Publicação
Microengineered scaffolds for three- dimensional cell culture applications
| Resumo: | Tissue engineering is a vast field that arose to fight several flaws inherent to transplants, replacement of damaged tissues, and generalized therapies. In this context, a cell supporting structure, called scaffold, plays a major role in recreating biological microenvironments. Thus, the design and manufacture of threedimensional (3D) scaffolds for cellular interaction studies are required to create complex architectures capable of mimicking specific tissues. An interesting technique for the development of 3D structures is the two-photon polymerization (2PP) method. This nonlinear process induces polymerization of a photosensitive material within extremely small volumes and allow the fabrication of complex architectures with a minimum feature of 200 nm. Resorting to three different 2PP setups, four materials with different stiffness are tested in the manufacture of 3D structures, including the IP-DIP and IP-S resins, the SZ2080 polymer, and the PEGDA 700 hydrogel. The chosen architectures follow a woodpile design, where pores closely match the size of specific cell lines. Various processing parameters, such as laser power, writing speed, and solvent evaporation, are optimized in order to guarantee mechanical stability and proper adhesion to the substrate. The strong intrinsic autofluorescence of the SZ2080 material, which can become problematic during cellular imaging assays, is reduced by up to 90 % with ultraviolet post-treatments. A nanoindenting machine auxiliaries the mechanical characterization of PEGDA 700 pedestals printed with different writing parameters. Preliminary results show a tendency of stiffness increase with extended laser exposure. The IP-S, SZ2080 and PEGDA 700 scaffolds’ biocompatibility is assessed with HeLa and bone marrow mesenchymal stem cell (BM-MSCs) lines, for their relevance in medicine. HeLa cells allow the in-depth study of cervical cancers, and BM-MSCs play key roles in several hematological mechanisms and associated diseases. The interactions between cell lines and developed 3D scaffolds reveal similar optimistic preliminary outcomes in terms of growth, adhesion, and proliferation, regardless the material used. Finally, SZ2080 scaffolds are integrated into culture chambers of microfluidic polydimethylsiloxane (PDMS) devices to allow dynamic cell culture under controlled conditions. After proper optimization, the final device allows the controlled circulation of fluids through the microchannels and culture chambers. Thus, it is demonstrated the potential of these 3D combined platforms, woodpile scaffolds and microfluidic technology, for further 3D cell culture assays in more biomimetic microenvironments with great prospective for developing disease models and more detailed studies of cellular behaviors. |
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| Autores principais: | Costa, Beatriz Neves Leal |
| Assunto: | Two-photon polymerization Three-dimensional scaffolds Cell culture Microfluidic Polimerização por dois fotões Scaffolds tridimensionais Cultivo celular Microfluídica |
| Ano: | 2021 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | Tissue engineering is a vast field that arose to fight several flaws inherent to transplants, replacement of damaged tissues, and generalized therapies. In this context, a cell supporting structure, called scaffold, plays a major role in recreating biological microenvironments. Thus, the design and manufacture of threedimensional (3D) scaffolds for cellular interaction studies are required to create complex architectures capable of mimicking specific tissues. An interesting technique for the development of 3D structures is the two-photon polymerization (2PP) method. This nonlinear process induces polymerization of a photosensitive material within extremely small volumes and allow the fabrication of complex architectures with a minimum feature of 200 nm. Resorting to three different 2PP setups, four materials with different stiffness are tested in the manufacture of 3D structures, including the IP-DIP and IP-S resins, the SZ2080 polymer, and the PEGDA 700 hydrogel. The chosen architectures follow a woodpile design, where pores closely match the size of specific cell lines. Various processing parameters, such as laser power, writing speed, and solvent evaporation, are optimized in order to guarantee mechanical stability and proper adhesion to the substrate. The strong intrinsic autofluorescence of the SZ2080 material, which can become problematic during cellular imaging assays, is reduced by up to 90 % with ultraviolet post-treatments. A nanoindenting machine auxiliaries the mechanical characterization of PEGDA 700 pedestals printed with different writing parameters. Preliminary results show a tendency of stiffness increase with extended laser exposure. The IP-S, SZ2080 and PEGDA 700 scaffolds’ biocompatibility is assessed with HeLa and bone marrow mesenchymal stem cell (BM-MSCs) lines, for their relevance in medicine. HeLa cells allow the in-depth study of cervical cancers, and BM-MSCs play key roles in several hematological mechanisms and associated diseases. The interactions between cell lines and developed 3D scaffolds reveal similar optimistic preliminary outcomes in terms of growth, adhesion, and proliferation, regardless the material used. Finally, SZ2080 scaffolds are integrated into culture chambers of microfluidic polydimethylsiloxane (PDMS) devices to allow dynamic cell culture under controlled conditions. After proper optimization, the final device allows the controlled circulation of fluids through the microchannels and culture chambers. Thus, it is demonstrated the potential of these 3D combined platforms, woodpile scaffolds and microfluidic technology, for further 3D cell culture assays in more biomimetic microenvironments with great prospective for developing disease models and more detailed studies of cellular behaviors. |
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