Abstract
In vitro systems that can effectively model liver function for long periods of time are fundamental tools for preclinical research. Nevertheless, the adoption of in vitro research tools at the earliest stages of drug development has been hampered by the lack of culture systems that offer the robustness, scalability, and flexibility necessary to meet industry’s demands. Bioreactor-based technologies, such as stirred tank bioreactors, constitute a feasible approach to aggregate hepatic cells and maintain long-term three-dimensional cultures. These three-dimensional cultures sustain the polarity, differentiated phenotype, and metabolic performance of human hepatocytes. Culture in computer-controlled stirred tank bioreactors allows the maintenance of physiological conditions, such as pH, dissolved oxygen, and temperature, with minimal fluctuations. Moreover, by operating in perfusion mode, gradients of soluble factors and metabolic by-products can be established, aiming at resembling the in vivo microenvironment. This chapter provides a protocol for the aggregation and culture of hepatocyte spheroids in stirred tank bioreactors by applying perfusion mode for the long-term culture of human hepatocytes. This in vitro culture system is compatible with feeding high-throughput screening platforms for the assessment of drug elimination pathways, being a useful tool for toxicology research and drug development in the preclinical phase.
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Acknowledgements
This work was financially supported by the grants PTDC/EBB-BIO/112786/2009 and SFRH/BD/70264/2010 provided by Fundação para a Ciência e Tecnologia, Portugal. The authors would like to acknowledge Janne Jensen and Peter Bjorquist from Cellartis and Rui Tostões, Sofia Leite, and Margarida Serra for implementation of the aggregation of primary cultures of human hepatocytes at iBET. The authors also acknowledge João Clemente for the implementation and support on the perfusion operation mode in the bioreactors.
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Rebelo, S.P., Costa, R., Sousa, M.F.Q., Brito, C., Alves, P.M. (2015). Establishing Liver Bioreactors for In Vitro Research. In: Vinken, M., Rogiers, V. (eds) Protocols in In Vitro Hepatocyte Research. Methods in Molecular Biology, vol 1250. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2074-7_13
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DOI: https://doi.org/10.1007/978-1-4939-2074-7_13
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