Using a Systems Approach to Investigate the Impact of in vitro culture conditions on Cellular Energetics and Biochemistry

Mitochondria are critical to the normal biochemical function of most mammalian cells in vivo. However, when cells are cultured in vitro there is often a fundamental shift in their dependence on mitochondria. Also, cellular and mitochondrial function is dramatically influenced by in vitro culture conditions which are not static and change over time as cells grow and respire in a fixed body of medium.

This dynamic interplay between culture conditions, mitochondrial function and biochemistry has profound implications for commonly used in vitro cell systems but is poorly understood. Two scenarios where this is of clear significance to the pharmaceutical industry are 1) the use of in vitro cultures to study mechanisms of drug action on mitochondrial function (for both toxicological and pharmacological applications) and 2) the use of transfected cells for the commercial production of proteins, eg bioengineered therapeutic antibodies.

This collaborative project will utilise expertise within the groups of Dr Karl Morten (Nuffield Department of Obstetrics and Gynaecology) and Associate Professor James McCullagh (Department of Chemistry) at the University of Oxford and GSK's Biopharmaceutical and Toxicology departments to develop fundamental understanding of how mitochondrial function and biochemistry are modified by culture conditions in vitro, and how this might be manipulated to improve cell models for drug screening (to avoid toxicity and discover potential new drugs) and to improve the production of therapeutic antibodies.

Mitochondrial biochemistry involves many interconnected pathways which feedback and compensate each other, producing a complex metabolic web. Given this complexity, we plan to integrate data from multiple different assay platforms; for example, Seahorse cellular respirometry data (and other key mitochondrial parameters) will be used to provide a detailed view of cellular respiration and mitochondrial function over time. Culture media samples and cell pellets will be analysed for biochemical changes using metabolomics and label free mass spectrometry. Adaptive changes in gene expression will be monitored by transcriptomic analysis of the cells. A highly novel perfused cell culture system being developed in Oxford may also be of value. Finally, the analysis would be supplemented with state of the art in silico modelling. These "virtual mitochondrion" computer models include hundreds of embedded biochemical reactions and enable us to simulate the effects of changing cellular or biochemical conditions on mitochondrial function, andvice versa.

The overall aim of this studentship is to provide significant new insights into the function and biochemistry of mitochondria in commonly used cell systems and to highlight how we might manipulate these to impact key cellular processes in vitro, such as recombinant protein production and adaptive stress responses (eg. when challenged with developmental drug). This in turn will enhance the value of in vitro systems for our specific applications; investigating mitochondria-drug interactions or improving commercial therapeutic antibody production.