Figure 5

Pretreatment of mouse cardiac mesoangioblasts increases myocardium regeneration. (a) Infarcted hearts, injected with cardiac mesoangioblasts pretreated with SDF-1 and transfected with the L-selectin-EGFP vector or with control cardiac mesoangioblasts, were collected after 6 weeks and subjected to real-time PCR for the presence of GFP. A mean of seven independent experiments run in triplicate is shown (^*α<0.02). (b) Infarcted hearts, injected with cardiac mesoangioblasts pretreated with SDF-1 and transfected with the L-selectin-EGFP vector or with control cardiac mesoangioblasts, were collected after 6 weeks and sorted for the GFP population after digestion. A representative plot of four independent experiments is shown. (c) Immunostaining of the free wall collected 6 weeks after injection with control cardiac mesoangioblasts or with cardiac mesoangioblasts pretreated with SDF-1 and transfected with the L-selectin-EGFP vector. Laminin is shown in red. Nuclei are stained with Hoechst (blue). Magnification: × 450. Scale bar, 100 μm. (d) Immunofluorescence for GFP and sarcomeric actin (red) from a heart region of CAL mice 6 weeks after injection with cardiac mesoangioblasts pretreated with SDF-1 and transfected with L-selectin-EGFP. Scale bar, 100 μm. (e) Higher magnification immunofluorescence for GFP and sarcomeric actin (red) from the hearts of CAL mice 6 weeks after injection with cardiac mesoangioblasts pretreated with SDF-1 and transfected with L-selectin-EGFP. Nuclei are stained with Hoechst (blue). Scale bar, 20 μm. (f) Western blot for GFP and sarcomeric actin of total GFP-sorted fractions recovered from the hearts of CAL mice 6 weeks after injection with control GFP-cardiac mesoangioblasts or with cardiac mesoangioblasts pretreated with SDF-1 and transfected with the L-selectin-EGFP vector. One representative out of five independent experiments is shown. (g) Real-time PCR for late cardiac differentiation genes on mature cardiomyocytes and GFP-sorted fraction coming from the hearts of CAL mice 6 weeks after injection with cardiac mesoangioblasts, both control or pretreated with SDF-1 and transfected with the L-selectin-EGFP vector. Quantities have been normalized. A mean of four independent experiments run in triplicate is shown