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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 does not attenuate FN content in the left carotid artery.(A) IHC for FN 14 days post-surgery. Bar, 100 μm. (B) Quantification of IHC intensity of FN in the intima-media area 7 (Sham: n = 6, Scrambled: n = 6, R1R2: n = 6) and 14 days (Sham: n = 7, Scrambled: n = 7, R1R2: n = 7) after ligating the vessel. * indicates p<0.05, ** p< 0.01 and *** p< 0.001.
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pone.0117356.g004: R1R2 does not attenuate FN content in the left carotid artery.(A) IHC for FN 14 days post-surgery. Bar, 100 μm. (B) Quantification of IHC intensity of FN in the intima-media area 7 (Sham: n = 6, Scrambled: n = 6, R1R2: n = 6) and 14 days (Sham: n = 7, Scrambled: n = 7, R1R2: n = 7) after ligating the vessel. * indicates p<0.05, ** p< 0.01 and *** p< 0.001.

Mentions: Our data show that R1R2 peptide inhibits the binding of collagen to FN (Fig. 1), which is consistent with published data in vitro [29,30]. To determine the inhibitory effect of R1R2 on ECM accumulation in vessels, we performed immunohistochemistry (IHC) and hydroxyproline assay for collagen type I. As aforementioned, no apparent difference in morphometry was observed among the groups of ligation, ligation with gel alone, and ligation with scrambled peptide (Fig. 2). Therefore, we used the experimental group treated with scrambled peptide as the control group. In Fig. 3A and 3B, IHC analysis showed a significant decrease of collagen type I intensity in R1R2-treated arteries compared to the control group on 7 and 14 days after the surgery (7 days: 0.117 ± 0.007, n = 6 vs. 0.198 ± 0.007, n = 6. 14 days: 0.136 ± 0.024, n = 5 vs. 0.151 ± 0.022, n = 6). Hydroxyproline assay (Fig. 3C, 1.522 ± 0.077, n = 3 vs. 2.133 ± 0.284, n = 3) showed a consistent reduction of collagen content in carotid vessels 14 days post-ligation. Previous in vitro studies have revealed that R1R2 inhibits the interaction between FN and collagen but does not affect FN fibril deposition into the ECM [29]. To determine whether R1R2 impairs FN accumulation in the vessel wall, we evaluated FN levels in the vessel wall after ligation using IHC (Fig. 4). Consistent with published in vitro results, there was no significant decrease of FN intensity between vessels treated with R1R2 and treated with scrambled peptide (7 days, 0.153 ± 0.012, n = 6 vs. 0.144 ± 0.005, n = 6. 14 days: 0.309 ± 0.005, n = 7 vs. 0.338 ± 0.016, n = 7). Collectively, these data show that R1R2 treatment in injured vessels significantly limits collagen type I deposition (Fig. 3), but not FN deposition (Fig. 4) 7 days and 14 days after ligation.


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 does not attenuate FN content in the left carotid artery.(A) IHC for FN 14 days post-surgery. Bar, 100 μm. (B) Quantification of IHC intensity of FN in the intima-media area 7 (Sham: n = 6, Scrambled: n = 6, R1R2: n = 6) and 14 days (Sham: n = 7, Scrambled: n = 7, R1R2: n = 7) after ligating the vessel. * indicates p<0.05, ** p< 0.01 and *** p< 0.001.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4326127&req=5

pone.0117356.g004: R1R2 does not attenuate FN content in the left carotid artery.(A) IHC for FN 14 days post-surgery. Bar, 100 μm. (B) Quantification of IHC intensity of FN in the intima-media area 7 (Sham: n = 6, Scrambled: n = 6, R1R2: n = 6) and 14 days (Sham: n = 7, Scrambled: n = 7, R1R2: n = 7) after ligating the vessel. * indicates p<0.05, ** p< 0.01 and *** p< 0.001.
Mentions: Our data show that R1R2 peptide inhibits the binding of collagen to FN (Fig. 1), which is consistent with published data in vitro [29,30]. To determine the inhibitory effect of R1R2 on ECM accumulation in vessels, we performed immunohistochemistry (IHC) and hydroxyproline assay for collagen type I. As aforementioned, no apparent difference in morphometry was observed among the groups of ligation, ligation with gel alone, and ligation with scrambled peptide (Fig. 2). Therefore, we used the experimental group treated with scrambled peptide as the control group. In Fig. 3A and 3B, IHC analysis showed a significant decrease of collagen type I intensity in R1R2-treated arteries compared to the control group on 7 and 14 days after the surgery (7 days: 0.117 ± 0.007, n = 6 vs. 0.198 ± 0.007, n = 6. 14 days: 0.136 ± 0.024, n = 5 vs. 0.151 ± 0.022, n = 6). Hydroxyproline assay (Fig. 3C, 1.522 ± 0.077, n = 3 vs. 2.133 ± 0.284, n = 3) showed a consistent reduction of collagen content in carotid vessels 14 days post-ligation. Previous in vitro studies have revealed that R1R2 inhibits the interaction between FN and collagen but does not affect FN fibril deposition into the ECM [29]. To determine whether R1R2 impairs FN accumulation in the vessel wall, we evaluated FN levels in the vessel wall after ligation using IHC (Fig. 4). Consistent with published in vitro results, there was no significant decrease of FN intensity between vessels treated with R1R2 and treated with scrambled peptide (7 days, 0.153 ± 0.012, n = 6 vs. 0.144 ± 0.005, n = 6. 14 days: 0.309 ± 0.005, n = 7 vs. 0.338 ± 0.016, n = 7). Collectively, these data show that R1R2 treatment in injured vessels significantly limits collagen type I deposition (Fig. 3), but not FN deposition (Fig. 4) 7 days and 14 days after ligation.

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