Abstract 4336: Design of a chamber for studying glucose metabolism by anoxic cancer cells

Abstract

Abstract Many tumors contain anoxic areas. In vitro studies of anoxia can easily be hampered by contamination with traces of oxygen. In order to find out how temporal depletion of extracellular glucose effects anoxic cell death in vitro we have developed a chamber in which 50 000-500 000 cells can be exposed to approximately 30 μl of medium, resulting in rapid depletion of oxygen and nutrients. Before the experiments, cancer cells were seeded on a glass plate, which was the lower glass plate after clamping with an upper plate that contained an 80-100 μm thick silicone spacer. The medium of the cavity between the two flat glass plates could be flushed through an inlet and outlet hole in the upper plate, located in the corners of the cavity. Samples were collected from the outlet hole for analysis. Most cancer cell lines use both anaerobic and aerobic metabolism to generate ATP, resulting in a more rapid depletion of oxygen than of glucose. We found that after 3 hours the glucose was completely consumed by the MCF-7 human breast cancer cells and that overnight exposure resulted in cell death, except in the immediate proximity of the holes, where oxygen could diffuse into the medium. An interesting observation was that the lactate produced was more than double the glucose used from the medium, suggesting that also intracellular glycogen stores had been consumed by the cancer cells. We are especially interested in using this system to simulate the effect of dietary restriction on individual dormant breast tumors that have not (yet) started angiogenesis. These are hypothesized to contain an outer ring of anoxic cells, that depend on and rapidly consume the lowest concentrations of glucose “at the end of the glucose gradient” from the nearest capillary blood vessel. A temporary decrease of blood glucose may kill these anoxic cells and perhaps affect non-Warburg cells less. The time dependence of cell killing on this decrease will be investigated using this system. Citation Format: Jan Lankelma, Mohamed Ahmed, Adrià López Nadal, Josephine C. Dorsman, Hans Joenje, Hans V. Westerhoff. Design of a chamber for studying glucose metabolism by anoxic cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4336. doi:10.1158/1538-7445.AM2014-4336