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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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Histological analysis for capillary density in ischemic myocardium.KSL, KL, SL, CD34+ cells and PBS control were systemically injected 3 days after MI induction. Heart samples were harvested following BS1 lectin systemic perfusion 28 days after cell injection. a, Capillaries were visualized as tubular structure perfused by BS1 lectin. b, The numbers of capillaries were counted in bilateral sides of the peri-infarct zone on LV cross sections and averaged in each group (n = 3), respectively.
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pone-0020219-g008: Histological analysis for capillary density in ischemic myocardium.KSL, KL, SL, CD34+ cells and PBS control were systemically injected 3 days after MI induction. Heart samples were harvested following BS1 lectin systemic perfusion 28 days after cell injection. a, Capillaries were visualized as tubular structure perfused by BS1 lectin. b, The numbers of capillaries were counted in bilateral sides of the peri-infarct zone on LV cross sections and averaged in each group (n = 3), respectively.

Mentions: In order to evaluate the size of MI, Masson's trichrome staining was performed with heart sections at day 28. Representative images indicated that fibrosis area was stained in blue and intact myocardium was stained in red (Figure 7a). Quantitative analysis revealed that both the percent of fibrosis area in entire LV cross-sectional area and the percent of scar length in entire internal LV circumference were significantly reduced in the CD34+ cell-injected group compared with the KSL cell-injected group, the SL cell-injected group and PBS group (Figure 7b and 7c). The staining of in vivo-perfused BS1 lectin and α-smooth muscle actin reflects angiogenesis in functional vessels in the peri-infarct myocardium four weeks after MI in all groups (Figure 8a and 9a). The averaged capillary density in the bilateral ischemic border zones of LV, an index of neovascularization, was significantly greater in the CD34+ cell-injected group compared to the KSL cell-inject group and PBS group (CD34: 165±40/mm2, KL: 150±25/mm2, SL: 103±35/mm2, KSL: 80±21/mm2, PBS: 70±13/mm2; P<0.05, PBS vs CD34 and KL, KSL vs CD34 and KL) (Figure 8b). Although the averaged number of arterioles was higher in the CD34+ cell-injected group, no significant difference could be found between any two groups (Figure 9b). The KL cell-injected group also showed an increase in capillary density (P<0.05, KL vs KSL and PBS), reduced fibrosis area (P<0.05, KL vs KSL and PBS) and reduced fibrosis length (P<0.05, KL vs KSL, SL and PBS).


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)

Histological analysis for capillary density in ischemic myocardium.KSL, KL, SL, CD34+ cells and PBS control were systemically injected 3 days after MI induction. Heart samples were harvested following BS1 lectin systemic perfusion 28 days after cell injection. a, Capillaries were visualized as tubular structure perfused by BS1 lectin. b, The numbers of capillaries were counted in bilateral sides of the peri-infarct zone on LV cross sections and averaged in each group (n = 3), respectively.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0020219-g008: Histological analysis for capillary density in ischemic myocardium.KSL, KL, SL, CD34+ cells and PBS control were systemically injected 3 days after MI induction. Heart samples were harvested following BS1 lectin systemic perfusion 28 days after cell injection. a, Capillaries were visualized as tubular structure perfused by BS1 lectin. b, The numbers of capillaries were counted in bilateral sides of the peri-infarct zone on LV cross sections and averaged in each group (n = 3), respectively.
Mentions: In order to evaluate the size of MI, Masson's trichrome staining was performed with heart sections at day 28. Representative images indicated that fibrosis area was stained in blue and intact myocardium was stained in red (Figure 7a). Quantitative analysis revealed that both the percent of fibrosis area in entire LV cross-sectional area and the percent of scar length in entire internal LV circumference were significantly reduced in the CD34+ cell-injected group compared with the KSL cell-injected group, the SL cell-injected group and PBS group (Figure 7b and 7c). The staining of in vivo-perfused BS1 lectin and α-smooth muscle actin reflects angiogenesis in functional vessels in the peri-infarct myocardium four weeks after MI in all groups (Figure 8a and 9a). The averaged capillary density in the bilateral ischemic border zones of LV, an index of neovascularization, was significantly greater in the CD34+ cell-injected group compared to the KSL cell-inject group and PBS group (CD34: 165±40/mm2, KL: 150±25/mm2, SL: 103±35/mm2, KSL: 80±21/mm2, PBS: 70±13/mm2; P<0.05, PBS vs CD34 and KL, KSL vs CD34 and KL) (Figure 8b). Although the averaged number of arterioles was higher in the CD34+ cell-injected group, no significant difference could be found between any two groups (Figure 9b). The KL cell-injected group also showed an increase in capillary density (P<0.05, KL vs KSL and PBS), reduced fibrosis area (P<0.05, KL vs KSL and PBS) and reduced fibrosis length (P<0.05, KL vs KSL, SL and PBS).

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