Mycoplasma contamination in human leukemia cell lines

Uphoff, Cord C.; Gignac, Suzanne M.; Drexler, Hans G.

doi:10.1016/s0022-1759(12)80047-0

Cited by 87 publications

(38 citation statements)

References 10 publications

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“…Cells were confirmed to be mycoplasma-free by 4′,6-diamidino-2-phenylindole (DAPI) method [33]. Molecular identity was confirmed by short tandem repeat analysis [34].…”

Section: Methodsmentioning

confidence: 99%

Therapeutic potential for phenytoin: targeting Nav1.5 sodium channels to reduce migration and invasion in metastatic breast cancer

Yang

Kozminski

Wold

et al. 2012

Breast Cancer Res Treat

177

223

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Voltage-gated Na+ channels (VGSCs) are heteromeric membrane protein complexes containing pore-forming α subunits and smaller, non-pore-forming β subunits. VGSCs are classically expressed in excitable cells, including neurons and muscle cells, where they mediate action potential firing, neurite outgrowth, pathfinding, and migration. VGSCs are also expressed in metastatic cells from a number of cancers. The Nav1.5 α subunit (encoded by SCN5A) is expressed in breast cancer (BCa) cell lines, where it enhances migration and invasion. We studied the expression of SCN5A in BCa array data, and tested the effect of the VGSC-blocking anticonvulsant phenytoin (5,5-diphenylhydantoin) on Na+ current, migration, and invasion in BCa cells. SCN5A was up-regulated in BCa samples in several datasets, and was more highly expressed in samples from patients who had a recurrence, metastasis, or died within 5 years. SCN5A was also overexpressed as an outlier in a subset of samples, and associated with increased odds of developing metastasis. Phenytoin inhibited transient and persistent Na+ current recorded from strongly metastatic MDA-MB-231 cells, and this effect was more potent at depolarized holding voltages. It may thus be an effective VGSC-blocking drug in cancer cells, which typically have depolarized membrane potentials. At a concentration within the therapeutic range used to treat epilepsy, phenytoin significantly inhibited the migration and invasion of MDA-MB-231 cells, but had no effect on weakly metastatic MCF-7 cells, which do not express Na+ currents. We conclude that phenytoin suppresses Na+ current in VGSC-expressing metastatic BCa cells, thus inhibiting VGSC-dependent migration and invasion. Together, our data support the hypothesis that SCN5A is up-regulated in BCa, favoring an invasive/metastatic phenotype. We therefore propose that repurposing existing VGSC-blocking therapeutic drugs should be further investigated as a potential new strategy to improve patient outcomes in metastatic BCa.

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“…Cells were confirmed to be mycoplasma-free by 4′,6-diamidino-2-phenylindole (DAPI) method [33]. Molecular identity was confirmed by short tandem repeat analysis [34].…”

Section: Methodsmentioning

confidence: 99%

Therapeutic potential for phenytoin: targeting Nav1.5 sodium channels to reduce migration and invasion in metastatic breast cancer

Yang

Kozminski

Wold

et al. 2012

Breast Cancer Res Treat

177

223

View full text Add to dashboard Cite

show abstract

“…For both biological and ecological reasons, it is important to eliminate these agents from cell cultures used for basic research, diagnosis, and biotechnological production. The most effective procedure for eliminating, inactivating, or suppressing mycoplasmas in cell cultures is treatment with antibiotics (6,22,27). In general, antibiotic therapies do not result in longlasting successful decontamination, and undesirable side effects on eukaryotic cells due to cytotoxic effects and the development of resistant mycoplasma strains have been observed (22,27).…”

mentioning

confidence: 99%

Antimycoplasma properties and application in cell culture of surfactin, a lipopeptide antibiotic from Bacillus subtilis

Vollenbroich

Pauli

Özel

et al. 1997

Appl Environ Microbiol

223

107

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Surfactin, a cyclic lipopeptide antibiotic and biosurfactant produced by Bacillus subtilis, is well-known for its interactions with artificial and biomembrane systems (e.g., bacterial protoplasts or enveloped viruses). To assess the applicability of this antiviral and antibacterial drug, we determined the cytotoxicity of surfactin with a 50% cytotoxic concentration of 30 to 64 M for a variety of human and animal cell lines in vitro. Concomitantly, we observed an improvement in proliferation rates and changes in the morphology of mycoplasmacontaminated mammalian cells after treatment with this drug. A single treatment over one passage led to complete removal of viable Mycoplasma hyorhinis cells from various adherent cell lines, and Mycoplasma orale was removed from nonadherent human T-lymphoid cell lines by double treatment. This effect was monitored by a DNA fluorescence test, an enzyme-linked immunosorbent assay, and two different PCR methods. Disintegration of the mycoplasma membranes as observed by electron microscopy indicated the mode of action of surfactin. Disintegration is obviously due to a physicochemical interaction of the membrane-active surfactant with the outer part of the lipid membrane bilayer, which causes permeability changes and at higher concentrations leads finally to disintegration of the mycoplasma membrane system by a detergent effect. The low cytotoxicity of surfactin for mammalian cells permits specific inactivation of mycoplasmas without significant deleterious effects on cell metabolism and the proliferation rate in cell culture. These results were used to develop a fast and simple method for complete and permanent inactivation of mycoplasmas in mammalian monolayer and suspension cell cultures.

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“…In this sense, previous studies on Mycoplasma sp. free eukaryotic cell lines do not provide any evidence of antibiotic effect on host cell viability toxicity [31]. However, we observed that Plasmocin TM and Ciprofloxacin at curative concentrations (25 and 10 µg/ml, respectively) significantly reduced H9 hESCs growth rate.…”

Section: Discussionmentioning

confidence: 65%

Effect of Antibiotics against Mycoplasma sp. on Human Embryonic Stem Cells Undifferentiated Status, Pluripotency, Cell Viability and Growth

et al. 2013

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Human embryonic stem cells (hESCs) are self-renewing pluripotent cells that can differentiate into specialized cells and hold great promise as models for human development and disease studies, cell-replacement therapies, drug discovery and in vitro cytotoxicity tests. The culture and differentiation of these cells are both complex and expensive, so it is essential to extreme aseptic conditions. hESCs are susceptible to Mycoplasma sp. infection, which is hard to detect and alters stem cell-associated properties. The purpose of this work was to evaluate the efficacy and cytotoxic effect of PlasmocinTM and ciprofloxacin (specific antibiotics used for Mycoplasma sp. eradication) on hESCs. Mycoplasma sp. infected HUES-5 884 (H5 884, stable hESCs H5-brachyury promoter-GFP line) cells were effectively cured with a 14 days PlasmocinTM 25 µg/ml treatment (curative treatment) while maintaining stemness characteristic features. Furthermore, cured H5 884 cells exhibit the same karyotype as the parental H5 line and expressed GFP, through up-regulation of brachyury promoter, at day 4 of differentiation onset. Moreover, H5 cells treated with ciprofloxacin 10 µg/ml for 14 days (mimic of curative treatment) and H5 and WA09 (H9) hESCs treated with PlasmocinTM 5 µg/ml (prophylactic treatment) for 5 passages retained hESCs features, as judged by the expression of stemness-related genes (TRA1-60, TRA1-81, SSEA-4, Oct-4, Nanog) at mRNA and protein levels. In addition, the presence of specific markers of the three germ layers (brachyury, Nkx2.5 and cTnT: mesoderm; AFP: endoderm; nestin and Pax-6: ectoderm) was verified in in vitro differentiated antibiotic-treated hESCs. In conclusion, we found that PlasmocinTM and ciprofloxacin do not affect hESCs stemness and pluripotency nor cell viability. However, curative treatments slightly diminished cell growth rate. This cytotoxic effect was reversible as cells regained normal growth rate upon antibiotic withdrawal.

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Mycoplasma contamination in human leukemia cell lines

Cited by 87 publications

References 10 publications

Therapeutic potential for phenytoin: targeting Nav1.5 sodium channels to reduce migration and invasion in metastatic breast cancer

Therapeutic potential for phenytoin: targeting Nav1.5 sodium channels to reduce migration and invasion in metastatic breast cancer

Antimycoplasma properties and application in cell culture of surfactin, a lipopeptide antibiotic from Bacillus subtilis

Effect of Antibiotics against Mycoplasma sp. on Human Embryonic Stem Cells Undifferentiated Status, Pluripotency, Cell Viability and Growth

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