Abstract 1695: Quantification of mitochondria in MCF-10A, MDA-MB-231, and SUM149 cells to understand potential defects in oxidative phosphorylation in cancer.
Authors: Ashrafzadeh Sepideh , Van Wassenhove Lauren D. , Merajver Sofia D. 
Published: August 2014

Abstract
Cancer cells are known to exhibit altered metabolism. The Warburg effect, whereby cancer cells preferentially use glycolysis instead of oxidative phosphorylation to produce ATP, plays an important role in the tumorigenic capacity of breast cancer. Previously, we found that SUM149 (inflammatory breast cancer) and MDA-MB-231 (invasive breast cancer) cells have decreased oxidative phosphorylation. Since mitochondria are the powerhouses of oxidative phosphorylation, examining their concentrations, phenotypes, and the distribution of their key enzymes can provide crucial information. These data can be used to further clarify whether the observed metabolic discrepancies stem from abundance differences or from other sources, such as defective enzymes and/or intermediate participation in alternate metabolic pathways. The purpose of our research was to elucidate if differential mitochondria concentrations accounted for the observed marked decrease in oxidative phosphorylation in cancer cells. Specifically, we focused on quantifying the number of mitochondria in two aggressive cancer cell lines, MDA-MB-231 and SUM149, and a normal-like breast cell line, MCF-10A.
In order to measure the number of mitochondria in the cancerous and noncancerous cells, a commercially available antibody conjugated to green fluorescent protein (GFP) and directed against the E1-alpha subunit of pyruvate dehydrogenase was used in each cell line. The resulting cells were imaged under high-resolution confocal microscopy and their relative fluorescence assessed. The average intensity of GFP in each cell was calculated and used as an indicator for the amount of mitochondria in each cell line. Additionally, these results were compared to transmission electron microscopy (TEM) images of each of these cell lines to validate the immunofluorescent quantifications.
Our results showed that there is no significant difference among the number of mitochondria in MDA-MB-231, SUM149, and MCF-10A cells. Additionally, the distribution of mitochondria among all three cell lines was relatively uniform, with a slightly greater abundance of mitochondria around the nuclei. These findings are surprising given that the published research on the Warburg effect, as well as our own prior observations on these three specific cell lines, suggested that cancer cells may have different numbers of mitochondria, although the previous work did not offer definitive proof of this interpretation. Therefore, these results contradict that hypothesis, and direct further studies to identify qualitative functional defects within mitochondria such as mutated or silenced mitochondrial genes, and to explore alternate pathways that breast cancer cells use to derive energy. Our investigation highlights the importance of taking a multi-facetted approach to studying cancer cell metabolism.
Citation Format: Sepideh Ashrafzadeh, Lauren D. Van Wassenhove, Sofia D. Merajver. Quantification of mitochondria in MCF-10A, MDA-MB-231, and SUM149 cells to understand potential defects in oxidative phosphorylation in cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1695. doi:10.1158/1538-7445.AM2013-1695