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Analysis of in situ and ex vivo αVβ3 integrin expression during experimental carotid atherogenesis.

Yao Y, Jiang Y, Sheng Z, Zhang Y, An Y, Yan F, Ma G, Liu N, Teng G, Cheng Z - Int J Nanomedicine (2012)

Bottom Line: Immunostaining confirmed the presence of mural α(V)β(3) integrin expression in macrophages in the neointima.Signal intensity increased in a macrophage density-dependent fashion in the stenotic segments.This expression can estimate the macrophage-bound inflammatory activity of atherosclerotic lesions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Zhongda Hospital, Medical School of Southeast University, Nanjing, Jiangsu, People's Republic of China.

ABSTRACT

Objective: Mural inflammation has been shown to contribute to the development of plaque, with the α(V)β(3) integrin highly expressed in atherosclerotic plaques. We herein examined α(V)β(3) integrin expression as a function of carotid atherosclerosis formation in the apolipoprotein E-deficient (apoE(-/-)) mouse.

Methods and results: Constrictive collars were placed around the left common carotid arteries of apo E(-/-) mice maintained on a high-fat diet (n = 14). Before and 21 days following collar placement, in vivo serial magnetic resonance imaging (MRI) measurements of the carotid aortic diameter were performed using a 7T magnetic resonance (MR) scanner. Near- infrared fluorescence (NIRF) imaging was performed (n = 6) using an in vivo imaging system 0-24 hours following administration of 1.0 nmol c(RGDyK)-Cy5.5 via the tail vein. A competition experiment was performed by the co-injection of a saturating dose of bicyclic RGD peptide H-Glu[cyclo(Arg-Gly-Asp-D-Tyr-Lys)]2 (n = 3). Following image acquisition and sacrifice at 24 hours after injection, carotid arteries were harvested for histological analyses. Neointima formation and arterial remodeling in the carotid arteries of apoE(-/-) mice were induced by the placement of a constrictive collar. Significantly greater fluorescent signals were obtained from constrictive collar left common carotid arteries as compared to uninvolved aortic segments in constrictive collar mice. Binding to stenotic lesions was efficiently blocked in competition experiments. Immunostaining confirmed the presence of mural α(V)β(3) integrin expression in macrophages in the neointima. Signal intensity increased in a macrophage density-dependent fashion in the stenotic segments.

Conclusion: Mural α(V)β(3) integrin expression, as determined using RGD-Cy5.5 near-infrared optical imaging, was increased in carotid arteries with constrictive collars in experimental mice. This expression can estimate the macrophage-bound inflammatory activity of atherosclerotic lesions.

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Accumulation of macrophages in the stenotic segment. (A) Representative micrograph of carotid artery sections stained with antibodies against αvβ3 integrin (400× magnification, scale bar = 100 μm). (B) Representative images of MAC-3 immunostaining in the stenotic lesion areas of the left carotid artery (400× magnification, scale bar = 100 μm). (C) Quantitative analyses of MAC-3-positive cells. (D) Colocalization in the proximal neointima, blue fluorescence of DAPI, the red fluorescence of Cy5.5 (probe) and the green fluorescence of FITC (macrophage) were examined (800× magnification).Notes: Upper panel shows representative photomicrographs of cross sections of left carotid artery proximal carotid site, white arrows show positive staining. Lower panel shows representative photomicrographs of cross sections of right carotid artery.Abbreviation: DAPI, diamidino-2-phenylindole dihydrochloride.
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f4-ijn-7-641: Accumulation of macrophages in the stenotic segment. (A) Representative micrograph of carotid artery sections stained with antibodies against αvβ3 integrin (400× magnification, scale bar = 100 μm). (B) Representative images of MAC-3 immunostaining in the stenotic lesion areas of the left carotid artery (400× magnification, scale bar = 100 μm). (C) Quantitative analyses of MAC-3-positive cells. (D) Colocalization in the proximal neointima, blue fluorescence of DAPI, the red fluorescence of Cy5.5 (probe) and the green fluorescence of FITC (macrophage) were examined (800× magnification).Notes: Upper panel shows representative photomicrographs of cross sections of left carotid artery proximal carotid site, white arrows show positive staining. Lower panel shows representative photomicrographs of cross sections of right carotid artery.Abbreviation: DAPI, diamidino-2-phenylindole dihydrochloride.

Mentions: We first explored αVβ3 integrin expression in the cross-sectional areas of the perivascular collar placement arterial segments. αVβ3 integrin expression was most apparent in the stenotic lesion areas of the left carotid artery. Figure 4A shows that αVβ3 integrin expression was significantly increased in the intima of the left stenotic carotid artery compared with the contralateral segment (localized to endothelial cells).


Analysis of in situ and ex vivo αVβ3 integrin expression during experimental carotid atherogenesis.

Yao Y, Jiang Y, Sheng Z, Zhang Y, An Y, Yan F, Ma G, Liu N, Teng G, Cheng Z - Int J Nanomedicine (2012)

Accumulation of macrophages in the stenotic segment. (A) Representative micrograph of carotid artery sections stained with antibodies against αvβ3 integrin (400× magnification, scale bar = 100 μm). (B) Representative images of MAC-3 immunostaining in the stenotic lesion areas of the left carotid artery (400× magnification, scale bar = 100 μm). (C) Quantitative analyses of MAC-3-positive cells. (D) Colocalization in the proximal neointima, blue fluorescence of DAPI, the red fluorescence of Cy5.5 (probe) and the green fluorescence of FITC (macrophage) were examined (800× magnification).Notes: Upper panel shows representative photomicrographs of cross sections of left carotid artery proximal carotid site, white arrows show positive staining. Lower panel shows representative photomicrographs of cross sections of right carotid artery.Abbreviation: DAPI, diamidino-2-phenylindole dihydrochloride.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3278228&req=5

f4-ijn-7-641: Accumulation of macrophages in the stenotic segment. (A) Representative micrograph of carotid artery sections stained with antibodies against αvβ3 integrin (400× magnification, scale bar = 100 μm). (B) Representative images of MAC-3 immunostaining in the stenotic lesion areas of the left carotid artery (400× magnification, scale bar = 100 μm). (C) Quantitative analyses of MAC-3-positive cells. (D) Colocalization in the proximal neointima, blue fluorescence of DAPI, the red fluorescence of Cy5.5 (probe) and the green fluorescence of FITC (macrophage) were examined (800× magnification).Notes: Upper panel shows representative photomicrographs of cross sections of left carotid artery proximal carotid site, white arrows show positive staining. Lower panel shows representative photomicrographs of cross sections of right carotid artery.Abbreviation: DAPI, diamidino-2-phenylindole dihydrochloride.
Mentions: We first explored αVβ3 integrin expression in the cross-sectional areas of the perivascular collar placement arterial segments. αVβ3 integrin expression was most apparent in the stenotic lesion areas of the left carotid artery. Figure 4A shows that αVβ3 integrin expression was significantly increased in the intima of the left stenotic carotid artery compared with the contralateral segment (localized to endothelial cells).

Bottom Line: Immunostaining confirmed the presence of mural α(V)β(3) integrin expression in macrophages in the neointima.Signal intensity increased in a macrophage density-dependent fashion in the stenotic segments.This expression can estimate the macrophage-bound inflammatory activity of atherosclerotic lesions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Zhongda Hospital, Medical School of Southeast University, Nanjing, Jiangsu, People's Republic of China.

ABSTRACT

Objective: Mural inflammation has been shown to contribute to the development of plaque, with the α(V)β(3) integrin highly expressed in atherosclerotic plaques. We herein examined α(V)β(3) integrin expression as a function of carotid atherosclerosis formation in the apolipoprotein E-deficient (apoE(-/-)) mouse.

Methods and results: Constrictive collars were placed around the left common carotid arteries of apo E(-/-) mice maintained on a high-fat diet (n = 14). Before and 21 days following collar placement, in vivo serial magnetic resonance imaging (MRI) measurements of the carotid aortic diameter were performed using a 7T magnetic resonance (MR) scanner. Near- infrared fluorescence (NIRF) imaging was performed (n = 6) using an in vivo imaging system 0-24 hours following administration of 1.0 nmol c(RGDyK)-Cy5.5 via the tail vein. A competition experiment was performed by the co-injection of a saturating dose of bicyclic RGD peptide H-Glu[cyclo(Arg-Gly-Asp-D-Tyr-Lys)]2 (n = 3). Following image acquisition and sacrifice at 24 hours after injection, carotid arteries were harvested for histological analyses. Neointima formation and arterial remodeling in the carotid arteries of apoE(-/-) mice were induced by the placement of a constrictive collar. Significantly greater fluorescent signals were obtained from constrictive collar left common carotid arteries as compared to uninvolved aortic segments in constrictive collar mice. Binding to stenotic lesions was efficiently blocked in competition experiments. Immunostaining confirmed the presence of mural α(V)β(3) integrin expression in macrophages in the neointima. Signal intensity increased in a macrophage density-dependent fashion in the stenotic segments.

Conclusion: Mural α(V)β(3) integrin expression, as determined using RGD-Cy5.5 near-infrared optical imaging, was increased in carotid arteries with constrictive collars in experimental mice. This expression can estimate the macrophage-bound inflammatory activity of atherosclerotic lesions.

Show MeSH
Related in: MedlinePlus