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  • 1.
    Rasti Boroojeni, Fatemeh
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Naeimipour, Sajjad
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Lifwergren, Philip
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Abrahamsson, Annelie
    Linköping University, Department of Biomedical and Clinical Sciences, Division of Surgery, Orthopedics and Oncology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Surgery, Orthopaedics and Cancer Treatment, Department of Oncology.
    Dabrosin, Charlotta
    Linköping University, Department of Biomedical and Clinical Sciences, Division of Surgery, Orthopedics and Oncology. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Surgery, Orthopaedics and Cancer Treatment, Department of Oncology.
    Selegård, Robert
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Aili, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Proteolytic remodeling of 3D bioprinted tumor microenvironments2024In: Biofabrication, ISSN 1758-5082, E-ISSN 1758-5090, Vol. 16, no 2, article id 025002Article in journal (Refereed)
    Abstract [en]

    In native tissue, remodeling of the pericellular space is essential for cellular activities and is mediated by tightly regulated proteases. Protease activity is dysregulated in many diseases, including many forms of cancer. Increased proteolytic activity is directly linked to tumor invasion into stroma, metastasis, and angiogenesis as well as all other hallmarks of cancer. Here we show a strategy for 3D bioprinting of breast cancer models using well-defined protease degradable hydrogels that can facilitate exploration of the multifaceted roles of proteolytic extracellular matrix remodeling in tumor progression. We designed a set of bicyclo[6.1.0]nonyne functionalized hyaluronan (HA)-based bioinks cross-linked by azide-modified poly(ethylene glycol) (PEG) or matrix metalloproteinase (MMP) degradable azide-functionalized peptides. Bioprinted structures combining PEG and peptide-based hydrogels were proteolytically degraded with spatial selectivity, leaving non-degradable features intact. Bioprinting of tumor-mimicking microenvironments using bioinks comprising human breast cancer cells (MCF-7) and fibroblast in hydrogels with different susceptibilities to proteolytic degradation shows that MCF-7 proliferation and spheroid size were significantly increased in protease degradable hydrogel compartments, but only in the presence of fibroblasts. In the absence of fibroblasts in the stromal compartment, cancer cell proliferation was reduced and did not differ between degradable and nondegradable hydrogels. The interactions between spatially separated fibroblasts and MCF-7 cells consequently resulted in protease-mediated remodeling of the bioprinted structures and a significant increase in cancer cell spheroid size, highlighting the close interplay between cancer cells and stromal cells in the tumor microenvironment and the influence of proteases in tumor progression.

  • 2.
    Naeimipour, Sajjad
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Rasti Boroojeni, Fatemeh
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Lifwergren, Philip
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Selegård, Robert
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Aili, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Multimodal and dynamic cross-linking of modular thiolated alginate-based bioinks2023In: Materials Today Advances, E-ISSN 2590-0498, Vol. 19, article id 100415Article in journal (Refereed)
    Abstract [en]

    Engineered extracellular matrix-mimicking hydrogels can facilitate 3D cell culture and fabrication of tissue-like constructs and biologically relevant disease models. Processing of cell-laden hydrogels using additive manufacturing techniques further allows for the development of tissue-mimetic structures with higher spatial complexity. Whereas a wide range of printable hydrogels is available, they tend to lack biological functionality and cell compatibility. Here we show an enzymatically mediated thiol-based cross-linking strategy for the design of modular and cytocompatible hydrogel-based bioinks for 3D bioprinting of dynamic multi-material architectures. Alginate is functionalized with cysteines modified with an enzyme-labile thiol protection group. Deprotection using penicillin G acylase (PGA) generates free thiols on-demand, enabling hydrogel cross-linking using thiol-reactive cross-linkers and intermolecular disulfides while avoiding undesired and uncontrolled thiol oxidation. Remaining free thiols can be used for post-printing hydrogel functionalization and lamination of multilayer structures. Moreover, the addition of PGA to a thermo-reversible hydrogel support bath enables the bioprinting of cell-laden 3D structures with high cell viability and excellent shape fidelity. The possibilities to enzymatically generate free thiols during bioprinting facilitate cross-linking and tuning of bioink properties using cytocompatible chemistries and allow for the printing of complex and dynamic cell-laden structures.

  • 3.
    Aronsson, Christopher
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Physics. Linköping University, Faculty of Science & Engineering.
    Jury, Michael
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Naeimipour, Sajjad
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Rasti Boroojeni, Fatemeh
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Christoffersson, Jonas
    Linköping University, Department of Physics, Chemistry and Biology, Biotechnology. Linköping University, Faculty of Science & Engineering. Univ Skovde, Sweden.
    Lifwergren, Philip
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Mandenius, Carl-Fredrik
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Selegård, Robert
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Aili, Daniel
    Linköping University, Department of Physics, Chemistry and Biology, Biophysics and bioengineering. Linköping University, Faculty of Science & Engineering.
    Dynamic peptide-folding mediated biofunctionalization and modulation of hydrogels for 4D bioprinting2020In: Biofabrication, ISSN 1758-5082, E-ISSN 1758-5090, Vol. 12, no 3, article id 035031Article in journal (Refereed)
    Abstract [en]

    Hydrogels are used in a wide range of biomedical applications, including three-dimensional (3D) cell culture, cell therapy and bioprinting. To enable processing using advanced additive fabrication techniques and to mimic the dynamic nature of the extracellular matrix (ECM), the properties of the hydrogels must be possible to tailor and change over time with high precision. The design of hydrogels that are both structurally and functionally dynamic, while providing necessary mechanical support is challenging using conventional synthesis techniques. Here, we show a modular and 3D printable hydrogel system that combines a robust but tunable covalent bioorthogonal cross-linking strategy with specific peptide-folding mediated interactions for dynamic modulation of cross-linking and functionalization. The hyaluronan-based hydrogels were covalently cross-linked by strain-promoted alkyne-azide cycloaddition using multi-arm poly(ethylene glycol). In addition, ade novodesigned helix-loop-helix peptide was conjugated to the hyaluronan backbone to enable specific peptide-folding modulation of cross-linking density and kinetics, and hydrogel functionality. An array of complementary peptides with different functionalities was developed and used as a toolbox for supramolecular tuning of cell-hydrogel interactions and for controlling enzyme-mediated biomineralization processes. The modular peptide system enabled dynamic modifications of the properties of 3D printed structures, demonstrating a novel route for design of more sophisticated bioinks for four-dimensional bioprinting.

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