Increased perivascular fibrosis and pro-fibrotic cellular transition in intramyocardial blood vessels in myocardial infarction patients

Figures

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  Fig. 1. Intramyocardial perivascular fibrosis in MI patients. Shown are examples of histological EVG staining of larger branches of the intramyocardial vasculature in the LV of control patients (A) and acute-phase MI patients (B), and a higher magnification of intramyocardial blood vessels in an acute-phase MI patient (C). The asterisks (*) indicate blood vessels. The black arrows indicate pink-stained perivascular collagen fibres, and the black double arrows indicate spherical or ovoid-shaped transparent perivascular adipose tissue. Quantification of perivascular fibrosis in controls (n = 14), acute-phase MI (n = 24) and subacute-phase MI patients (n = 12) (D). Data are presented as mean [SD], each distinct point corresponds to the scores of individual patient: circles for controls, squares for acute-phase MI patients, triangles for the border zones and inverted triangles for the infarct cores of subacute-phase MI patients. The data followed a Gaussian distribution, one-way ANOVA was employed to assess the differences among the groups. A paired samples t-test was conducted to compare the data between the infarct core and border zone in subacute-phase MI. Error bars represent SD. Levels…

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  Fig. 2. FAP expression in the myocardium of MI patients. Shown are examples of immunohistochemical FAP staining in the myocardium of acute-phase (A) and sub-acute-phase (B) MI patients. Asterisks (*) indicate blood vessels; black arrows indicate perivascular FAP staining and red arrow-heads indicate FAP staining in the granulation tissue; IC and BZ indicate infarct cores and border zones respectively; Quantification of FAP expression in acute-phase and subacute-phase MI patients. Data are presented as median [IQR] in the graph, each distinct point corresponds to the scores of individual patient: controls (n = 14; circles), acute-phase MI patients (n = 24; squares) and subacute-phase MI patients (n = 12; triangles for border zones, inverted triangles for infarct cores). A Kruskal-Wallis H test was used for statistical comparisons, due to the non-Gaussian distribution; a Wilcoxon matched-paired signed rank test was conducted to evaluate differences between the infarct core and border zone in subacute-phase MI. Error bars represent IQR. Levels of significance were indicated as *p ≤ 0.05, ***p ≤ 0.001, ****p ≤ 0.0001. (For interpretation of the references to colour in this figure legend, the…

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  Fig. 3. FAP expression in intramyocardial vascular cells of MI patients. Shown are examples of immunohistochemical staining for FAP, highlighting its expression in vascular cells in acute-phase (A and B) and sub-acute-phase (C) MI patients. Asterisks (*) indicate blood vessels; black arrows indicate FAP-positive endothelial cells (A and C); black double arrows indicate vascular smooth muscle cells (B and C). Panel (D) displays an example of immunofluorescent staining for CD31 (purple), SMA (red) and FAP (green) in the myocardium of an acute-phase MI patient. Nuclei were stained with Hoechst (blue). Colocalization of CD31 + FAP (white) in the endothelial layer is indicated in the merged image (white arrow), while colocalization of SMA + FAP (yellow) in the smooth muscle layer is shown (white double arrow). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

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  Fig. 4. Pro-fibrotic cellular transition in intramyocardial blood vessels in MI patients. (A) Example of immunofluorescent staining for SMA (red), S100A4 (green) in the myocardium of an acute-phase MI patient. Nuclei were stained with Hoechst (blue). Colocalisation of SMA + S100A4 in the smooth muscle layer, indicating the VSMC-to-mesenchymal transition, is shown in the merged image (yellow). (B) Examples of immunofluorescent staining for CD31 (magenta), S100A4 (green) in the myocardium of an acute-phase MI patient (top row: arterioles; bottom row: capillaries). Nuclei were stained with Hoechst (blue). Colocalisation of CD31 + S100A4 in the endothelial layer, indicating endothelial-to-mesenchymal transition, is shown in the merged image (white). (C) Larger magnified example of immunofluorescent staining for SMA (red), CD31 (magenta), S100A4 (green) in the myocardium of an acute-phase MI patient (top row). Nuclei were stained with Hoechst (blue). Colocalization of SMA + S100A4 (yellow) and CD31 + S100A4 (white) in the medial- and endothelial layers respectively is shown in bottom row. (For interpretation of the references to colour in this figure legend, the reader is referred to the web…

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  Fig. 5. Quantification of pro-fibrotic vascular cellular transition in MI patients. Quantification of fibroblast-like transitioning of SMA + S100A4+ vascular smooth muscle cells (A) and CD31 + S100A4+ endothelial cells (B) in controls (n = 14), acute-phase (n = 24) and subacute-phase MI (n = 12) patients. Data are presented as median [IQR] in the graph, each distinct point corresponds to the scores of individual patient: circles for controls, squares for acute-phase MI patients, triangles for the border zones and inverted triangles for the infarct cores of subacute-phase MI patients. A Kruskal-Wallis H test was used for statistical comparisons, due to the non-Gaussian distribution; a Wilcoxon matched-paired signed rank test was conducted to evaluate differences between the infarct core and border zone in subacute-phase MI. Error bars represent IQR. Levels of significance were indicated as *p ≤ 0.05, ***p ≤ 0.001, ****p ≤ 0.0001.

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  Fig. 6. Gender-specific quantification of intramyocardial perivascular fibrosis, FAP expression and pro-fibrotic vascular cellular transition in MI patients. Shown are the quantifications of intramyocardial perivascular fibrosis (A), FAP expression (B), fibroblast-like transitioning of SMA + S100A4+ vascular smooth muscle cells (C) and CD31 + S100A4+ endothelial cells (D) in male (M) versus female (F) controls (n = 14), acute-phase (n = 24) and subacute-phase MI (n = 12) patients. Data are presented as median [IQR] in the graph. Each distinct point corresponds to the scores of individual patient: circles for controls, squares for acute-phase MI patients, triangles for the border zones and inverted triangles for the infarct cores of subacute-phase MI patients. A Mann-Whitney U test was used for statistical comparisons; Error bars represent IQR, ns indicated no statistic difference.