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Mesenchymal-endothelial transition contributes to cardiac neovascularization.

Ubil E, Duan J, Pillai IC, Rosa-Garrido M, Wu Y, Bargiacchi F, Lu Y, Stanbouly S, Huang J, Rojas M, Vondriska TM, Stefani E, Deb A - Nature (2014)

Bottom Line: We show that the transcription factor p53 regulates such a switch in cardiac fibroblast fate.Loss of p53 in cardiac fibroblasts severely decreases the formation of fibroblast-derived endothelial cells, reduces post-infarct vascular density and worsens cardiac function.These observations demonstrate that mesenchymal-to-endothelial transition contributes to neovascularization of the injured heart and represents a potential therapeutic target for enhancing cardiac repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology &Physiology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599, USA.

ABSTRACT
Endothelial cells contribute to a subset of cardiac fibroblasts by undergoing endothelial-to-mesenchymal transition, but whether cardiac fibroblasts can adopt an endothelial cell fate and directly contribute to neovascularization after cardiac injury is not known. Here, using genetic fate map techniques, we demonstrate that cardiac fibroblasts rapidly adopt an endothelial-cell-like phenotype after acute ischaemic cardiac injury. Fibroblast-derived endothelial cells exhibit anatomical and functional characteristics of native endothelial cells. We show that the transcription factor p53 regulates such a switch in cardiac fibroblast fate. Loss of p53 in cardiac fibroblasts severely decreases the formation of fibroblast-derived endothelial cells, reduces post-infarct vascular density and worsens cardiac function. Conversely, stimulation of the p53 pathway in cardiac fibroblasts augments mesenchymal-to-endothelial transition, enhances vascularity and improves cardiac function. These observations demonstrate that mesenchymal-to-endothelial transition contributes to neovascularization of the injured heart and represents a potential therapeutic target for enhancing cardiac repair.

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RITA enhances MEndoT after cardiac injuryHearts from PBS or RITA treated Col1a2CreERT:R26RtdTomato mice (a,b) p53 immunostaining in labeled fibroblasts (arrowheads) and (c) quantitation of p53 expression (* p<0.005). (d–e) VECAD immunostaining in labeled fibroblasts (arrowheads). (f) Labeled fibroblasts(%) expressing VECAD (* p<0.05). (g) Endothelial cells/hpf (* p<0.05). (h) Masson’s Trichrome staining (blue, arrowheads) near (i) apex, (ii) between mid-ventricle and apex and (iii) mid-ventricle (n=14 animals) (i) Quantitation of fibrosis (* p<0.001, n=14 animals). (j,k) Cardiac function prior to and 7 days after injury. (* p<0.05, n=8 animals). (n=4 animals unless mentioned, All graphs show mean ± S.E.M., scale bar: 10μm).
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Figure 4: RITA enhances MEndoT after cardiac injuryHearts from PBS or RITA treated Col1a2CreERT:R26RtdTomato mice (a,b) p53 immunostaining in labeled fibroblasts (arrowheads) and (c) quantitation of p53 expression (* p<0.005). (d–e) VECAD immunostaining in labeled fibroblasts (arrowheads). (f) Labeled fibroblasts(%) expressing VECAD (* p<0.05). (g) Endothelial cells/hpf (* p<0.05). (h) Masson’s Trichrome staining (blue, arrowheads) near (i) apex, (ii) between mid-ventricle and apex and (iii) mid-ventricle (n=14 animals) (i) Quantitation of fibrosis (* p<0.001, n=14 animals). (j,k) Cardiac function prior to and 7 days after injury. (* p<0.05, n=8 animals). (n=4 animals unless mentioned, All graphs show mean ± S.E.M., scale bar: 10μm).

Mentions: Endothelial cells are known to undergo endothelial-mesenchymal-transition (EndMT) and adopt a fibroblast phenotype after cardiac injury5. We investigated whether endothelial cells undergoing EndMT would exhibit tdTomato fluorescence and be mistakenly included in our analysis. We performed immunofluorescent staining for Col1 and observed that approximately 6±1% (mean±S.E.M.) of endothelial cells after injury expressed Col1 but none of these cells exhibited tdTomato fluorescence (Extended Data Fig. 3d). Conversely tdTomato labeled cells in the injury region that expressed VECAD did not stain for Col1 (Extended Data Fig. 3e). Next, we crossed Col1a2CreERT:R26RtdTomato mice with the Col1-GFP transgenic mice, which have GFP expression directly driven by the Col1a1 promoter. GFP expression thus serves as a useful real-time reporter of Col1 expression21,22. Progeny mice (Col1a2CreERT:R26RtdTomato:Col1-GFP) were administered tamoxifen and subjected to ischemic myocardial injury. At 3 days post-injury, tdTomato labeled cells that expressed VECAD did not exhibit GFP fluorescence (Extended Data Fig. 4a). Approximately 3±1% (mean±S.E.M.) of VECAD positive cells were GFP positive but with the exception of rare cells, VECAD+GFP+ cells did not exhibit tdTomato fluorescence (Extended Fig. 4b).


Mesenchymal-endothelial transition contributes to cardiac neovascularization.

Ubil E, Duan J, Pillai IC, Rosa-Garrido M, Wu Y, Bargiacchi F, Lu Y, Stanbouly S, Huang J, Rojas M, Vondriska TM, Stefani E, Deb A - Nature (2014)

RITA enhances MEndoT after cardiac injuryHearts from PBS or RITA treated Col1a2CreERT:R26RtdTomato mice (a,b) p53 immunostaining in labeled fibroblasts (arrowheads) and (c) quantitation of p53 expression (* p<0.005). (d–e) VECAD immunostaining in labeled fibroblasts (arrowheads). (f) Labeled fibroblasts(%) expressing VECAD (* p<0.05). (g) Endothelial cells/hpf (* p<0.05). (h) Masson’s Trichrome staining (blue, arrowheads) near (i) apex, (ii) between mid-ventricle and apex and (iii) mid-ventricle (n=14 animals) (i) Quantitation of fibrosis (* p<0.001, n=14 animals). (j,k) Cardiac function prior to and 7 days after injury. (* p<0.05, n=8 animals). (n=4 animals unless mentioned, All graphs show mean ± S.E.M., scale bar: 10μm).
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Figure 4: RITA enhances MEndoT after cardiac injuryHearts from PBS or RITA treated Col1a2CreERT:R26RtdTomato mice (a,b) p53 immunostaining in labeled fibroblasts (arrowheads) and (c) quantitation of p53 expression (* p<0.005). (d–e) VECAD immunostaining in labeled fibroblasts (arrowheads). (f) Labeled fibroblasts(%) expressing VECAD (* p<0.05). (g) Endothelial cells/hpf (* p<0.05). (h) Masson’s Trichrome staining (blue, arrowheads) near (i) apex, (ii) between mid-ventricle and apex and (iii) mid-ventricle (n=14 animals) (i) Quantitation of fibrosis (* p<0.001, n=14 animals). (j,k) Cardiac function prior to and 7 days after injury. (* p<0.05, n=8 animals). (n=4 animals unless mentioned, All graphs show mean ± S.E.M., scale bar: 10μm).
Mentions: Endothelial cells are known to undergo endothelial-mesenchymal-transition (EndMT) and adopt a fibroblast phenotype after cardiac injury5. We investigated whether endothelial cells undergoing EndMT would exhibit tdTomato fluorescence and be mistakenly included in our analysis. We performed immunofluorescent staining for Col1 and observed that approximately 6±1% (mean±S.E.M.) of endothelial cells after injury expressed Col1 but none of these cells exhibited tdTomato fluorescence (Extended Data Fig. 3d). Conversely tdTomato labeled cells in the injury region that expressed VECAD did not stain for Col1 (Extended Data Fig. 3e). Next, we crossed Col1a2CreERT:R26RtdTomato mice with the Col1-GFP transgenic mice, which have GFP expression directly driven by the Col1a1 promoter. GFP expression thus serves as a useful real-time reporter of Col1 expression21,22. Progeny mice (Col1a2CreERT:R26RtdTomato:Col1-GFP) were administered tamoxifen and subjected to ischemic myocardial injury. At 3 days post-injury, tdTomato labeled cells that expressed VECAD did not exhibit GFP fluorescence (Extended Data Fig. 4a). Approximately 3±1% (mean±S.E.M.) of VECAD positive cells were GFP positive but with the exception of rare cells, VECAD+GFP+ cells did not exhibit tdTomato fluorescence (Extended Fig. 4b).

Bottom Line: We show that the transcription factor p53 regulates such a switch in cardiac fibroblast fate.Loss of p53 in cardiac fibroblasts severely decreases the formation of fibroblast-derived endothelial cells, reduces post-infarct vascular density and worsens cardiac function.These observations demonstrate that mesenchymal-to-endothelial transition contributes to neovascularization of the injured heart and represents a potential therapeutic target for enhancing cardiac repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology &Physiology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599, USA.

ABSTRACT
Endothelial cells contribute to a subset of cardiac fibroblasts by undergoing endothelial-to-mesenchymal transition, but whether cardiac fibroblasts can adopt an endothelial cell fate and directly contribute to neovascularization after cardiac injury is not known. Here, using genetic fate map techniques, we demonstrate that cardiac fibroblasts rapidly adopt an endothelial-cell-like phenotype after acute ischaemic cardiac injury. Fibroblast-derived endothelial cells exhibit anatomical and functional characteristics of native endothelial cells. We show that the transcription factor p53 regulates such a switch in cardiac fibroblast fate. Loss of p53 in cardiac fibroblasts severely decreases the formation of fibroblast-derived endothelial cells, reduces post-infarct vascular density and worsens cardiac function. Conversely, stimulation of the p53 pathway in cardiac fibroblasts augments mesenchymal-to-endothelial transition, enhances vascularity and improves cardiac function. These observations demonstrate that mesenchymal-to-endothelial transition contributes to neovascularization of the injured heart and represents a potential therapeutic target for enhancing cardiac repair.

Show MeSH
Related in: MedlinePlus