http://hdl.handle.net/10362/4555 Wed, 22 May 2013 12:06:59 GMT 2013-05-22T12:06:59Z http://hdl.handle.net/10362/8555 Title: Novel approaches for culturing hepatocytes for drug testing applications Authors: Leite, Sofia Batista Abstract: Drug Development is a long and costly process that starts with thousands of drug candidates and ends with one compound, which often fails in clinical trials. This high percentage of failure of the drug development process results from the lack of highly predictable models in the pre-clinical tests. Being metabolism a bottleneck in in vitro drug testing, special efforts have been made towards the development of metabolic competent liver cell cultures. The major drawback of standard hepatocyte cultivation systems is the spontaneous cell dedifferentiation that results in the decrease of metabolic competency and other liver specific functions and ultimately will compromise the effective biotransformation of drugs. Therefore, novel culture strategies presenting an improved hepatocyte viability and functionality for extended periods are required. Within this context, the main goal of this PhD project was to develop an efficient strategy for culturing hepatocytes as 3D spheroid structures since this better mimics the in vivo tissue. Within these 3D structures, cells can re-establish cell-cell interactions and specific microenvironments resulting in a better retention of important hepatic functions. The work here depicted has focused on the use of primary cultures of hepatocytes, since they better retain the liver functional profile. Additionally, the developed strategy has been applied to a human hepatocarcinoma cell line with the aim of generating a more physiological model with cells that are easier to obtain then fresh isolated hepatocytes. Within this context, and also to confirm the robustness of the 3D culturing strategy, three relevant cellular models were used namely, human hepatocytes, rat hepatocytes and the HepaRG cell line.(...) Description: Dissertation presented to obtain the Ph.D degree in Biochemisry, Biotechnology Thu, 01 Mar 2012 00:00:00 GMT http://hdl.handle.net/10362/8555 2012-03-01T00:00:00Z http://hdl.handle.net/10362/6849 Title: Metabolic flux analysis of neural cell metabolism in primary cultures Authors: Amaral, Ana Isabel Porém Abstract: Brain energy metabolism results from a complex group of pathways and trafficking mechanisms between all cellular components in the brain, and importantly provides the energy for sustaining most brain functions. In recent decades, 13C nuclear magnetic resonance (NMR) spectroscopy and metabolic modelling tools allowed quantifying the main cerebral metabolic fluxes in vitro and in vivo. These investigations contributed significantly to elucidate neuro-glial metabolic interactions, cerebral metabolic compartmentation and the individual contribution of neurons and astrocytes to brain energetics. However, many issues in this field remain unclear and/or under debate. Description: Dissertation presented to obtain the Ph.D degree in Biochemistry, Neuroscience Sat, 01 Oct 2011 00:00:00 GMT http://hdl.handle.net/10362/6849 2011-10-01T00:00:00Z http://hdl.handle.net/10362/6176 Title: Systems biotechnology of baculovirus-producing insect cells Authors: Carinhas, Nuno Abstract: critical step in the biopharmaceutical industry is the capacity to properly manufacture biological products in animal cell cultures. The ability to exploit the full potential of the underlying “cellular factories” is essential to reduce manufacturing costs and improve yield and product quality, thus freeing up resources to pursue other added value activities, such as clinical development and pharmacovigilance of new products. Bioprocess optimization efforts have mostly relied on empirical experience and rational, hypothesis-driven studies, rather than by fully leveraging the knowledge of how complex cells behave in culture. Despite the vast amount of information available on different cellular features, specific productivity improvements have been essentially incremental. The relatively recent developments in Systems Biology and computational methodologies present a paradigm-shifting opportunity to develop biotechnology processes with high efficiencies in a consistent, systematic way. (...) Description: Dissertation presented to obtain the Ph.D degree in Ciências da Engenharia e Tecnologia, especialidade Biotecnologia Thu, 01 Sep 2011 00:00:00 GMT http://hdl.handle.net/10362/6176 2011-09-01T00:00:00Z http://hdl.handle.net/10362/5770 Title: Expression of antibodies and retroviralVectors from defined chromosomal sites: strategies towards reliable production systems Authors: Norton, Leonor da Gama Carvalho Abstract: Technologies enabling expression of recombinant genes in mammalian cells belong to the tools of the trade in molecular biology. These technologies can serve a wide variety of purposes, ranging from basic research to elucidate gene functions e.g., in transgenic mouse models, to biotechnological applications such as manufacturing antibodies or producing viral vectors. In cell line development, the integration of transgenes into the chromosomal DNA of the host cell is a crucial step, since chromosomal surroundings have a major impact on the expression of the transgene. With regard to safety, product consistency and operational transparency, expression of biotechnological relevant products from single copy DNA integrants is preferable over a scenario of multi‐gene integrants.(...) Description: Dissertation presented to obtain a Ph.D. degree in Sciences of Engineering and Technology, Cell Technology, at the Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa Tue, 01 Jun 2010 00:00:00 GMT http://hdl.handle.net/10362/5770 2010-06-01T00:00:00Z