Use antibiotics in cell culture with caution: genome-wide identification of antibiotic-induced changes in gene expression and regulation

Abstract

Standard cell culture guidelines often use media supplemented with antibiotics to prevent cell contamination. However, relatively little is known about the effect of antibiotic use in cell culture on gene expression and the extent to which this treatment could confound results. To comprehensively characterize the effect of antibiotic treatment on gene expression, we performed RNA-seq and ChIP-seq for H3K27ac on HepG2 cells, a human liver cell line commonly used for pharmacokinetic, metabolism and genomic studies, cultured in media supplemented with penicillin-streptomycin (PenStrep) or vehicle control. We identified 209 PenStrep-responsive genes, including transcription factors such as ATF3 that are likely to alter the regulation of other genes. Pathway analyses found a significant enrichment for “xenobiotic metabolism signaling” and “PXR/RXR activation” pathways. Our H3K27ac ChIP-seq identified 9,514 peaks that are PenStrep responsive. These peaks were enriched near genes that function in cell differentiation, tRNA modification, nuclease activity and protein dephosphorylation. Our results suggest that PenStrep treatment can significantly alter gene expression and regulation in a common liver cell type such as HepG2, advocating that antibiotic treatment should be taken into account when carrying out genetic, genomic or other biological assays in cultured cells.

A cyclopeptide and three oligomycin-class polyketides produced by an underexplored actinomycete of the genus Pseudosporangium

“…Microbial secondary metabolites have been used as therapeutic drugs [1], veterinary medicines [2], agrochemicals [3], food preservatives/colorings [4,5], medium supplements for selective microbial/cell culture [6][7][8], or biochemical reagents for pharmacological/chemical biology studies [9] and continue to be indispensable to support and improve human welfare and social life. In recent years, in accordance with the advancement of genome and in silico analytical technologies, the searching process for new microbial secondary metabolites became faster and more efficient [10].…”

Serum-free media for the growth of primary bovine myoblasts

“…A second problem with large-scale cell and tissue culture for food application is the routine addition of antibiotics and fungostatics to reduce the risk of infection. Ryu et al have shown that PenStrep treatment of HepG2 cells during in vitro culture can significantly alter gene expression and regulation (Ryu et al 2017) and Skubis et al have shown that use of antibiotics has an impact on the phenotype of adipose derived stem cells, affecting both their proliferation and differentiation, but still little is known about the consequences of antibiotics-free culture of mammalian cells (Skubis et al 2017). Therefore, the secondary aim of this study was to investigate the possibility and effects of myoblast cell culture in serum containing and serum-free serum medium without using antibiotics.…”

Conservation in the wake of myrtle rust – a case study on two critically endangered Australian rainforest plants

“…Plant Preservative Mixture TM (a broad-spectrum microbicide) is sometimes used in tissue culture media to reduce contamination but this mixture is not always effective in controlling endogenous microbes and can be phytotoxic (Thomas et al 2017). The use of antibiotics in culture can also lead to changes in gene expression and regulation (Ryu et al 2017).…”