TY - JOUR
T1 - Versatile click alginate hydrogels with protease-sensitive domains as cell responsive/instructive 3D microenvironments
AU - Neves, Mariana I.
AU - Magalhães, Mariana V.
AU - Bidarra, Sílvia J.
AU - Moroni, Lorenzo
AU - Barrias, Cristina C.
N1 - Funding Information:
This work was supported by project EndoSWITCH ( PTDC/BTM-ORG/5154/2020 ) funded by FCT (Portuguese Foundation for Science and Technology ). The authors thank FCT for CCB's IF research position (Grant No: IF/00296/2015), MIN's scholarship (Grant No: SFRH/BD/129855/2017 and COVID/BD/151886/2022 ), MVM's scholarship (Grant No: SFRH/BD/10184/2022 ), and SJB's research contract DL 57/2016/CP1360/CT0006 .
Funding Information:
This work was supported by project EndoSWITCH (PTDC/BTM-ORG/5154/2020) funded by FCT (Portuguese Foundation for Science and Technology). The authors thank FCT for CCB's IF research position (Grant No: IF/00296/2015), MIN's scholarship (Grant No: SFRH/BD/129855/2017 and COVID/BD/151886/2022), MVM's scholarship (Grant No: SFRH/BD/10184/2022), and SJB's research contract DL 57/2016/CP1360/CT0006.The authors acknowledge the support of i3S Scientific Platforms: “Bioimaging” member of the PPBI (Grant No: PPBI-POCI-01-0145-FEDER-022122), “Biointerfaces and Nanotechnology” (Grant No: UID/BIM/04293/2019), the “Proteomics Scientific Platform”, in particular Hugo Osório, financed by the Portuguese Mass Spectrometry Network, integrated in the National Roadmap of Research Infrastructures of Strategic Relevance (ROTEIRO/0028/2013; LISBOA-01-0145-FEDER-022125). The authors also acknowledge Paulo Aguiar, Group Leader of Neuroengineering and Computational Neuroscience Group at i3S for the development of the Fiji macro for the quantification of cell-cell interconnectivity degree. Finally, the authors acknowledge the Laboratory for Structural Elucidation (LAE) and the Image, Microstructure and Microanalysis Unit (IMCROS) from the Materials Centre of the University of Porto (CEMUP).
Funding Information:
The authors acknowledge the support of i3S Scientific Platforms: “Bioimaging” member of the PPBI (Grant No: PPBI-POCI-01-0145-FEDER-022122 ), “Biointerfaces and Nanotechnology” (Grant No: UID/BIM/04293/2019 ), the “Proteomics Scientific Platform”, in particular Hugo Osório, financed by the Portuguese Mass Spectrometry Network , integrated in the National Roadmap of Research Infrastructures of Strategic Relevance ( ROTEIRO/0028/2013 ; LISBOA-01-0145-FEDER-022125 ). The authors also acknowledge Paulo Aguiar, Group Leader of Neuroengineering and Computational Neuroscience Group at i3S for the development of the Fiji macro for the quantification of cell-cell interconnectivity degree. Finally, the authors acknowledge the Laboratory for Structural Elucidation (LAE) and the Image, Microstructure and Microanalysis Unit (IMCROS) from the Materials Centre of the University of Porto (CEMUP).
Publisher Copyright:
© 2023 Elsevier Ltd
PY - 2023/11/15
Y1 - 2023/11/15
N2 - Alginate (ALG) is a widely used biomaterial to create artificial extracellular matrices (ECM) for tissue engineering. Since it does not degrade in the human body, imparting proteolytic sensitivity to ALG hydrogels leverages their properties as ECM-mimics. Herein, we explored the strain-promoted azide-alkyne cycloaddition (SPAAC) as a biocompatible and bio-orthogonal click-chemistry to graft cyclooctyne-modified alginate (ALG-K) with bi-azide-functionalized PVGLIG peptides. These are sensitive to matrix metalloproteinase (MMP) and may act as crosslinkers. The ALG-K-PVGLIG conjugates (50, 125, and 250 µM PVGLIG) were characterized for peptide incorporation, crosslinking ability (double-end grafting), and enzymatic liability. For producing cell-permissive multifunctional 3D matrices for dermal fibroblast culture, oxidized ALG-K was grafted with PVGLIG and with RGD peptides for cell-adhesion. SPAAC reactions were performed immediately before cell-laden hydrogel formation by secondary ionic-crosslinking, considerably reducing the steps and time of preparation. Hydrogels with intermediate PVGLIG concentration (125 µM) presented slightly higher stiffness while promoting extensive cell spreading and higher degree of cell-cell interconnections, likely favored by cell-driven proteolytic remodeling of the network. The hydrogel-embedded cells were able to produce their own pericellular ECM, expressed MMP-2 and 14, and secreted PVGLIG-degrading enzymes. By recapitulating key ECM-like features, these hydrogels provide biologically relevant 3D matrices for soft tissue regeneration.
AB - Alginate (ALG) is a widely used biomaterial to create artificial extracellular matrices (ECM) for tissue engineering. Since it does not degrade in the human body, imparting proteolytic sensitivity to ALG hydrogels leverages their properties as ECM-mimics. Herein, we explored the strain-promoted azide-alkyne cycloaddition (SPAAC) as a biocompatible and bio-orthogonal click-chemistry to graft cyclooctyne-modified alginate (ALG-K) with bi-azide-functionalized PVGLIG peptides. These are sensitive to matrix metalloproteinase (MMP) and may act as crosslinkers. The ALG-K-PVGLIG conjugates (50, 125, and 250 µM PVGLIG) were characterized for peptide incorporation, crosslinking ability (double-end grafting), and enzymatic liability. For producing cell-permissive multifunctional 3D matrices for dermal fibroblast culture, oxidized ALG-K was grafted with PVGLIG and with RGD peptides for cell-adhesion. SPAAC reactions were performed immediately before cell-laden hydrogel formation by secondary ionic-crosslinking, considerably reducing the steps and time of preparation. Hydrogels with intermediate PVGLIG concentration (125 µM) presented slightly higher stiffness while promoting extensive cell spreading and higher degree of cell-cell interconnections, likely favored by cell-driven proteolytic remodeling of the network. The hydrogel-embedded cells were able to produce their own pericellular ECM, expressed MMP-2 and 14, and secreted PVGLIG-degrading enzymes. By recapitulating key ECM-like features, these hydrogels provide biologically relevant 3D matrices for soft tissue regeneration.
KW - Bio-orthogonal
KW - Bioactive
KW - Matrix remodeling
KW - Protease sensitive
U2 - 10.1016/j.carbpol.2023.121226
DO - 10.1016/j.carbpol.2023.121226
M3 - Article
SN - 0144-8617
VL - 320
JO - Carbohydrate Polymers
JF - Carbohydrate Polymers
IS - 1
M1 - 121226
ER -