Browsing Ph.D. Biology by Subject "Cell culture, Mitochondria, Metabolism"
Now showing items 1-1 of 1
Investigating the Effect of Cell Culture Compositions on Mitochondrial Metabolism, Dynamics, and Transcriptome and Proteome of cellsThe phytoestrogen Resveratrol (RES) is a natural polyphenol that has been detected in more than 70 plant species. RES has structural similarity to mammalian estrogens and can bind to estrogen receptors, eliciting genomic and non-genomic effects. Both RES and physiological estrogens like 17-β-estradiol (E2) have wide-ranging effects on mitochondria. In this thesis, I began by investigating RES’s effects on mitochondrial network dynamics (Chapter 2) and discovered a pro-fusion activity apparently mediated by the mitofusin enzyme Mfn2. RES stimulated mitochondrial network hyper-fusion morphology in all three cell lines investigated (C2C12 (mouse myoblast), PC3 (prostate cancer), and MEFs (mouse embryonic fibroblast)), but the effect was absent in Mfn2-null MEFs. As this work was being completed; several research groups introduced ‘physiologic cell culture media’ that are modeled on the human plasma metabolome. I co-authored a study (not in this thesis) demonstrating that RES’s effects on mitochondrial dynamics are dependent on cell culture conditions. To follow up on this, I investigated whether E2’s mitochondrial effects might also be dependent on the cell culture environment, and showed conclusively that this is indeed the case, using C2C12 cells as a model system (Chapter 3). These results and those published by others in 2017-2019 suggested that medium composition can profoundly affect cellular functions. In Chapter 4, I followed this up by studying how culture conditions affect mitochondrial bioenergetics and network morphology using four cancer cell lines and showed that this is a significant issue. Finally, to gain a more complete understanding of this phenomenon, I completed a full transcriptomic and proteomic analysis of media effects using MCF7 breast cancer cells as a model (Chapter 5). I showed that hundreds of transcripts and proteins are affected according to culture conditions. Taken together, the results presented in this thesis emphasize the significant extent to which the cell culture environment affects experimental outcomes, particularly with respect to mitochondrial bioenergetics and dynamics. This information contributes to the development of cell culture experiments providing results that can be translated in vivo.