Effects of Hypoxia on Cell Growth and Metabolism in the Leukemia Cell Line JJN-3 and Bladder Cancer Cell Line UM-UC-3

Abstract

Hypoxia is a condition in which the oxygen (O2) access is extremely reduced and thus limits the O2 available for cellular respiration. The hypoxic condition induces metabolic adjustments in the cells that permit further cell survival and growth. In this study, effects of hypoxia on cell growth and metabolism were studied in the human leukemia cell line JJN-3 and the bladder cancer cell line UM-UC-3. Initial studies showed an effect of hypoxia within 24 h of incubation with faster initial cell growth, altered glucose/glutamine uptake and lactate excretion fluxes, and altered cell cycle progression in both cell lines. Especially interesting was the increased glutamine consumption in UM-UC-3 24 h after hypoxic incubation, which was the opposite from the decreased glutamine consumption in JJN-3. A HIF-1 assay was used to confirm that the experimental conditions were hypoxic. HIF-1, a transcription factor involved in hypoxic adaptation, reached maximum level in JJN-3 8 h after incubation, indicating its role in the immediate hypoxic response. The second part of the project involved the study of metabolic adaptation to hypoxia in JJN-3 before, at and after the HIF-1 peak. Cells incubated in hypoxic conditions had increased glucose consumption and lactate production the first 24 h. The glutamine consumption was strongly reduced after the HIF-1 peak. The hypoxic effect on the endometabolome was studied using target mass spectrometric (MS) metabolic profiling methods (capIC-MS/MS and LC-MS/MS). Metabolite pools of the glycolysis, the PPP and TCA cycle, as well as the nucleoside phosphate pool, were all strongly increased immediately after hypoxic incubation. Following this immediate hypoxic response, the metabolite pools were adjusted to a level closer to the normoxic at and after the HIF-1 peaked. The exception was -ketoglutarate (KG) which was strongly reduced in the hypoxic cells at all sampling points. Altogether, the findings show that the cells have an immediate response to hypoxia with increased metabolism before it is adjusted to a level more similar to the normoxic state when the cells are adapting to the hypoxic conditions. Hypoxia seems to induce some different adjustments in JJN-3 and UM-UC-3, which makes it of interest to study metabolic adaptation at the endometabolome level of UM-UC-3 as well.