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miRNA-21 is dysregulated in response to vein grafting in multiple models and genetic ablation in mice attenuates neointima formation.

McDonald RA, White KM, Wu J, Cooley BC, Robertson KE, Halliday CA, McClure JD, Francis S, Lu R, Kennedy S, George SJ, Wan S, van Rooij E, Baker AH - Eur. Heart J. (2013)

Bottom Line: Identifying novel strategies to prevent neointimal thickening is important.Thus, this study aimed to identify microRNAs (miRNAs) that are dysregulated during neointimal formation and determine their pathophysiological relevance following miRNA manipulation.Furthermore, pharmacological knockdown of miR-21 in human veins resulted in target gene de-repression and a significant reduction in neointimal formation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow G12 8TA, UK.

ABSTRACT

Aims: The long-term failure of autologous saphenous vein bypass grafts due to neointimal thickening is a major clinical burden. Identifying novel strategies to prevent neointimal thickening is important. Thus, this study aimed to identify microRNAs (miRNAs) that are dysregulated during neointimal formation and determine their pathophysiological relevance following miRNA manipulation.

Methods and results: We undertook a microarray approach to identify dysregulated miRNAs following engraftment in an interpositional porcine graft model. These profiling experiments identified a number of miRNAs which were dysregulated following engraftment. miR-21 levels were substantially elevated following engraftment and these results were confirmed by quantitative real-time PCR in mouse, pig, and human models of vein graft neointimal formation. Genetic ablation of miR-21 in mice or grafted veins dramatically reduced neointimal formation in a mouse model of vein grafting. Furthermore, pharmacological knockdown of miR-21 in human veins resulted in target gene de-repression and a significant reduction in neointimal formation.

Conclusion: This is the first report demonstrating that miR-21 plays a pathological role in vein graft failure. Furthermore, we also provided evidence that knockdown of miR-21 has therapeutic potential for the prevention of pathological vein graft remodelling.

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Related in: MedlinePlus

Immunolocalization of cells expressing miR-21 in murine vein grafts. (A) In situ hybridization with a scrambled LNA probe. (B) In situ hybridization for miR-21 in a mouse vein graft at 28 days. (C–F) Immunohistochemistry staining for proliferating cell nuclear antigens, smooth muscle cells (SMA), macrophages (mac-2), and control IgG, respectively. Scale bar represents 100 μm.
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EHT105F4: Immunolocalization of cells expressing miR-21 in murine vein grafts. (A) In situ hybridization with a scrambled LNA probe. (B) In situ hybridization for miR-21 in a mouse vein graft at 28 days. (C–F) Immunohistochemistry staining for proliferating cell nuclear antigens, smooth muscle cells (SMA), macrophages (mac-2), and control IgG, respectively. Scale bar represents 100 μm.

Mentions: In a second mouse model of vein grafting, the isogenic graft model (vena cava into the right common carotid artery), miR-21 levels were undetectable in the ungrafted inferior vena cava and low in the carotid artery compared with the wide spread and high-level expression in grafted tissue (Figure 3). We performed immunohistochemistry for α-actin and mac-2 to determine whether the cells expressing miR-21 were SMCs or macrophages, respectively. Comparison of the staining pattern for α-actin, mac-2, and proliferating cell nuclear antigen (PCNA) revealed that miR-21 is expressed in regions of the graft which stained positive for both SMC actin (α-actin) and macrophages (mac-2), although not all α-actin-positive cells expressed miR-21 (Figure 4A–D). These studies on sequential serial sections suggest that miR-21 is expressed in proliferating SMCs in the neointima and macrophages and actin-positive cells in the adventitial layer of the vein grafts. We next performed immunohistochemistry for the fibroblast markers vimentin and FSP-1.27 Cells staining positive for vimentin and FSP-1 were found extensively in the adventitia and a large proportion of cells in the neointimal layer, suggesting that myofibroblasts also contribute to miR-21 expression in the neointimal layer in this mouse model (see Supplementary material online, Figure S2). However, further detailed co-localization studies are needed to definitively demonstrate the cell types responsible for miR-21 expression.Figure 3


miRNA-21 is dysregulated in response to vein grafting in multiple models and genetic ablation in mice attenuates neointima formation.

McDonald RA, White KM, Wu J, Cooley BC, Robertson KE, Halliday CA, McClure JD, Francis S, Lu R, Kennedy S, George SJ, Wan S, van Rooij E, Baker AH - Eur. Heart J. (2013)

Immunolocalization of cells expressing miR-21 in murine vein grafts. (A) In situ hybridization with a scrambled LNA probe. (B) In situ hybridization for miR-21 in a mouse vein graft at 28 days. (C–F) Immunohistochemistry staining for proliferating cell nuclear antigens, smooth muscle cells (SMA), macrophages (mac-2), and control IgG, respectively. Scale bar represents 100 μm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3675389&req=5

EHT105F4: Immunolocalization of cells expressing miR-21 in murine vein grafts. (A) In situ hybridization with a scrambled LNA probe. (B) In situ hybridization for miR-21 in a mouse vein graft at 28 days. (C–F) Immunohistochemistry staining for proliferating cell nuclear antigens, smooth muscle cells (SMA), macrophages (mac-2), and control IgG, respectively. Scale bar represents 100 μm.
Mentions: In a second mouse model of vein grafting, the isogenic graft model (vena cava into the right common carotid artery), miR-21 levels were undetectable in the ungrafted inferior vena cava and low in the carotid artery compared with the wide spread and high-level expression in grafted tissue (Figure 3). We performed immunohistochemistry for α-actin and mac-2 to determine whether the cells expressing miR-21 were SMCs or macrophages, respectively. Comparison of the staining pattern for α-actin, mac-2, and proliferating cell nuclear antigen (PCNA) revealed that miR-21 is expressed in regions of the graft which stained positive for both SMC actin (α-actin) and macrophages (mac-2), although not all α-actin-positive cells expressed miR-21 (Figure 4A–D). These studies on sequential serial sections suggest that miR-21 is expressed in proliferating SMCs in the neointima and macrophages and actin-positive cells in the adventitial layer of the vein grafts. We next performed immunohistochemistry for the fibroblast markers vimentin and FSP-1.27 Cells staining positive for vimentin and FSP-1 were found extensively in the adventitia and a large proportion of cells in the neointimal layer, suggesting that myofibroblasts also contribute to miR-21 expression in the neointimal layer in this mouse model (see Supplementary material online, Figure S2). However, further detailed co-localization studies are needed to definitively demonstrate the cell types responsible for miR-21 expression.Figure 3

Bottom Line: Identifying novel strategies to prevent neointimal thickening is important.Thus, this study aimed to identify microRNAs (miRNAs) that are dysregulated during neointimal formation and determine their pathophysiological relevance following miRNA manipulation.Furthermore, pharmacological knockdown of miR-21 in human veins resulted in target gene de-repression and a significant reduction in neointimal formation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow G12 8TA, UK.

ABSTRACT

Aims: The long-term failure of autologous saphenous vein bypass grafts due to neointimal thickening is a major clinical burden. Identifying novel strategies to prevent neointimal thickening is important. Thus, this study aimed to identify microRNAs (miRNAs) that are dysregulated during neointimal formation and determine their pathophysiological relevance following miRNA manipulation.

Methods and results: We undertook a microarray approach to identify dysregulated miRNAs following engraftment in an interpositional porcine graft model. These profiling experiments identified a number of miRNAs which were dysregulated following engraftment. miR-21 levels were substantially elevated following engraftment and these results were confirmed by quantitative real-time PCR in mouse, pig, and human models of vein graft neointimal formation. Genetic ablation of miR-21 in mice or grafted veins dramatically reduced neointimal formation in a mouse model of vein grafting. Furthermore, pharmacological knockdown of miR-21 in human veins resulted in target gene de-repression and a significant reduction in neointimal formation.

Conclusion: This is the first report demonstrating that miR-21 plays a pathological role in vein graft failure. Furthermore, we also provided evidence that knockdown of miR-21 has therapeutic potential for the prevention of pathological vein graft remodelling.

Show MeSH
Related in: MedlinePlus