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Browsing by Subject "3D cell culture"

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  • Koskinen, Elisabeth (2023)
    Three-dimensional (3D) cellular cultures have been shown to represent tissue formations and functions more accurately than two-dimensional (2D) cultures and have been successfully utilized more accurately in model organisms, e.g., to understand cellular modular functions. However, the applications in non-model organisms are limited, and to our knowledge have not been implemented in ectotherms. At an ecological scale, the technique can enhance our understanding by providing insights on cellular and tissue level molecular mechanisms. A potential implementation of this method in Atlantic salmon is to elucidate the molecular function of the vestigial-like 3 (vgll3) gene, which plays a central role in salmonid maturity, potentially by regulating energy allocation via regulating adipogenesis. In this thesis, a proof of principle study was implemented, where the feasibility of obtaining and maintaining a suspension 3D adipose tissue culture in Atlantic salmon (Salmo salar) was assessed. The harvested visceral white adipose tissue from around the intestinal tissue of mature Atlantic salmon (salmon past smolt stage) was first separated into stromal vascular fraction (SVFs) and mature adipose fraction (MAFs). SVFs contain preadipocytes (precursors of adipocyte cells) in addition to a variety of other cell types. MAFs are mature adipocytes. Both MAFs and SVFs were successfully maintained in-vitro for over four weeks. SVFs were then successfully differentiated into mature adipocytes, demonstrating the feasibility of studying adipogenesis in Atlantic salmon. Proof of this methodology and its further implications may help us to understand the cellular functions of vgll3 and may subsequently help to better understand its causal relation to the maturation process in Atlantic salmon.
  • Heilala, Maria (2019)
    Despite the advances in the management of breast cancer, discovery of novel and targeted drugs remains a challenge. It has been suggested that drug failure rates in clinical trials might be diminished by improving the predictive potential of preclinical cancer models. Three-dimensional (3D) scaffold-based cell culture has emerged as an attractive platform for mimicking tissue-like microenvironment, since it is well-known that cells respond to the cues in the extracellular matrix (ECM). The aim of this thesis was to develop fibrin-based hydrogels and evaluate their performance in 3D cell culture of breast cancer cells. The fibrin gel formulation was first optimized by testing the effect of different buffers on gel properties. Structural properties were examined with scanning electron microscopy and mechanical properties measured with oscillatory rheometry. Three different fibrin concentrations of the optimized formulation were then used as scaffolds for DU4475 breast cancer cells. After seven days of culture, the morphology, phenotype and proliferation of the resulting cell structures were assessed by using techniques such as light microscopy, immunofluorescent confocal microscopy and Western blot analysis. The desired properties for 3D cell culture were obtained by preparing fibrin gels at high pH in the absence of calcium. The main finding of the thesis was that fibrin concentration strongly affected the phenotype of DU4475 cells, with cells cultured in the softest gel retaining their original characteristics to the greatest extent. In the future, the developed scaffold could possibly be used in drug discovery and personalized medicine by culturing tumor explants from patients. However, the methods used in the study must be further optimized and the results validated with other breast cancer cell lines and with primary tissues.