angiogenic potential contributing to neovascularization. However, a clear definition of mouse EPCs by cell surface markers still remains elusive. We hypothesized that CD34 could be used for identification and isolation of functional EPCs from mouse bone marrow.

Methodology/principal findings: CD34(+) cells, c-Kit(+)/Sca-1(+)/Lin(-) (KSL) cells, c-Kit(+)/Lin(-) (KL) cells and Sca-1(+)/Lin(-) (SL) cells were isolated from mouse bone marrow mononuclear cells (BMMNCs) using fluorescent activated cell sorting. EPC colony forming capacity and differentiation capacity into endothelial lineage were examined in the cells. Although CD34(+) cells showed the lowest EPC colony forming activity, CD34(+) cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1. Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34(+) cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34(+) cells. Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34(+) cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others.

Conclusion: These findings suggest that mouse CD34(+) cells may represent a functional EPC population in bone marrow, which could benefit the investigation of therapeutic EPC biology.

Show MeSH
Related in: MedlinePlus