Evaluation of ELP, collagen and laminin hydrogel composites to support in vitro differentiation of hADSCs toward adipocytes and β-cells

Examensarbete för masterexamen

Please use this identifier to cite or link to this item: https://hdl.handle.net/20.500.12380/217875
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Type: Examensarbete för masterexamen
Master Thesis
Title: Evaluation of ELP, collagen and laminin hydrogel composites to support in vitro differentiation of hADSCs toward adipocytes and β-cells
Authors: Åström, Maria
Dahlén, Robin
Abstract: Drug discovery typically involves studies on animal models as proxies for human beings followed by three phases of clinical studies, which result in frequent late rejections due to interspecies differences, present moral issues and requires extensive administration. Hence, large efforts are made to develop in vitro drug screening assays based on human cells, which provide detailed information on intracellular processes and enable high-throughput analysis of multiple drugs in parallel. Assays mimicking metabolic diseases such as diabetes and obesity are of particular interest, due to their increasing prevalence and due to important interspecies differences in metabolic pathways. However, it becomes increasingly clear that cells grown in vitro on conventional flat surfaces have difficulties maintaining their native, mature functionalities and eventually develop artificial phenotypes. For that reason, we have evaluated tissue-mimicking, protein-composite hydrogels for three-dimensional growth and differentiation of human adipocytes-derived stem cells (hADSCs) toward (i) adipocytes and (ii) β-cells. The proteins collagen I, laminin isoforms 411 and 511 and recombinant-engineered elastin-like proteins (ELP) have been investigated in detail, individually as well as in mixes. Their different amino-acid sequences coding for cell attachment/interaction, attractive material properties (i.e. soft and hydrated) together with the three-dimensional encapsulation are intended to mimic the native microenviroment, and thereby drive the differentiation and cellular maturation. The metabolic activity, viability and function of the differentiated cells were analyzed with cellular assays. Further, the tissue-mimicking samples were evaluated with non-linear microscopy; multi-photon excitation fluorescence (MPEF, autofluorescence and stains), coherent anti-Stokes Raman scattering (CARS, cellular lipid content) and second harmonic generation (SHG, collagen fibers). The assays and MPEF microscopy revealed that cells do not thrive in the ELP hydrogels crosslinked with amine-reactive THPC, probably due to cytotoxicity of the crosslinker and as the hydrogel seems not to permit cellular expansion. In contrast, cells in collagen and collagen/ELP matrices were successfully differentiated into adipocyte-like cells, confirmed by the presence of excessive amounts of lipid stores co-localized with perilipin. Insulin-positive cells were found in the collagen and collagen/laminin matrices. Hence, different collagen mixes seem to be promising hydrogel materials for three-dimensional growth of hADSCs for metabolic drug screening.
Keywords: Livsvetenskaper;Biokemi och molekylärbiologi;Molekylärbiologi;Medicinsk bioteknologi (med inriktning mot cellbiologi);Biomedicinsk laboratorievetenskap/teknologi;Life Science;Biochemistry and Molecular Biology;Molecular biology;Medical Biotechnology (with a focus on Cell Biology);Biomedical Laboratory Science/Technology
Issue Date: 2015
Publisher: Chalmers tekniska högskola / Institutionen för biologi och bioteknik, Kemisk biologi
Chalmers University of Technology / Department of Biology and Biological Engineering, Chemical Biology
URI: https://hdl.handle.net/20.500.12380/217875
Collection:Examensarbeten för masterexamen // Master Theses



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