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>High-throughput combinatorial analysis of three-dimensional biomaterials behavior using superhydrophobic patterned platforms =Análise combinatória expedita do comportamento de biomateriais tridimensionais usando plataforma superhidrofóbicas padronizadas
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High-throughput combinatorial analysis of three-dimensional biomaterials behavior using superhydrophobic patterned platforms =Análise combinatória expedita do comportamento de biomateriais tridimensionais usando plataforma superhidrofóbicas padronizadas
One of the still unaccomplished struggles in the maintenance of population life quality is related to the current need for effective biomaterials. The optimization of tissue engineering (TE) strategies by combining biomaterials, cells and soluble factors usually relies on time-consuming iterative processes. Rapid and low-cost high-throughput testing is needed to accelerate the discovery of ideal TE systems. The main hypothesis of this thesis was that superhydrophobic surfaces patterned with wettable spots were amenable to be used as platforms for high-throughput complete testing of 3D biomaterials. Indeed, such platforms allowed taking advantage of wettability contrast to pattern biomaterials with precise shape and pre-determined height, by controlling the volume dispensed in each spot. The superhydrophobic chips were first used to pattern ionic alginate-based cell-laden hydrogels in the wettable spots. The chemical composition of each biomaterial was evaluated by FTIR and the cellular response of fibroblast and osteoblast-like cell lines was assessed on-chip by image-based analysis. Image-based non-destructive assessment was validated by comparison with conventional biochemical colorimetric tests. Superhydrophobic chips were later used to produce and study miniaturized porous scaffolds. The size of the spots in the milimetric range allowed having porous biomaterial structures with significant pore size for cell migration and growth. Chitosan/alginate scaffolds were processed by polyelectrolyte complexation and freeze-drying, followed by fibronectin adsorption. Cell number and viability were assessed using two cell lines. DMA and muCT techniques were adapted to be used on-chip, in dry conditions, to characterize the scaffolds mechanically and morphologically. The on-chip DMA method was upgraded to be performed under physiological-like conditions using chitosan/bioactive glass nanoparticles hydrogels. The selective adhesion and proliferation of a pre-osteoblast cell line allowed hit-spotting favorable in vitro biomaterial formulations. After demonstrating their adequacy for in vitro cell-3D biomaterials interactions assessment, superhydrophobic chips containing 36 biomaterials were implanted in single Wistar rats, allowing the high-throughput in vivo study of inflammatory response caused by biomaterials. An important aspect in TE is the dependency of tissue regeneration on prolonged action of bioactive agents. Superhydrophobic chips were imprinted with ring-shaped spots with concentric superhydrophobic regions where polymeric protein-loaded spheres were deposited. The acquisition of sequential images of each spot over time using microscopy methods allowed monitoring protein release. Finally, cell suspension droplets were fixed in the wettable regions of the chips to produce cell spheroids/microtissues for drug screening by the hanging drop methodology in a robot-free automated manner. In conclusion, the superhydrophobic platforms patterned with wettable spots used in this thesis proved to be compatible with a complete study of 3D biomaterials-cells interactions, comprising a wide set of factors as biomaterials characterization, in vitro testing, innovative in vivo assessment and bioactive molecules-related tests.
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