Abstract
Cancer cell culture is routinely performed under superphysiologic O2 levels and in media such as Dulbecco’s Modified Eagle Medium (DMEM) with nutrient composition dissimilar to mammalian extracellular fluid. Recently developed cell culture media (e.g., Plasmax, Human Plasma-Like Medium (HPLM)), which are modeled on the metabolite composition of human blood plasma, have been shown to shift key cellular activities in several cancer cell lines. Similar effects have been reported with respect to O2 levels in cell culture. Given these observations, we investigated how media composition and O2 levels affect cellular energy metabolism and mitochondria network structure in MCF7, SaOS2, LNCaP, and Huh7 cells. Cells were cultured in physiologic (5%) or standard (18%) O2 levels, and in physiologic (Plasmax) or standard cell culture media (DMEM). We show that both O2 levels and media composition significantly affect mitochondrial abundance and network structure, concomitantly with changes in cellular bioenergetics. Extracellular acidification rate (ECAR), a proxy for glycolytic activity, was generally higher in cells cultured in DMEM while oxygen consumption rates (OCR) were lower. This effect of media on energy metabolism is an important consideration for the study of cancer drugs that target aspects of energy metabolism, including lactate dehydrogenase activity.
Highlights
Standard cell culture procedures originating in the mid-20th century remain widely used today for investigating cell biology, including toxicity testing, and drug development.it has become increasingly clear that media composition, including oxygen, affects many aspects of cell biology, including metabolism and drug efficacy [1,2,3]
Consistent with that, we have shown that the concentration of even one single constituent in culture medium significantly affected the effects of two wellcharacterized anti-cancer molecules: resveratrol and rapamycin on multiple human cancer cell lines (e.g., LNCaP, Huh7) [5]
Cells were cultured for a minimum of two weeks to acclimatize them to one of four different cell culture conditions: (1) the most common condition used for cancer cell culture, i.e., high-glucose Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and unregulated O2 ( ~18%); (2) this same medium but with O2 regulated at 5%; (3) the Plasmax formulation of VandeVoorde et al [4]; (4) this same medium but with O2 regulated at 5%
Summary
Standard cell culture procedures originating in the mid-20th century remain widely used today for investigating cell biology, including toxicity testing, and drug development. The metabolic profile of TNBC spheroids grown in Plasmax for only four days resembled the metabolic landscape of orthotopic xenografts more closely than those in RPMI [4] These results demonstrate that physiologic media such as HPLM and Plasmax alter cellular metabolism and the response to specific drugs. Cells were cultured for a minimum of two weeks to acclimatize them to one of four different cell culture conditions: (1) the most common condition used for cancer cell culture, i.e., high-glucose DMEM supplemented with 10% FBS and unregulated O2 ( ~18%); (2) this same medium but with O2 regulated at 5%; (3) the Plasmax formulation of VandeVoorde et al [4]; (4) this same medium but with O2 regulated at 5% Under these four conditions, we evaluated cellular bioenergetics, mitochondrial abundance, and mitochondrial network morphology in all four cell lines. These results indicate the importance of considering culture conditions in studies of energy metabolism and mitochondrial function in vitro
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