Culturing conditions highly affect DNA methylation and gene expression levels in MCF7 breast cancer cell line
- 194 Downloads
The levels of DNA methylation and their role in gene expression are key factors that could affect diagnosis, prognosis, and treatment options of different diseases. In this study, the methylation levels of 22 genes that are mostly correlated to breast cancer were determined using EpiTect methyl II PCR array. This analysis was performed to determine the effect of cells’ passage number and the use of antibiotics in the culturing media on gene methylation levels in MCF7 cell line. DNA methylation levels of PTGS2, ADAM23, HIC1, and PYCARD were found to be significantly different among different passages. While the DNA methylation levels of CCNA1, RASSF1, and THBS1 were found to be affected by the use of 1% of penicillin/streptomycin in the culture media. Gene expression analysis after demethylation using 5-Aza-2′-deoxycytidine showed that the gene expression levels of the hypermethylated genes varied between different passage numbers. This study shows that the presence of antibiotic within cultured media and cell line’s passage number could greatly affect the methylation levels that need to be considered in future studies on cell lines.
KeywordsDNA methylation Breast cancer mcf-7 cell line Passage number Penicillin/streptomycin
This study received a financial support from Al-Zaytoonah University of Jordan.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
- Benevolenskaya EV, Islam AB, Ahsan H, Kibriya MG, Jasmine F, Wolff B, Al-Alem U, Wiley E, Kajdacsy-Balla A, Macias V, Rauscher GH (2016) DNA methylation and hormone receptor status in breast cancer. Clin Epigenetics 8:17. https://doi.org/10.1186/s13148-016-0184-7 eCollection 2016CrossRefPubMedPubMedCentralGoogle Scholar
- Heitzer E, Artl M, Filipits M, Resel M, Graf R, Weißenbacher B, Lax S, Gnant M, Wrba F, Greil R, Dietze O, Hofbauer F, Böhm G, Höfler G, Samonigg H, Schaberl-Moser R, Balic M, Dandachi N (2014) Differential survival trends of stage II colorectal cancer patients relate to promoter methylation status of PCDH10, SPARC, and UCHL1. Mod Pathol 27(6):906–915. https://doi.org/10.1038/modpathol.2013.204 CrossRefPubMedGoogle Scholar
- Hinshelwood RA, Huschtscha LI, Melki J, Stirzaker C, Abdipranoto A, Vissel B, Ravasi T, Wells CA, Hume DA, Reddel RR, Clark SJ (2007) Concordant epigenetic silencing of transforming growth factor-beta signaling pathway genes occurs early in breast carcinogenesis. Cancer Res 67(24):11517–11527CrossRefPubMedGoogle Scholar
- Klajic J, Busato F, Edvardsen H, Touleimat N, Fleischer T, Bukholm I, Børresen-Dale AL, Lønning PE, Tost J, Kristensen VN (2014) DNA methylation status of key cell-cycle regulators such as CDKNA2/p16 and CCNA1 correlates with treatment response to doxorubicin and 5-fluorouracil in locally advanced breast tumors. Clin Cancer Res 20(24):6357–6366. https://doi.org/10.1158/1078-0432.CCR-14-0297 CrossRefPubMedGoogle Scholar
- Shames DS, Girard L, Gao B, Sato M, Lewis CM, Shivapurkar N, Jiang A, Perou CM, Kim YH, Pollack JR, Fong KM, Lam CL, Wong M, Shyr Y, Nanda R, Olopade OI, Gerald W, Euhus DM, Shay JW, Gazdar AF, Minna JD (2006) A genome-wide screen for promoter methylation in lung cancer identifies novel methylationmarkers for multiple malignancies. PLoS Med 3(12):e486CrossRefPubMedPubMedCentralGoogle Scholar