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Many cells are cultured in media that contains an antibiotic to prevent bacterial contamination. Mycoplasma and other bacterial contamination is a serious problem for those involved in cell culture. Antibiotics in the media helps prevent this contamination and make life easier for the investigators; as performing cell culture experiments in antibiotic free media is difficult and requires vigorous sterile technique. There are many reports of antibiotics causing mitochondrial damage. In this study, we tested the effect of gentamicin in culture media on human mammary epithelial MCF-12A and breast cancer MCF-7 and MDA-MB-231 cell lines by real time PCR, immunofluorescent microscopy, lactate assay, DNA damage assay. We found that the addition of gentamicin in media upregulated the gene expression of hypoxia inducer factor 1 alpha (HIF1a), glycolytic enzymes and glucose transporters, compared to the cells cultured in gentamicin free media. Gentamicin also increased the lactate production and inhibited mitochondrial membrane potential of the cell lines. Furthermore, the antibiotics in media induced mitochondrial reactive oxygen species causing DNA damage. We found an increase of 8-hydroxy-2'-deoxyguanosine a product of DNA oxidative damage in the media of MCF-12A, MCF-7 and MDA-MB-231 cell lines. These results showed that normal epithelial and breast cancer cells cultured in the media with gentamicin had increased HIF1a, aerobic glycolysis and DNA oxidative damage. If we use these unhealthy cells in the experiment, all data will be different, compared to cells grown in gentamicin free media. We have studied the detrimental effects of three antibiotics on mitochondrial function in the untransformed MCF-12A human mammary cell line and two human mammary cancer cell lines, MCF-7 and MB-MDA-231. The metabolic changes in all cell lines were dramatically different between those in antibiotic free media versus antibiotic containing media. There was a marked difference in gene expression of glycolytic enzymes, reactive oxygen species production and effects on membrane potential. Ironically, our first studies were done in media containing gentamicin, and repeated studies were done in gentamicin free media. The results were very different. The purpose of this report is to emphasize that metabolic cell culture data may be inaccurate because experiments were performed in cell culture media containing antibiotics. We will present evidence to support this theory.
This article was published in the following journal.
Name: PloS one
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Antibiotic produced by Micromonospora inyoensis. It is closely related to gentamicin C1A, one of the components of the gentamicin complex (GENTAMICINS).
Liver disease lasting six months or more, caused by an adverse drug effect. The adverse effect may result from a direct toxic effect of a drug or metabolite, or an idiosyncratic response to a drug or metabolite.
Liver disease lasting six months or more, caused by an adverse effect of a drug or chemical. The adverse effect may be caused by drugs, drug metabolites, chemicals from the environment, or an idiosyncratic response.
The highest dosage administered that does not produce toxic effects. The NOAEL will depend on how closely dosages are spaced (lowest-observed-adverse-effect level and no-observed-effect level) and the number of animals examined. The ultimate objective is usually to determine not the "safe" dosage in laboratory animals but the "safe" dosage for humans. Therefore, the extrapolation most often required of toxicologists is from high-dosage studies in laboratory animals to low doses in humans. (Casarett and Doull's Toxicology: The Basic Science of Poisons, 4th ed)
An adverse effect occurring with a medical treatment that is not attributable to the actions of the treatment.
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