Figure 1
Figure 1. cAng-1 enhances vasculogenesis and SDF-1 expression in ischemic muscle. (A) Capillary density was significantly increased in the cAng-1 injection group (*P < .05, **P < .02) after ischemic surgery of C57BL mouse hind limb. Local administration of 100 μg of cAng-1 by 3 injections into thigh muscle (n = 7 each). (B) Immunoblotting for SDF-1 and α-tubulin using limb muscle protein of 3 days of ischemia demonstrated that cAng-1 significantly increased SDF-1 protein. (C) Murine peripheral blood samples were obtained with heparin tube through cardiac puncture at postoperative day 3. ELISA tests revealed a rise in the SDF-1 plasma levels after induction of ischemia, and showed a further elevation of SDF-1 in the cAng-1 injection group (n = 4 each). (D) Quantitative analysis for capillaries expressing SDF-1. Cells double positive for BS-1-lectin+/SDF-1+ were counted in 10 different microscopic fields of at least 3 different sections from each animal. BS-1-lectin+/SDF-1+ (yellow) numbers significantly increased in cAng-1 injection group (*P < .05, **P < .05). (E) Confocal microscopy for BS-1-lectin (green) and SDF-1 (red) in ischemic limb tissues at postoperative day 7. Capillary endothelial cells stained with BS-1 lectin are located around skeletal myocytes (phase bright) and are colocalized with SDF-1 immunofluorescence. No fluorescence signal in the IgG control group. Magnification ×100; scale bar, 50μM. At least 3 different sections from each animal (n = 7) were analyzed. (F) The 3-dimensional confocal microscopic image of fluorescence staining against transverse section shows conclusive colocalization of SDF-1 and BS-1 lectin. Magnification ×630 with immersion oil scale bar, 10 μM. (G) Immunohistochemical staining for SDF-1 and VE-cadherin in ischemic hindlimb tissues at postoperative day 7. Most SDF-1 staining corresponds to VE-cadherin staining in capillary endothelial cells (reddish brown; arrowheads). Unstained cells are skeletal myocytes. No signal in the IgG control group. Magnification ×200, ×400; (C) scale bar, 50 μM. Representative images are shown.

cAng-1 enhances vasculogenesis and SDF-1 expression in ischemic muscle. (A) Capillary density was significantly increased in the cAng-1 injection group (*P < .05, **P < .02) after ischemic surgery of C57BL mouse hind limb. Local administration of 100 μg of cAng-1 by 3 injections into thigh muscle (n = 7 each). (B) Immunoblotting for SDF-1 and α-tubulin using limb muscle protein of 3 days of ischemia demonstrated that cAng-1 significantly increased SDF-1 protein. (C) Murine peripheral blood samples were obtained with heparin tube through cardiac puncture at postoperative day 3. ELISA tests revealed a rise in the SDF-1 plasma levels after induction of ischemia, and showed a further elevation of SDF-1 in the cAng-1 injection group (n = 4 each). (D) Quantitative analysis for capillaries expressing SDF-1. Cells double positive for BS-1-lectin+/SDF-1+ were counted in 10 different microscopic fields of at least 3 different sections from each animal. BS-1-lectin+/SDF-1+ (yellow) numbers significantly increased in cAng-1 injection group (*P < .05, **P < .05). (E) Confocal microscopy for BS-1-lectin (green) and SDF-1 (red) in ischemic limb tissues at postoperative day 7. Capillary endothelial cells stained with BS-1 lectin are located around skeletal myocytes (phase bright) and are colocalized with SDF-1 immunofluorescence. No fluorescence signal in the IgG control group. Magnification ×100; scale bar, 50μM. At least 3 different sections from each animal (n = 7) were analyzed. (F) The 3-dimensional confocal microscopic image of fluorescence staining against transverse section shows conclusive colocalization of SDF-1 and BS-1 lectin. Magnification ×630 with immersion oil scale bar, 10 μM. (G) Immunohistochemical staining for SDF-1 and VE-cadherin in ischemic hindlimb tissues at postoperative day 7. Most SDF-1 staining corresponds to VE-cadherin staining in capillary endothelial cells (reddish brown; arrowheads). Unstained cells are skeletal myocytes. No signal in the IgG control group. Magnification ×200, ×400; (C) scale bar, 50 μM. Representative images are shown.

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