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Light-triggered in vivo activation of adhesive peptides regulates cell adhesion, inflammation and vascularization of biomaterials.

Lee TT, García JR, Paez JI, Singh A, Phelps EA, Weis S, Shafiq Z, Shekaran A, Del Campo A, García AJ - Nat Mater (2014)

Bottom Line: Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control.We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material.This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

View Article: PubMed Central - PubMed

Affiliation: 1] Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332, USA [2] Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.

ABSTRACT
Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control. Although materials with temporally regulated presentation of bioadhesive ligands using external triggers, such as light and electric fields, have recently been realized for cells in culture, the impact of in vivo temporal ligand presentation on cell-material responses is unknown. Here, we present a general strategy to temporally and spatially control the in vivo presentation of bioligands using cell-adhesive peptides with a protecting group that can be easily removed via transdermal light exposure to render the peptide fully active. We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material. This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

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Related in: MedlinePlus

Light-based activation of cell adhesive peptide promotes vascularization of implanted biomaterialsa, Immunostaining images for hydrogels presenting caged RGD or caged RDG peptide for different UV exposure conditions. Top: green = CD31 [endothelial cell], magenta = αSMA [smooth muscle cell], blue = DAPI [DNA], scale bar, 100 μm. Bottom: magenta = CD68 [macrophage], blue = DAPI [DNA], scale bar, 100 μm. b, Fluorescent images of blood vessel ingrowth (green) into PEG-maleimide hydrogels implanted subcutaneously at 14 days (scale bar, 100 μm). PEG-maleimide hydrogels presenting peptides were implanted subcutaneously and exposed to UV transdermally at selected time points. Mice were perfused with DyLight488-conjugated tomato lectin at sacrifice to label functional blood vessels. Hydrogels presenting caged RGD peptides which were exposed to UV transdermally at Day 0 and Day 7 exhibited robust blood vessel growth, similar to gels presenting control RGD. Hydrogels functionalized with scrambled RDG peptide or caged RGD peptide that was not exposed to UV displayed minimal blood vessel infiltration. c, Blood vessel density, box-whisker plot (minimum, 25th percentile, median, 75th percentile, and maximum) for 4 mice per group for caged RGD conditions, 3 mice per group for control peptides. Kruskal-Wallis p < 0.01, * p < 0.01 vs. RDG, † p < 0.05 vs. No UV Caged RGD.
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Figure 5: Light-based activation of cell adhesive peptide promotes vascularization of implanted biomaterialsa, Immunostaining images for hydrogels presenting caged RGD or caged RDG peptide for different UV exposure conditions. Top: green = CD31 [endothelial cell], magenta = αSMA [smooth muscle cell], blue = DAPI [DNA], scale bar, 100 μm. Bottom: magenta = CD68 [macrophage], blue = DAPI [DNA], scale bar, 100 μm. b, Fluorescent images of blood vessel ingrowth (green) into PEG-maleimide hydrogels implanted subcutaneously at 14 days (scale bar, 100 μm). PEG-maleimide hydrogels presenting peptides were implanted subcutaneously and exposed to UV transdermally at selected time points. Mice were perfused with DyLight488-conjugated tomato lectin at sacrifice to label functional blood vessels. Hydrogels presenting caged RGD peptides which were exposed to UV transdermally at Day 0 and Day 7 exhibited robust blood vessel growth, similar to gels presenting control RGD. Hydrogels functionalized with scrambled RDG peptide or caged RGD peptide that was not exposed to UV displayed minimal blood vessel infiltration. c, Blood vessel density, box-whisker plot (minimum, 25th percentile, median, 75th percentile, and maximum) for 4 mice per group for caged RGD conditions, 3 mice per group for control peptides. Kruskal-Wallis p < 0.01, * p < 0.01 vs. RDG, † p < 0.05 vs. No UV Caged RGD.

Mentions: To examine triggerable control over in vivo vascularization, PEG-maleimide hydrogels presenting caged RGD or RDG peptides, protease-degradable crosslinks, and VEGF vasculogenic protein were polymerized directly into subcutaneous pockets in mice. At 0 or 7 days post-implantation, implants were exposed to UV light transdermally. Following sacrifice at day 14, hydrogels were explanted, stained for different cell markers, and analyzed by confocal microscopy to examine cell infiltration and vascularization within the hydrogel. As shown by cell nuclei staining, all implanted hydrogels exhibited high cell numbers within the implant with no gross differences for cell infiltration among hydrogel type (RGD, RDG) or UV exposure condition (Fig. 5a). This observation is in agreement with previous reports 26–28. Staining for inflammatory cell markers revealed that the large majority of cells were macrophages and no staining for neutrophils was evident, which is the expected inflammatory cell profile at this time point 23,25. Importantly, hydrogels presenting caged RGD peptide that had been UV-exposed transdermally at day 0 or day 7 contained many tubular structures that stained positive for the endothelial cell marker CD31 and smooth muscle cell marker αSMA (Fig. 5a and S9), indicating vascularization of these hydrogels. In contrast, hydrogels presenting caged RGD peptide which had not been exposed to UV light exhibited diffuse staining for CD31 and αSMA, and no tubular structures resembling blood vessels were observed (Fig. 5a and S9). Likewise, hydrogels presenting caged scrambled RDG peptide showed diffuse CD31 and αSMA staining and no evidence of blood vessels for any UV exposure condition. These results demonstrate that UV-light triggered activation of RGD peptides promotes vascularization of these hydrogels.


Light-triggered in vivo activation of adhesive peptides regulates cell adhesion, inflammation and vascularization of biomaterials.

Lee TT, García JR, Paez JI, Singh A, Phelps EA, Weis S, Shafiq Z, Shekaran A, Del Campo A, García AJ - Nat Mater (2014)

Light-based activation of cell adhesive peptide promotes vascularization of implanted biomaterialsa, Immunostaining images for hydrogels presenting caged RGD or caged RDG peptide for different UV exposure conditions. Top: green = CD31 [endothelial cell], magenta = αSMA [smooth muscle cell], blue = DAPI [DNA], scale bar, 100 μm. Bottom: magenta = CD68 [macrophage], blue = DAPI [DNA], scale bar, 100 μm. b, Fluorescent images of blood vessel ingrowth (green) into PEG-maleimide hydrogels implanted subcutaneously at 14 days (scale bar, 100 μm). PEG-maleimide hydrogels presenting peptides were implanted subcutaneously and exposed to UV transdermally at selected time points. Mice were perfused with DyLight488-conjugated tomato lectin at sacrifice to label functional blood vessels. Hydrogels presenting caged RGD peptides which were exposed to UV transdermally at Day 0 and Day 7 exhibited robust blood vessel growth, similar to gels presenting control RGD. Hydrogels functionalized with scrambled RDG peptide or caged RGD peptide that was not exposed to UV displayed minimal blood vessel infiltration. c, Blood vessel density, box-whisker plot (minimum, 25th percentile, median, 75th percentile, and maximum) for 4 mice per group for caged RGD conditions, 3 mice per group for control peptides. Kruskal-Wallis p < 0.01, * p < 0.01 vs. RDG, † p < 0.05 vs. No UV Caged RGD.
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Related In: Results  -  Collection

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Figure 5: Light-based activation of cell adhesive peptide promotes vascularization of implanted biomaterialsa, Immunostaining images for hydrogels presenting caged RGD or caged RDG peptide for different UV exposure conditions. Top: green = CD31 [endothelial cell], magenta = αSMA [smooth muscle cell], blue = DAPI [DNA], scale bar, 100 μm. Bottom: magenta = CD68 [macrophage], blue = DAPI [DNA], scale bar, 100 μm. b, Fluorescent images of blood vessel ingrowth (green) into PEG-maleimide hydrogels implanted subcutaneously at 14 days (scale bar, 100 μm). PEG-maleimide hydrogels presenting peptides were implanted subcutaneously and exposed to UV transdermally at selected time points. Mice were perfused with DyLight488-conjugated tomato lectin at sacrifice to label functional blood vessels. Hydrogels presenting caged RGD peptides which were exposed to UV transdermally at Day 0 and Day 7 exhibited robust blood vessel growth, similar to gels presenting control RGD. Hydrogels functionalized with scrambled RDG peptide or caged RGD peptide that was not exposed to UV displayed minimal blood vessel infiltration. c, Blood vessel density, box-whisker plot (minimum, 25th percentile, median, 75th percentile, and maximum) for 4 mice per group for caged RGD conditions, 3 mice per group for control peptides. Kruskal-Wallis p < 0.01, * p < 0.01 vs. RDG, † p < 0.05 vs. No UV Caged RGD.
Mentions: To examine triggerable control over in vivo vascularization, PEG-maleimide hydrogels presenting caged RGD or RDG peptides, protease-degradable crosslinks, and VEGF vasculogenic protein were polymerized directly into subcutaneous pockets in mice. At 0 or 7 days post-implantation, implants were exposed to UV light transdermally. Following sacrifice at day 14, hydrogels were explanted, stained for different cell markers, and analyzed by confocal microscopy to examine cell infiltration and vascularization within the hydrogel. As shown by cell nuclei staining, all implanted hydrogels exhibited high cell numbers within the implant with no gross differences for cell infiltration among hydrogel type (RGD, RDG) or UV exposure condition (Fig. 5a). This observation is in agreement with previous reports 26–28. Staining for inflammatory cell markers revealed that the large majority of cells were macrophages and no staining for neutrophils was evident, which is the expected inflammatory cell profile at this time point 23,25. Importantly, hydrogels presenting caged RGD peptide that had been UV-exposed transdermally at day 0 or day 7 contained many tubular structures that stained positive for the endothelial cell marker CD31 and smooth muscle cell marker αSMA (Fig. 5a and S9), indicating vascularization of these hydrogels. In contrast, hydrogels presenting caged RGD peptide which had not been exposed to UV light exhibited diffuse staining for CD31 and αSMA, and no tubular structures resembling blood vessels were observed (Fig. 5a and S9). Likewise, hydrogels presenting caged scrambled RDG peptide showed diffuse CD31 and αSMA staining and no evidence of blood vessels for any UV exposure condition. These results demonstrate that UV-light triggered activation of RGD peptides promotes vascularization of these hydrogels.

Bottom Line: Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control.We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material.This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

View Article: PubMed Central - PubMed

Affiliation: 1] Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332, USA [2] Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.

ABSTRACT
Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control. Although materials with temporally regulated presentation of bioadhesive ligands using external triggers, such as light and electric fields, have recently been realized for cells in culture, the impact of in vivo temporal ligand presentation on cell-material responses is unknown. Here, we present a general strategy to temporally and spatially control the in vivo presentation of bioligands using cell-adhesive peptides with a protecting group that can be easily removed via transdermal light exposure to render the peptide fully active. We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material. This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

Show MeSH
Related in: MedlinePlus