Figure 2

Hoechst increases IGF-1 localization to infarcted myocardium.

(A) Rats were subjected to ischemia-reperfusion (IR) surgery. One day later Hoechst-biotin was injected and allowed to circulate for 1 h. Frozen sections were made and tissue was examined via red autofluorescence of the myocardium. Sham operated animals showed cell membrane integrity in the left ventricle (LV) and little blue fluorescence. IR-operated animals showed necrosis in the LV and diffuse blue staining suggesting release of DNA (Scale bar = 200 μm). (B) Imaging of hearts harvested from mice at the indicated time-point. Hoechst increased IGF-1 accumulation at 18 hours compared to S-IGF1 (n = 4–7 per group; * p < 0.05 vs. S-IGF1, ** p < 0.01 vs. SIGF1, two-way ANOVA followed by Bonferroni post-test). (C) Imaging was used to quantify IGF-1 as %ID per gram of tissue at 18 h. While other organs were similar, there was a significant increase in the heart of H-IGF1 treated mice compared to S-IGF1 treatment (n ≥ 5 per group, *p < 0.05; t-test). (D) Western blots showing phosphorylation of Akt in the LV of mice treated with S-IGF1 or H-IGF1. By 18 hours, Akt phosphorylation in H-IGF1 treated mice was significantly greater than S-IGF1 treated mice. Line represents PBS treated IR mice. (* p < 0.05 vs. S-IGF1 at 18 hours, n = 3–7 per group, two-way ANOVA followed by Bonferroni post-test). (E) Mice were treated with S-IGF1, H-IGF1, or H-IGF1 pre-incubated with 1000× excess of double-stranded DNA. Pre-incubation with DNA significantly decreased IGF-1 levels. (*p < 0.05 vs. S-IGF1, # p < 0.05 vs. H-IGF1, n = 5–7 per group, one-way ANOVA followed by Bonferroni post-test).