Skip to main content
SpringerLink
Log in

Benzo[k]fluoranthene-induced changes in miRNA-mRNA interactions in human hepatocytes

  • Research Article
  • Published:
Toxicology and Environmental Health Sciences Aims and scope Submit manuscript

Abstract

Toxicology studies assessing the risk of environmental toxicants in humans frequently use in vitro systems in combination with transcriptomics to characterize toxic responses. Thus far, changes have mostly been investigated at the mRNA level. Recently, microRNAs (miRNAs) have attracted attention because they are powerful negative regulators of mRNA levels and thus may be responsible for the modulation of important mRNA networks implicated in toxicity. This study aimed to identify possible miRNA-mRNA networks as novel interactions at the gene expression level after exposure to environmental toxicants. Benzo[k]fluoranthene (BF), a polycyclic aromatic hydrocarbon that is ubiquitously distributed throughout the environment, was used. We analyzed mRNA and miRNA profiles in HepG2 cells, a human liver cell line, using a human oligonucleotide chip. Changes in miRNA expression in response to BF, in combination with multiple alterations of mRNA levels, were observed. Many of the altered mRNAs were targets of altered miRNAs. Using gene ontology (GO) and KEGG pathway analysis, we determined the relevance of such miRNA deregulation to carcinogenicity. This revealed five miRNAs that appear to participate in specific BFresponsive pathways relevant to genotoxicity and carcinogenicity, such as DNA damage repair, apoptosis, cancer, VEGF signaling, and Jak-STAT signaling. Our results indicate that miR-146a, miR-365, let-7f, miR-199b-5p, and miR-30c-1* are novel players in the BF response. Therefore, this study demonstrates the added value of an integrated miRNA-mRNA approach for identification of the molecular mechanisms induced by BF in an in vitro human model.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Rana, T. M. Illuminating the silence: understanding the structure and function of small RNAs. Nat. Rev. Mol. Cell Biol. 8, 23–36 (2007).

    Article  PubMed  CAS  Google Scholar 

  2. Lu, J. et al. MicroRNA expression profiles classify human cancers. Nature 435, 834–838 (2005).

    Article  PubMed  CAS  Google Scholar 

  3. Volinia, S. et al. A microRNA expression signature of human solid tumors defines cancer gene targets. Proc. Natl. Acad. Sci. USA 103, 2257–2261 (2006).

    Article  PubMed  CAS  Google Scholar 

  4. Taylor, E. L. & Gant, T. W. Emerging fundamental roles for non-coding RNA species in toxicology. Toxicology 246, 34–39 (2008).

    Article  PubMed  CAS  Google Scholar 

  5. Brueckner, B. et al. The human let-7a-3 locus contains an epigenetically regulated MicroRNA gene with oncogenic function. Cancer Res. 67, 1419–1423 (2007).

    Article  PubMed  CAS  Google Scholar 

  6. Johnson, S., Lin, S. & Slack, F. The time of appearance of the C. elegans let-7 microRNA is transcriptionally controlled utilizing a temporal regulatory element in its promoter. Dev. Biol. 259, 364–379 (2003).

    Article  PubMed  CAS  Google Scholar 

  7. Meng, F. et al. Involvement of human micro-RNA in growth and response to chemotherapy in human cholangiocarcinoma cell lines. Gastroenterology 130, 2113–2129 (2006).

    Article  PubMed  CAS  Google Scholar 

  8. Pogribny, I. P. et al. Induction of microRNAome deregulation in rat liver by long-term tamoxifen exposure. Mutat. Res. 619, 30–37 (2007).

    Article  PubMed  CAS  Google Scholar 

  9. Sathyan, P., Golden, H. B. & Miranda, R. C. Competing interactions between micro-RNAs determine neural progenitor survival and proliferation after ethanol exposure: evidence from an ex vivo model of the fetal cerebral cortical neuroepithelium. J. Neurosci. 27, 8546–8557 (2007).

    Article  PubMed  CAS  Google Scholar 

  10. He, L. & Hannon, G. J. MicroRNAs: small RNAs with a big role in gene regulation. Nat. Rev. Genet. 5, 522–531 (2004).

    Article  PubMed  CAS  Google Scholar 

  11. Pothof, J. et al. MicroRNA-mediated gene silencing modulates the UV-induced DNA-damage response. EMBO J. 28, 2090–2099 (2009).

    Article  PubMed  CAS  Google Scholar 

  12. Kumar, M. S., Lu, J., Mercer, K. L., Golub, T. R. & Jacks, T. Impaired microRNA processing enhances cellular transformation and tumorigenesis. Nat. Genet. 39, 673–677 (2007).

    Article  PubMed  CAS  Google Scholar 

  13. Guo, H., Ingolia, N. T., Weissman, J. S. & Bartel, D. P. Mammalian microRNAs predominantly act to decrease target mRNA levels. Nature 466, 835–840 (2010).

    Article  PubMed  CAS  Google Scholar 

  14. Sun, B. K. & Tsao, H. Small RNAs in development and disease. J. Am. Acad. Dermatol. 59, 725–737 (2008).

    Article  PubMed  Google Scholar 

  15. Wang, Y., Liang, Y. & Lu, Q. MicroRNA epigenetic alterations: Predicting biomarkers and therapeutic targets in human diseases. Clin. Genet. 74, 307–315 (2008).

    Article  PubMed  CAS  Google Scholar 

  16. Lynam-Lennon, N., Maher, S. G. & Reynolds, J. V. The roles of micro-RNA in cancer and apoptosis. Biol. Rev. Camb. Philos. Soc. 84, 55–71 (2009).

    Article  PubMed  Google Scholar 

  17. Liu, H. & Kohane, I. S. Tissue and process specific microRNA-mRNA co-expression in mammalian development and malignancy. PLoS One 4, e5436 (2009).

    Article  PubMed  Google Scholar 

  18. Lafferty-Whyte, K., Cairney, C. J., Jamieson, N. B., Oien, K. A. & Keith, W. N. Pathway analysis of sene-scence-associated miRNA targets reveals common processes to different senescence induction mechanisms. Biochim. Biophys. Acta. 1792, 341–352 (2009).

    Article  PubMed  CAS  Google Scholar 

  19. Fukushima, T., Hamada, Y., Yamada, H. & Horii, I. Changes of micro-RNA expression in rat liver treated by acetaminophen or carbon tetrachloride-regulating role of micro-RNA for RNA expression. J. Toxicol. Sci. 32, 401–409 (2007).

    Article  PubMed  CAS  Google Scholar 

  20. Wang, K. et al. Circulating microRNAs, potential biomarkers for drug-induced liver injury. Proc. Natl. Acad. Sci. USA 106, 4402–4407 (2009).

    Article  PubMed  CAS  Google Scholar 

  21. Agency for Toxic Substances and Disease Registry. Toxicological Profile for Polycyclic Aromatic Hydrocarbons (PAHs). Atlanta, GA: ATSDR (1995).

    Google Scholar 

  22. Mastrangelo, G., Fadda, E. & Marzia, V. Polycyclic aromatic hydrocarbons and cancer in man. Environ. Health Perspect. 104, 1166–1170 (1996).

    Article  PubMed  CAS  Google Scholar 

  23. Song, M. K. et al. Gene expression analysis identifies DNA damage-related markers of benzo[a]pyrene exposure in HepG2 human hepatocytes. Toxicol. Environ. Health. Sci. 4, 19–29 (2012).

    Article  Google Scholar 

  24. Izzotti, A. et al. Downregulation of microRNA expression in the lungs of rats exposed to cigarette smoke. FASEB J. 23, 806–812 (2009).

    Article  PubMed  CAS  Google Scholar 

  25. Schembri, F. et al. MicroRNAs as modulators of smoking-induced gene expression changes in human airway epithelium. Proc. Natl. Acad. Sci. USA 106, 2319–2324 (2009).

    Article  PubMed  CAS  Google Scholar 

  26. Bolleyn, J. et al. Effect of Trichostatin A on miRNA expression in cultures of primary rat hepatocytes. Toxicol. in Vitro 25, 1173–1182 (2011).

    Article  PubMed  CAS  Google Scholar 

  27. Yauk, C. L., Jackson, K., Malowany, M. & Williams, A. Lack of change in microRNA expression in adult mouse liver following treatment with benzo(a)pyrene despite robust mRNA transcriptional response. Mutat. Res. 722, 131–139 (2010).

    PubMed  Google Scholar 

  28. Halappanavar, S. et al. Pulmonary gene and microRNA expression changes in mice exposed to benzo(a)pyrene by oral gavage. Toxicology 285, 133–141 (2011).

    Article  PubMed  CAS  Google Scholar 

  29. Duan, H., Jiang, Y., Zhang, H. & Wu, Y. MiR-320 and miR-494 affect cell cycles of primary murine bronchial epithelial cells exposed to benzo[a]pyrene. Toxicol. in Vitro 24, 928–935 (2010).

    Article  PubMed  CAS  Google Scholar 

  30. Lema, C. & Cunningham, M. J. MicroRNAs and their implications in toxicological research. Toxicol. Lett. 198, 100–105 (2010).

    Article  PubMed  CAS  Google Scholar 

  31. Yokoi, T. & Nakajima, M. Toxicological implications of modulation of gene expression by microRNAs. Toxicol. Sci. 123, 1–14 (2011).

    Article  PubMed  CAS  Google Scholar 

  32. Audebert, M. et al. Use of the gammaH2AX assay for assessing the genotoxicity of polycyclic aromatic hydrocarbons in human cell lines. Toxicol. Lett. 199, 182–192 (2010).

    Article  PubMed  CAS  Google Scholar 

  33. Song, M. K., Kim, Y. J., Song, M., Choi, H. S. & Ryu, J. C. Dose-response functional gene analysis by exposure to 3 different polycyclic aromatic hydrocarbons in human hepatocytes. Mol. Cell. Toxicol. 7, 221–232 (2011).

    Article  CAS  Google Scholar 

  34. Yokoi, T. & Nakajima, M. Toxicological implications of modulation of gene expression by microRNAs. Toxicol. Sci. 123, 1–14 (2011).

    Article  PubMed  CAS  Google Scholar 

  35. Sonkoly, E. & Pivarcsi, A. MicroRNAs in inflammation and response to injuries induced by environmental pollution. Mutat. Res. 17, 46–53 (2011).

    Google Scholar 

  36. Moffat, I. D. et al. microRNAs in adult rodent liver are refractory to dioxin treatment. Toxicol. Sci. 99, 470–487 (2007).

    Article  PubMed  CAS  Google Scholar 

  37. Malik, A. I., Williams, A., Lemieux, C. L., White, P. A. & Yauk, C. L. Hepatic mRNA, microRNA, and miR-34a-Target responses in mice after 28 days exposure to doses of benzo(a)pyrene that elicit DNA damage and mutation. Environ. Mol. Mutagen. 53, 10–21 (2012).

    Article  PubMed  CAS  Google Scholar 

  38. Farh, K. K. et al. The widespread impact of mammalian MicroRNAs on mRNA repression and evolution. Science 310, 1817–1821 (2005).

    Article  PubMed  CAS  Google Scholar 

  39. Favaro, E. et al. MicroRNA-210 regulates mitochondrial free radical response to hypoxia and krebs cycle in cancer cells by targeting iron sulfur cluster protein ISCU. PLoS One 5, e10345 (2010).

    Article  PubMed  Google Scholar 

  40. Sethi, J. K. & Vidal-Puig, A. Wnt signalling and the control of cellular metabolism. Biochem. J. 427, 1–17 (2010).

    Article  PubMed  CAS  Google Scholar 

  41. Hutcheson, J. et al. Combined deficiency of proapoptotic regulators Bim and Fas results in the early onset of systemic autoimmunity. Immunity 28, 206–217 (2008).

    Article  PubMed  CAS  Google Scholar 

  42. Zhang, L. et al. Integrative genomic analysis of phosphatidylinositol 3′-kinase family identifies PIK3R3 as a potential therapeutic target in epithelial ovarian cancer. Clin. Cancer Res. 13, 5314–5321 (2007).

    Article  PubMed  CAS  Google Scholar 

  43. Wang, H. Q. et al. Deregulated miR-155 promotes Fasmediated apoptosis in human intervertebral disc degeneration by targeting FADD and caspase-3. J. Pathol. 225, 32–42 (2011).

    Google Scholar 

  44. Crowder, R. J. et al. PIK3CA and PIK3CB inhibition produce synthetic lethality when combined with estrogen deprivation in estrogen receptor-positive breast cancer. Cancer Res. 69, 3955–3962 (2009).

    Article  PubMed  CAS  Google Scholar 

  45. Qin, B. et al. MicroRNAs expression in ox-LDL treated HUVECs: MiR-365 moudlates apoptosis and Bcl-2 expression. Biochem. Biophys. Res. Commun. 410, 127–133 (2011).

    Article  PubMed  CAS  Google Scholar 

  46. Li, J. et al. miR-146a, an IL-1β responsive miRNA, induces vascular endothelial growth factor and chondrocyte apoptosis by targeting Smad4. Arthritis Res. Ther. 14, R75 (2012).

    Article  PubMed  CAS  Google Scholar 

  47. Dang, C. V. c-Myc target genes involved in cell growth, apoptosis, and metabolism. Mol. Cell. Biol. 19, 1–11 (1999).

    PubMed  CAS  Google Scholar 

  48. Cazzalini, O., Scovassi, A. I., Savio, M., Stivala, L. A. & Prosperi, E. Multiple roles of the cell cycle inhibitor p21 (CDKN1A) in the DNA damage response. Mutat. Res. 704, 12–20 (2010).

    Article  PubMed  CAS  Google Scholar 

  49. Sujobert, P. et al. Essential role for the p110d isoform in phosphoinositide 3-kinase activation and cell proliferation in acute myeloid leukemia. Blood 106, 1063–1066 (2005).

    Article  PubMed  CAS  Google Scholar 

  50. Mosmann, T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Methods 65, 55–63 (1983).

    Article  PubMed  CAS  Google Scholar 

  51. Park, H. W., Kim, S. J. & Oh, M. J. Gene expression patterns of environmental chemicals in human cell lines using HazChem human array. BioChip J. 3, 65–70 (2009).

    Google Scholar 

  52. Yim, W. C. et al. Identification of novel 17β-estradiol (E2) target genes using crossexperiment gene expression datasets. Toxicol. Environ. Health. Sci. 2, 25–38 (2010).

    Article  Google Scholar 

  53. Benjamini, Y. & Hochberg, Y. Controlling the false discovery rate: A practical and powerful approach to multiple testing. J. R. Statist. Soc. B 57, 289–300 (1995).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Center for Integrated Risk Research, Korea Institute of Science & Technology, P.O. Box 131, Cheongryang, Seoul, 130-650, Korea

    Mi-Kyung Song, Mee Song, Han-Seam Choi & Jae-Chun Ryu

Authors
  1. Mi-Kyung Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mee Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Han-Seam Choi
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jae-Chun Ryu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mi-Kyung Song.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Song, MK., Song, M., Choi, HS. et al. Benzo[k]fluoranthene-induced changes in miRNA-mRNA interactions in human hepatocytes. Toxicol. Environ. Health Sci. 4, 143–153 (2012). https://doi.org/10.1007/s13530-012-0129-2

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13530-012-0129-2

Keywords

Search

Navigation