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. 2018 Oct 11;7(20):e010025. doi: 10.1161/JAHA.118.010025

Figure 6.

Figure 6

SWT stimulates cardiac vasculogenesis in chronic ischemic heart failure by recruitment of bone marrow–derived endothelial cells. A, GFP bone marrow transplantation model for evaluation of myocardial vasculogenesis. WT animals underwent sublethal irradiation and received a bone marrow transplantation from GFP donor mice. We performed LAD ligation and after 3 weeks cardial SWT. Four weeks later, hearts were harvested and analyzed for GFP‐positive endothelial cells (representing bone marrow–derived endothelial cells). B, SWT enhances recruitment and homing of bone marrow–derived endothelial cells to ischemic myocardium. Quantification of cells double‐positive for rhodamine‐labeled isolectin (red, endothelial cell marker) and for GFP (green, bone marrow–derived cells) revealed significantly higher numbers of BMECs in the treatment group compared with untreated controls. (n=6 per group, ****P<0.0001 vs CTR). C and D, SDF‐1 upregulation after SWT. Animals were euthanized 72 h after SWT. Hearts were analyzed via Western blot, and SDF‐1 serum levels were measured using an ELISA. We found increased myocardial SDF‐1 protein levels with concomitant increase of SDF‐1 serum levels. (n=6 per group, *P<0.05 vs CTR). E, SWT causes increased CXCR4 expression. Myocardial mRNA levels of the specific SDF‐1 receptor CXCR4 were upregulated 72 h after SWT indicating involvement of the SDF‐1‐CXCR4 axis in SW‐induced vasculogenesis (n=6 per group, *P<0.05 vs CTR). F, SWT triggers release of chemoattractants. HUVECs were treated with SWT and supernatant analyzed via a cytokine profiler for amount of chemoattractant proteins. Treatment resulted in increased levels of IL‐6, IL‐8, MCP‐1, and MIF, all potent chemoattractants for BMEC recruitment. All experiments were performed at least in triplicate. G, Enhanced endothelial cell migration after SWT. Culture medium of ischemic cardiomyocytes treated with SWT‐induced chemotaxis of endothelial cells in a modified Boyden chamber migration assay. (green: WGA, blue: DAPI; scale bar=100 μm). (*P<0.05 vs CTR). All experiments were performed at least in triplicate. BMEC indicates bone marrow–derived endothelial cell; CTR, control; CXCR4, C‐X‐C chemokine receptor type 4; DAPI, 4′,diamidino‐2‐phenylindole; GFP, green fluorescent protein; HUVEC, human umbilical vein endothelial cell; IL‐6, interleukin 6; IL‐8, interleukin 8; Isch‐CM, ischemic cardiomyocytes; LAD, left anterior descending; MCP‐1, monocyte chemoattractant protein 1; MIF, macrophage migration inhibitory factor; SDF‐1, stromal cell‐derived factor; SWT, shock wave therapy; WGA, wheat germ agglutinin.