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Collagen inhibitory peptide R1R2 mediates vascular remodeling by decreasing inflammation and smooth muscle cell activation.

Lee TH, Sottile J, Chiang HY - PLoS ONE (2015)

Bottom Line: Morphometric analysis demonstrated that R1R2 reduced intima-media thickening compared to the control groups.This decrease was accompanied by decreased VCAM-1 and ICAM-1 expression.Our in vitro studies revealed that R1R2 attenuated SMC proliferation and migration, and also decreased monocyte adhesion and transendothelial migration through endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan.

ABSTRACT
The extracellular matrix (ECM) is a major constituent of the vessel wall. In addition to providing a structural scaffold, the ECM controls numerous cellular functions in both physiologic and pathologic settings. Vascular remodeling occurs after injury and is characterized by endothelial cell activation, inflammatory cell infiltration, phenotypic modulation of smooth muscle cells (SMCs), and augmented deposition of collagen-rich ECM. R1R2, a peptide derived from the bacterial adhesin SFS, with sequence homology to collagen, is known to inhibit collagen type I deposition in vitro by inhibiting the binding of fibronectin to collagen. However, the inhibitory effects of R1R2 during vascular remodeling have not been explored. We periadventitially delivered R1R2 to carotid arteries using pluronic gel in a vascular remodeling mouse model induced by blood flow cessation, and evaluated its effects on intima-media thickening, ECM deposition, SMC activation, and inflammatory cell infiltration. Morphometric analysis demonstrated that R1R2 reduced intima-media thickening compared to the control groups. R1R2 treatment also decreased collagen type I deposition in the vessel wall, and maintained SMC in the contractile phenotype. Interestingly, R1R2 dramatically reduced inflammatory cell infiltration into the vessel by ∼ 78%. This decrease was accompanied by decreased VCAM-1 and ICAM-1 expression. Our in vitro studies revealed that R1R2 attenuated SMC proliferation and migration, and also decreased monocyte adhesion and transendothelial migration through endothelial cells. Together, these data suggest that R1R2 attenuates vascular remodeling responses by decreasing inflammation and by modulating SMC proliferation and migration, and suggest that the R1R2 peptide may have therapeutic potential in treating occlusive vascular diseases.

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R1R2 decreases inflammatory cell accumulation and VCAM-1 and ICAM-1 levels.(A) Photographs showing representative immunostaining of CD-45, VCAM-1 and ICAM-1 in the left carotid artery from the animals subjected to ligation 7 days after the surgery. Arrows indicate CD45 (+) leukocytes. Bar, 50 μm. (B-D) Percentage of the area which is CD45 (+) (Sham: n = 7, Scrambled: n = 6, R1R2: n = 5) (B), VCAM-1 (+) (Sham: n = 7, Scrambled: n = 9, R1R2: n = 7) (C) or ICAM-1 (+) (Sham: n = 5, Scrambled: n = 8, R1R2: n = 7) (D) were assessed in the intima-media of the vessel. (E) Western blot analysis of ICAM-1 and VCAM-1 expression in ligated carotid artery at 7 days. Equal protein loading was confirmed with GAPDH. * indicates p<0.05 and ** p<0.01.
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pone.0117356.g007: R1R2 decreases inflammatory cell accumulation and VCAM-1 and ICAM-1 levels.(A) Photographs showing representative immunostaining of CD-45, VCAM-1 and ICAM-1 in the left carotid artery from the animals subjected to ligation 7 days after the surgery. Arrows indicate CD45 (+) leukocytes. Bar, 50 μm. (B-D) Percentage of the area which is CD45 (+) (Sham: n = 7, Scrambled: n = 6, R1R2: n = 5) (B), VCAM-1 (+) (Sham: n = 7, Scrambled: n = 9, R1R2: n = 7) (C) or ICAM-1 (+) (Sham: n = 5, Scrambled: n = 8, R1R2: n = 7) (D) were assessed in the intima-media of the vessel. (E) Western blot analysis of ICAM-1 and VCAM-1 expression in ligated carotid artery at 7 days. Equal protein loading was confirmed with GAPDH. * indicates p<0.05 and ** p<0.01.

Mentions: Inflammatory cell infiltration is a key event in neointimal formation after vascular injury [42,46]. Consistent with the data shown by other labs [35], reduced flow induced an increase of CD45(+) leukocytes in the intima and media 7 days post-surgery (Fig. 7). Of note, we observed a dramatic decrease in the number of CD45 (+) cells in response to R1R2 treatment (0.094 ± 0.035%, n = 5 vs. 0.441 ± 0.049%, n = 6). Furthermore, R1R2 also decreased the expression of cell adhesion molecules. IHC analysis showed that R1R2 application resulted in a dramatic reduction of vascular cell adhesion molecule-1 (VCAM-1) (Fig. 7A and 7C, 10.36 ± 1.802%, n = 7 vs. 25.72 ± 1.966%, n = 9) and intercellular cell adhesion molecule-1 (ICAM-1) (Fig. 7A and 7D, 11.58 ± 2.387%, n = 7 vs. 30.39 ± 3.016%, n = 8) expression in comparison with control peptide-treated vessels 7 days after ligation. Additionally, immunoblotting of VCAM-1 and ICAM-1 also showed a similar reduction in protein levels in the carotid artery following surgery (Fig. 7E). Taken in aggregate, our in vivo findings uncover an inhibitory effect of R1R2 peptide on leukocyte infiltration and CAM expression in the arterial wall after injury.


Collagen inhibitory peptide R1R2 mediates vascular remodeling by decreasing inflammation and smooth muscle cell activation.

Lee TH, Sottile J, Chiang HY - PLoS ONE (2015)

R1R2 decreases inflammatory cell accumulation and VCAM-1 and ICAM-1 levels.(A) Photographs showing representative immunostaining of CD-45, VCAM-1 and ICAM-1 in the left carotid artery from the animals subjected to ligation 7 days after the surgery. Arrows indicate CD45 (+) leukocytes. Bar, 50 μm. (B-D) Percentage of the area which is CD45 (+) (Sham: n = 7, Scrambled: n = 6, R1R2: n = 5) (B), VCAM-1 (+) (Sham: n = 7, Scrambled: n = 9, R1R2: n = 7) (C) or ICAM-1 (+) (Sham: n = 5, Scrambled: n = 8, R1R2: n = 7) (D) were assessed in the intima-media of the vessel. (E) Western blot analysis of ICAM-1 and VCAM-1 expression in ligated carotid artery at 7 days. Equal protein loading was confirmed with GAPDH. * indicates p<0.05 and ** p<0.01.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4326127&req=5

pone.0117356.g007: R1R2 decreases inflammatory cell accumulation and VCAM-1 and ICAM-1 levels.(A) Photographs showing representative immunostaining of CD-45, VCAM-1 and ICAM-1 in the left carotid artery from the animals subjected to ligation 7 days after the surgery. Arrows indicate CD45 (+) leukocytes. Bar, 50 μm. (B-D) Percentage of the area which is CD45 (+) (Sham: n = 7, Scrambled: n = 6, R1R2: n = 5) (B), VCAM-1 (+) (Sham: n = 7, Scrambled: n = 9, R1R2: n = 7) (C) or ICAM-1 (+) (Sham: n = 5, Scrambled: n = 8, R1R2: n = 7) (D) were assessed in the intima-media of the vessel. (E) Western blot analysis of ICAM-1 and VCAM-1 expression in ligated carotid artery at 7 days. Equal protein loading was confirmed with GAPDH. * indicates p<0.05 and ** p<0.01.
Mentions: Inflammatory cell infiltration is a key event in neointimal formation after vascular injury [42,46]. Consistent with the data shown by other labs [35], reduced flow induced an increase of CD45(+) leukocytes in the intima and media 7 days post-surgery (Fig. 7). Of note, we observed a dramatic decrease in the number of CD45 (+) cells in response to R1R2 treatment (0.094 ± 0.035%, n = 5 vs. 0.441 ± 0.049%, n = 6). Furthermore, R1R2 also decreased the expression of cell adhesion molecules. IHC analysis showed that R1R2 application resulted in a dramatic reduction of vascular cell adhesion molecule-1 (VCAM-1) (Fig. 7A and 7C, 10.36 ± 1.802%, n = 7 vs. 25.72 ± 1.966%, n = 9) and intercellular cell adhesion molecule-1 (ICAM-1) (Fig. 7A and 7D, 11.58 ± 2.387%, n = 7 vs. 30.39 ± 3.016%, n = 8) expression in comparison with control peptide-treated vessels 7 days after ligation. Additionally, immunoblotting of VCAM-1 and ICAM-1 also showed a similar reduction in protein levels in the carotid artery following surgery (Fig. 7E). Taken in aggregate, our in vivo findings uncover an inhibitory effect of R1R2 peptide on leukocyte infiltration and CAM expression in the arterial wall after injury.

Bottom Line: Morphometric analysis demonstrated that R1R2 reduced intima-media thickening compared to the control groups.This decrease was accompanied by decreased VCAM-1 and ICAM-1 expression.Our in vitro studies revealed that R1R2 attenuated SMC proliferation and migration, and also decreased monocyte adhesion and transendothelial migration through endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan.

ABSTRACT
The extracellular matrix (ECM) is a major constituent of the vessel wall. In addition to providing a structural scaffold, the ECM controls numerous cellular functions in both physiologic and pathologic settings. Vascular remodeling occurs after injury and is characterized by endothelial cell activation, inflammatory cell infiltration, phenotypic modulation of smooth muscle cells (SMCs), and augmented deposition of collagen-rich ECM. R1R2, a peptide derived from the bacterial adhesin SFS, with sequence homology to collagen, is known to inhibit collagen type I deposition in vitro by inhibiting the binding of fibronectin to collagen. However, the inhibitory effects of R1R2 during vascular remodeling have not been explored. We periadventitially delivered R1R2 to carotid arteries using pluronic gel in a vascular remodeling mouse model induced by blood flow cessation, and evaluated its effects on intima-media thickening, ECM deposition, SMC activation, and inflammatory cell infiltration. Morphometric analysis demonstrated that R1R2 reduced intima-media thickening compared to the control groups. R1R2 treatment also decreased collagen type I deposition in the vessel wall, and maintained SMC in the contractile phenotype. Interestingly, R1R2 dramatically reduced inflammatory cell infiltration into the vessel by ∼ 78%. This decrease was accompanied by decreased VCAM-1 and ICAM-1 expression. Our in vitro studies revealed that R1R2 attenuated SMC proliferation and migration, and also decreased monocyte adhesion and transendothelial migration through endothelial cells. Together, these data suggest that R1R2 attenuates vascular remodeling responses by decreasing inflammation and by modulating SMC proliferation and migration, and suggest that the R1R2 peptide may have therapeutic potential in treating occlusive vascular diseases.

Show MeSH
Related in: MedlinePlus