Glucagon-like peptide-1 receptor expression after myocardial infarction: Imaging study using 68Ga-NODAGA-exendin-4 positron emission tomography

Figures

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  Figure 1. Histology of rat myocardium 1 week after coronary ligation or sham-operation. A Masson’s trichrome staining showing myocytes as red color in the left ventricle myocardium of a sham-operated rat, whereas collagen fibers (blue) are detected in the infarcted area and in the remote, non-infarcted myocardium. There are numerous CD68-positive macrophages, α-SMA-positive and GLP-1R-positive cells in the infarcted area, and scattered α-SMA- and GLP-1R-positive interstitial cells in the remote myocardium (brown color). Scale bar 20 μm. B Bars show the areas of collagen (Masson’s trichrome staining), CD68-positive staining and α-SMA-positive staining. Values are mean ± SD; Student’s t-test for unpaired measurements and ANOVA for comparisons of MI 3 day, 1 week and 12 week; n = 6-9 per each bar. (α-SMA, alpha-smooth muscle actin; d, day; GLP-1R, glucagon-like peptide-1 receptor; MI, myocardial infarction; wk, week)

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  Figure 2. In vivo imaging of 68Ga-NODAGA-exendin-4 uptake. A Horizontal long axis positron emission tomography (PET) and corresponding contrast-enhanced computed tomography (CT) images showing focally increased 68Ga-NODAGA-exendin-4 uptake in the anterior wall of the left ventricle at 3 days and 1 week after myocardial infarction (MI) (red arrows) compared with remote myocardium (white arrows). After pre-injection of unlabeled exendin-4 peptide (MI 1 week block), uptake is as low as in the sham-operated rats. Scale; SUVmean (50-60 minutes), *scale from 0.3 to 3.7 at 12 weeks. B Bars show the mean ± SD standardized uptake values (SUVmean) 50 to 60 minutes after injection. Student’s t-test for unpaired and paired (MI vs remote) measurements; 3 d n = 3, Sham 1 wk n = 8, MI 1 wk n = 7, MI block n = 3, Sham 12 wk n = 6, MI 12 wk n = 8. (d, day; wk, week)

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  Figure 3. Kinetic analysis of 68Ga-NODAGA-exendin-4 uptake. A Horizontal long axis Ki-generated positron emission tomography and corresponding contrast-enhanced computed tomography image showing clear and focal 68Ga-NODAGA-exendin-4 uptake in the anterior wall of the left ventricle 1 week after myocardial infarction (MI) (red arrows) compared with remote myocardium (white arrows). B Representative Patlak plots of the MI and sham-operated rat. The slope of the linear portion represents the tracer influx rate constant (Ki), which refers to irreversible receptor binding and/or internalization of 68Ga-NODAGA-exendin-4. C Bars show the quantifications of Ki values. Values are mean ± SD; Student’s t-test for unpaired and paired (MI vs remote) measurements; 3 d n = 3, Sham 1 wk n = 7, MI 1 wk n = 7, MI block n = 3, Sham 12 wk n = 6, MI 12 wk n = 6. (d, day; wk, week)

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  Figure 4. Ex vivo myocardial uptake of 68Ga-NODAGA-exendin-4. A Autoradiographs and corresponding Masson’s trichrome stainings of the left ventricle cross sections (scale bar 1 mm) showing increased 68Ga-NODAGA-exendin-4 uptake in the infarcted area (arrows) at 3 days, 1 week and 12 weeks after myocardial infarction (MI), whereas uptake is low after sham-operation or injection of unlabeled exendin-4 (MI 1 wk block). Red arrows indicate border zone and arrowhead non-specific residual activity in the area of necrotic myocyte debris. B Quantification of 68Ga-NODAGA-exendin-4 uptake as photo-stimulated luminescence per square millimeter (PSL/mm2, mean ± SD) in infarcted area, border zone, remote myocardium and myocardium of sham-operated rats. Student’s t-test for unpaired and paired (MI vs remote) measurements; 3 d n = 6, Sham 1 wk n = 9, MI 1 wk n = 7, Sham 12 wk n = 9, MI 12 wk n = 9. (d, day; wk, week)

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  Figure 5. Localization of GLP-1R 1 week after coronary ligation. Double immunofluorescence staining for GLP-1R (green) and CD68-positive macrophages (red) showing complete co-localization (yellow, merge), whereas GLP-1R (green) do not co-localize (merge) with α-SMA-positive cells (red) in the infarcted area. Scale bar 10 μm. (α-SMA, alpha-smooth muscle actin; GLP-1R, glucagon-like peptide-1 receptor)

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  Figure 6. Histological correlations of 68Ga-NODAGA-exendin-4 uptake. Scatter plots showing positive correlation between CD68 staining and 68Ga-NODAGA-exendin-4 uptake in the infarcted area both by in vivo PET and autoradiography, whereas there is a negative correlation with α-SMA staining in the infarcted area. In the remote myocardium, 68Ga-NODAGA-exendin-4 uptake correlates positively with α-SMA staining. r = Spearman’s rank correlation coefficient. (α-SMA, alpha-smooth muscle actin; d, day; MI, myocardial infarction; PSL/mm2, photo-stimulated luminescence per square millimeter; wk, week)