Lactate Metabolism and the Response to Nanoparticle Exposure in Normal and Cancerous Breast Cells

To understand lactate metabolism and the response of normal and cancerous breast cells to nanoparticle exposure, we performed two studies using breast cancer cell lines MCF-7 and MDA-MB-231, and a normal breast cell line HMEC 184./nIn my first study I examined the expression and the localization of the lactate shuttle proteins monocarboxylate transporter (MCT) and lactate dehydrogenase (LDH) isoforms in two breast cancer cell lines MCF-7, MDA-MB-231, compared to the normal breast cell line HMEC 184. I hypothesized that there are changes in the localization and expression of MCTs and LDH isoforms in cancerous breast cells when compared to normal breast cells, and that these changes are associated with the Warburg Effect and correspond to the oxidative capacity of the cancerous cells. My data show that MCT (1, 2, and 4), and LDH isoforms (A and B) are expressed in both normal and cancerous breast cell lines, except that MDA-MB-231 did not express MCT1. MCT1 was highly expressed in normal cells when compared to cancer cell lines. MCT4 was highly expressed in MDA-MB-231, and MCT2 was highly expressed in MCF-7. LDH was highly expressed in both cancerous cell lines compared to the normal cell line, and MCF-7 expressed mainly LDH5 (LDHB), while MDA-MB-231 and HMEC 184 expressed mainly LDH1 (LDHA). Using confocal laser scanning microscopy, I found that MCT2, MCT4, and LDH are localized in mitochondria in addition to their localization in the plasma membrane and cytosol, whereas MCT1 is mainly localized in the plasma membrane. This localization was the same in cancerous and normal cell lines. The changes in the expression of MCT and LDH isoforms corresponded to the metabolic status of each cell line. Both cell lines MCF-7 and MDA-MB-231 produced higher amounts of lactate than the HMEC 184 cell line, but have less endogenous and maximum respiration than the HMEC 184 cell line. In conclusion, I reported changes in the expression of MCT and LDH in breast cancer cells with no change in their localization. These changes corresponded to the breast cancer cells' oxidative capacity. My data support the existence of the previously reported lactate shuttle in cancer, and add a new explanation of its function./nMy next project examined the effect of co-polymer nanoparticles, Eudragit® RS 100 (ENPs), increasingly being used to coat and deliver drugs including chemotherapy agents, on the metabolic activity and proliferation of the human epithelial breast cells (HMEC 184, MCF-7, MDA-MB-231). I reported novel responses of human epithelial breast cells when exposed to Eudragit nanoparticles. I showed that cells displayed dose-dependent increases in metabolic activity and growth, but lower proliferation rates, than control cells, as evidenced in tetrazolium salt (WST-1) and 5-bromo-2'-deoxyuridine (BrdU) assays. Using mass spectrometry and micrroarry analyses I found that the mechanism for this behaviour stems from the ability of Eudragit nanoparticles to bind to certain proteins in culture media and to bring them closer to the surface of cells. Those proteins are involved in cell adhesion, growth, differentiation, and migration. The effect of nanoparticle treatment in increasing cancer and normal human epithelial breast cell metabolic activity and growth has not been reported previously, and this project highlighted the need for further research to address the potentially counter-productive effects of using nanoparticles in cancer chemotherapy.

Advisor: Brooks, George A