Skip to main content
SpringerLink
Log in

Altered microRNome Profiling in Statin-Induced HepG2 Cells: A Pilot Study Identifying Potential new Biomarkers Involved in Lipid-Lowering Treatment

  • ORIGINAL ARTICLE
  • Published:
Cardiovascular Drugs and Therapy Aims and scope Submit manuscript

Abstract

Purpose

Statins are widely prescribed drugs to manage hypercholesterolemia. Despite they are considered effective lipid-lowering agents, significant inter-individual variability has been reported in relation to drug response. Among the reasons explaining this variation, genetic factors are known to partially contribute. Nonetheless, poor evidence exists regarding epigenetic factors involved.

Methods

We investigated if atorvastatin can modulate the cholesterol related miR-33 family. Furthermore, we analyzed the microRNA expression profiles in HepG2 cells treated for 24 hours with atorvastatin or simvastatin using a microarray platform.

Results

Our results indicate that atorvastatin does not influence the expression of the miR-33 family. In addition, microarray examination revealed that atorvastatin modulated thirteen miRs, whilst simvastatin only affected two miRs. All significantly modulated miRs after simvastastin therapy were also modulated by atorvastatin. In addition, four novel miRs with previously unreported functions were identified as statin-modulated.

Conclusion

We identified several novel miRs affected by statin treatment. Additional research is needed to determine the biological significance of differentially expressed miRs identified in statins-induced HepG2 cells.

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

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  1. Ross SD, Allen IE, Connelly JE, Korenblat BM, Smith ME, Bishop D, et al. Clinical outcomes in statin treatment trials: a meta-analysis. Arch Intern Med. 1999;159(15):1793–802.

    Article  CAS  PubMed  Google Scholar 

  2. LaRosa JC, He J, Vupputuri S. Effect of statins on risk of coronary disease: a meta-analysis of randomized controlled trials. JAMA. 1999;282(24):2340–6.

    Article  CAS  PubMed  Google Scholar 

  3. Davidson MH, Toth PP. Comparative effects of lipid-lowering therapies. Prog Cardiovasc Dis. 2004;47(2):73–104.

    Article  CAS  PubMed  Google Scholar 

  4. Alagona Jr P. Pitavastatin: evidence for its place in treatment of hypercholesterolemia. Core Evid. 2010;5:91–105.

  5. Liao JK, Laufs U. Pleiotropic effects of statins. Annu Rev Pharmacol Toxicol. 2005;45:89–118.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  6. Wang CY, Liu PY, Liao JK. Pleiotropic effects of statin therapy: molecular mechanisms and clinical results. Trends Mol Med. 2008;14(1):37–44.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  7. Golomb BA, Evans MA. Statin adverse effects : a review of the literature and evidence for a mitochondrial mechanism. Am J Cardiovasc Drugs. 2008;8(6):373–418.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  8. Reiner Z Resistance and intolerance to statins. Nutr Metab Cardiovasc Dis. 2014;24(10):1057–66.

    Article  CAS  PubMed  Google Scholar 

  9. Mangravite LM, Thorn CF, Krauss RM. Clinical implications of pharmacogenomics of statin treatment. Pharmacogenomics J. 2006;6(6):360–74..

    Article  CAS  PubMed  Google Scholar 

  10. Rodrigues AC, Hirata MH, Hirata RD. The genetic determinants of atorvastatin response. Curr Opin Mol Ther. 2007;9(6):545–53.

    CAS  PubMed  Google Scholar 

  11. Postmus I, Verschuren JJ, de Craen AJ, Slagboom PE, Westendorp RG, Jukema JW, et al. Pharmacogenetics of statins: achievements, whole-genome analyses and future perspectives. Pharmacogenomics. 2012;13(7):831–40.

    Article  CAS  PubMed  Google Scholar 

  12. Postmus I, Trompet S, Deshmukh HA, Barnes MR, Li X, Warren HR, et al. Pharmacogenetic meta-analysis of genome-wide association studies of LDL cholesterol response to statins. Nat Commun. 2014;5:5068.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  13. Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004;116(2):281–97.

    Article  CAS  PubMed  Google Scholar 

  14. Esau C, Davis S, Murray SF, Yu XX, Pandey SK, Pear M, et al. miR-122 regulation of lipid metabolism revealed by in vivo antisense targeting. Cell Metab. 2006;3(2):87–98.

    Article  CAS  PubMed  Google Scholar 

  15. Krutzfeldt J, Rajewsky N, Braich R, Rajeev KG, Tuschl T, Manoharan M, et al. Silencing of microRNAs in vivo with ‘antagomirs’. Nature. 2005;438(7068):685–9.

    Article  PubMed  Google Scholar 

  16. Fichtlscherer S, De Rosa S, Fox H, Schwietz T, Fischer A, Liebetrau C, et al. Circulating microRNAs in patients with coronary artery disease. Circ Res. 2010;107(5):677–84.

    Article  CAS  PubMed  Google Scholar 

  17. Hoekstra M, van der Lans CA, Halvorsen B, Gullestad L, Kuiper J, Aukrust P, et al. The peripheral blood mononuclear cell microRNA signature of coronary artery disease. Biochem Biophys Res Commun. 2010;394(3):792–7.

    Article  CAS  PubMed  Google Scholar 

  18. Tan KS, Armugam A, Sepramaniam S, Lim KY, Setyowati KD, Wang CW, et al. Expression profile of MicroRNAs in young stroke patients. PLoS One. 2009;4(11):e7689.

    Article  PubMed Central  PubMed  Google Scholar 

  19. Najafi-Shoushtari SH, Kristo F, Li Y, Shioda T, Cohen DE, Gerszten RE, et al. MicroRNA-33 and the SREBP host genes cooperate to control cholesterol homeostasis. Science. 2010;328(5985):1566–9.

    Article  CAS  PubMed  Google Scholar 

  20. Rayner KJ, Suarez Y, Davalos A, Parathath S, Fitzgerald ML, Tamehiro N, et al. MiR-33 contributes to the regulation of cholesterol homeostasis. Science. 2010;328(5985):1570–3.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Minami Y, Satoh M, Maesawa C, Takahashi Y, Tabuchi T, Itoh T, et al. Effect of atorvastatin on microRNA 221/222 expression in endothelial progenitor cells obtained from patients with coronary artery disease. Eur J Clin Investig. 2009;39(5):359–67.

    Article  CAS  Google Scholar 

  22. Tabuchi T, Satoh M, Itoh T, Nakamura M. MicroRNA-34a regulates the longevity-associated protein SIRT1 in coronary artery disease: effect of statins on SIRT1 and microRNA-34a expression. Clin Sci (Lond). 2012;123(3):161–71.

    Article  CAS  Google Scholar 

  23. Wang H, Lu HM, Yang WH, Luo C, Lu SH, Zhou Y, et al. The influence of statin therapy on circulating microRNA-92a expression in patients with coronary heart disease. Zhongguo Wei Zhong Bing Ji Jiu Yi Xue. 2012;24(4):215–8.

    PubMed  Google Scholar 

  24. Takahashi Y, Satoh M, Minami Y, Tabuchi T, Itoh T, Nakamura M. Expression of miR-146a/b is associated with the toll-like receptor 4 signal in coronary artery disease: effect of renin-angiotensin system blockade and statins on miRNA-146a/b and toll-like receptor 4 levels. Clin Sci (Lond). 2010;119(9):395–405.

    Article  CAS  Google Scholar 

  25. Takwi AA, Li Y, Becker Buscaglia LE, Zhang J, Choudhury S, Park AK, et al. A statin-regulated microRNA represses human c-Myc expression and function. EMBO Mol Med. 2012;4(9):896–909.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  26. Tu Y, Wan L, Bu L, Zhao D, Dong D, Huang T, et al. MicroRNA-22 downregulation by atorvastatin in a mouse model of cardiac hypertrophy: a new mechanism for antihypertrophic intervention. Cell Physiol Biochem. 2013;31(6):997–1008.

    Article  CAS  PubMed  Google Scholar 

  27. Hu JR, Lv GH, Yin BL. Altered microRNA expression in the ischemic-reperfusion spinal cord with atorvastatin therapy. J Pharmacol Sci. 2013;121(4):343–6.

    Article  CAS  PubMed  Google Scholar 

  28. Lennernas H, Fager G. Pharmacodynamics and pharmacokinetics of the HMG-CoA reductase inhibitors. Similarities and differences. Clin Pharmacokinet. 1997;32(5):403–25.

    Article  CAS  PubMed  Google Scholar 

  29. Cerda A, Genvigir FD, Rodrigues AC, Willrich MA, Dorea EL, Bernik MM, et al. Influence of polymorphisms and cholesterol-lowering treatment on SCARB1 mRNA expression. J Atheroscler Thromb. 2011;18(8):640–51.

    Article  CAS  PubMed  Google Scholar 

  30. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(−delta delta C(T)) method. Methods. 2001;25(4):402–8.

    Article  CAS  PubMed  Google Scholar 

  31. Bolstad BM, Irizarry RA, Astrand M, Speed TP. A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics. 2003;19(2):185–93.

    Article  CAS  PubMed  Google Scholar 

  32. Randomised trial of cholesterol lowering in 4444 patients with coronary heart disease: the Scandinavian Simvastatin Survival Study (4S). Lancet. 1994;344(8934):1383–9.

  33. Vrecer M, Turk S, Drinovec J, Mrhar A. Use of statins in primary and secondary prevention of coronary heart disease and ischemic stroke. Meta-analysis of randomized trials. Int J Clin Pharmacol Ther. 2003;41(12):567–77.

    Article  CAS  PubMed  Google Scholar 

  34. Naci H, Brugts JJ, Fleurence R, Tsoi B, Toor H, Ades AE. Comparative benefits of statins in the primary and secondary prevention of major coronary events and all-cause mortality: a network meta-analysis of placebo-controlled and active-comparator trials. Eur J Prev Cardiol. 2013;20(4):641–57.

    Article  PubMed  Google Scholar 

  35. Marquart TJ, Allen RM, Ory DS, Baldan A. miR-33 links SREBP-2 induction to repression of sterol transporters. Proc Natl Acad Sci U S A. 2010;107(27):12228–32.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  36. Niesor EJ, Schwartz GG, Perez A, Stauffer A, Durrwell A, Bucklar-Suchankova G, et al. Statin-induced decrease in ATP-binding cassette transporter A1 expression via microRNA33 induction may counteract cholesterol efflux to high-density lipoprotein. Cardiovasc Drugs Ther. 2015;29(1):7–14.

    Article  CAS  PubMed  Google Scholar 

  37. Stormo C, Kringen MK, Lyle R, Olstad OK, Sachse D, Berg JP, et al. RNA-sequencing analysis of HepG2 cells treated with atorvastatin. PLoS One. 2014;9(8):e105836.

    Article  PubMed Central  PubMed  Google Scholar 

  38. Barter P CETP and atherosclerosis. Arterioscler Thromb Vasc Biol. 2000;20(9):2029–31.

    Article  CAS  PubMed  Google Scholar 

  39. Genvigir FD, Rodrigues AC, Cerda A, Arazi SS, Willrich MA, Oliveira R, et al. Effects of lipid-lowering drugs on reverse cholesterol transport gene expressions in peripheral blood mononuclear and HepG2 cells. Pharmacogenomics. 2010;11(9):1235–46.

    Article  CAS  PubMed  Google Scholar 

  40. Istvan ES, Deisenhofer J. Structural mechanism for statin inhibition of HMG-CoA reductase. Science. 2001;292(5519):1160–4.

    Article  CAS  PubMed  Google Scholar 

  41. Istvan E Statin inhibition of HMG-CoA reductase: a 3-dimensional view. Atheroscler Suppl. 2003;4(1):3–8.

    Article  CAS  PubMed  Google Scholar 

  42. da Costa RF, Freire VN, Bezerra EM, Cavada BS, Caetano EW, de Lima Filho JL, et al. Explaining statin inhibition effectiveness of HMG-CoA reductase by quantum biochemistry computations. Phys Chem Chem Phys. 2012;14(4):1389–98.

    Article  PubMed  Google Scholar 

  43. Dansette PM, Jaoen M, Pons C. HMG-CoA reductase activity in human liver microsomes: comparative inhibition by statins. Exp Toxicol Pathol. 2000;52(2):145–8.

    Article  CAS  PubMed  Google Scholar 

  44. Leszczynska A, Gora M, Plochocka D, Hoser G, Szkopinska A, Koblowska M, et al. Different statins produce highly divergent changes in gene expression profiles of human hepatoma cells: a pilot study. Acta Biochim Pol. 2011;58(4):635–9.

    CAS  PubMed  Google Scholar 

  45. Schachter M Chemical, pharmacokinetic and pharmacodynamic properties of statins: an update. Fundam Clin Pharmacol. 2005;19(1):117–25.

    Article  CAS  PubMed  Google Scholar 

  46. Connolly E, Melegari M, Landgraf P, Tchaikovskaya T, Tennant BC, Slagle BL, et al. Elevated expression of the miR-17-92 polycistron and miR-21 in hepadnavirus-associated hepatocellular carcinoma contributes to the malignant phenotype. Am J Pathol. 2008;173(3):856–64.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  47. Shan SW, Fang L, Shatseva T, Rutnam ZJ, Yang X, Du W, et al. Mature miR-17-5p and passenger miR-17-3p induce hepatocellular carcinoma by targeting PTEN, GalNT7 and vimentin in different signal pathways. J Cell Sci. 2013;126(Pt 6):1517–30.

    Article  CAS  PubMed  Google Scholar 

  48. Gao Y, Lu XC, Yang HY, Liu XF, Cao J, Fan L. The molecular mechanism of the anticancer effect of atorvastatin: DNA microarray and bioinformatic analyses. Int J Mol Med. 2012;30(4):765–74.

    CAS  PubMed  Google Scholar 

  49. Lakshminarayana Reddy CN, Vyjayanti VN, Notani D, Galande S, Kotamraju S. Down-regulation of the global regulator SATB1 by statins in COLO205 colon cancer cells. Mol Med Rep. 2010;3(5):857–61.

    CAS  PubMed  Google Scholar 

  50. Mistafa O, Stenius U. Statins inhibit Akt/PKB signaling via P2X7 receptor in pancreatic cancer cells. Biochem Pharmacol. 2009;78(9):1115–26.

    Article  CAS  PubMed  Google Scholar 

  51. Lee YM, Chen HW, Maurya PK, Su CM, Tzeng CR. MicroRNA regulation via DNA methylation during the morula to blastocyst transition in mice. Mol Hum Reprod. 2012;18(4):184–93.

    Article  CAS  PubMed  Google Scholar 

  52. Engel T, Lorkowski S, Lueken A, Rust S, Schluter B, Berger G, et al. The human ABCG4 gene is regulated by oxysterols and retinoids in monocyte-derived macrophages. Biochem Biophys Res Commun. 2001;288(2):483–8.

    Article  CAS  PubMed  Google Scholar 

  53. Murphy AJ, Bijl N, Yvan-Charvet L, Welch CB, Bhagwat N, Reheman A, et al. Cholesterol efflux in megakaryocyte progenitors suppresses platelet production and thrombocytosis. Nat Med. 2013;19(5):586–94.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  54. Fan HM, Sun XY, Guo W, Zhong AF, Niu W, Zhao L, et al. Differential expression of microRNA in peripheral blood mononuclear cells as specific biomarker for major depressive disorder patients. J Psychiatr Res. 2014;59:45–52.

    Article  CAS  PubMed  Google Scholar 

  55. Bianchessi V, Badi I, Bertolotti M, Nigro P, D’Alessandra Y, Capogrossi MC, et al. The mitochondrial lncRNA ASncmtRNA-2 is induced in aging and replicative senescence in endothelial cells. J Mol Cell Cardiol. 2015;81:62–70.

    Article  CAS  PubMed  Google Scholar 

  56. Xu W, Liu M, Peng X, Zhou P, Zhou J, Xu K, et al. miR-24-3p and miR-27a-3p promote cell proliferation in glioma cells via cooperative regulation of MXI1. Int J Oncol. 2013;42(2):757–66.

    PubMed  Google Scholar 

  57. Camps C, Saini HK, Mole DR, Choudhry H, Reczko M, Guerra-Assuncao JA, et al. Integrated analysis of microRNA and mRNA expression and association with HIF binding reveals the complexity of microRNA expression regulation under hypoxia. Mol Cancer. 2014;13:28.

    Article  PubMed Central  PubMed  Google Scholar 

  58. Lewis GF, Rader DJ. New insights into the regulation of HDL metabolism and reverse cholesterol transport. Circ Res. 2005;96(12):1221–32.

    Article  CAS  PubMed  Google Scholar 

  59. Vaughan AM, Oram JF. ABCA1 and ABCG1 or ABCG4 act sequentially to remove cellular cholesterol and generate cholesterol-rich HDL. J Lipid Res. 2006;47(11):2433–43.

    Article  CAS  PubMed  Google Scholar 

  60. Wang N, Lan D, Chen W, Matsuura F, Tall AR. ATP-binding cassette transporters G1 and G4 mediate cellular cholesterol efflux to high-density lipoproteins. Proc Natl Acad Sci U S A. 2004;101(26):9774–9.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  61. Barter PJ, Brandrup-Wognsen G, Palmer MK, Nicholls SJ. Effect of statins on HDL-C: a complex process unrelated to changes in LDL-C: analysis of the VOYAGER database. J Lipid Res. 2010;51(6):1546–53.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  62. Xiang J, Wu J. Feud or friend? The role of the miR-17-92 cluster in tumorigenesis. Curr Genomics. 2010;11(2):129–35.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  63. Mogilyansky E, Rigoutsos I. The miR-17/92 cluster: a comprehensive update on its genomics, genetics, functions and increasingly important and numerous roles in health and disease. Cell Death Differ. 2013;20(12):1603–14.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  64. Hayashita Y, Osada H, Tatematsu Y, Yamada H, Yanagisawa K, Tomida S, et al. A polycistronic microRNA cluster, miR-17-92, is overexpressed in human lung cancers and enhances cell proliferation. Cancer Res. 2005;65(21):9628–32.

    Article  CAS  PubMed  Google Scholar 

  65. Cloonan N, Brown MK, Steptoe AL, Wani S, Chan WL, Forrest AR, et al. The miR-17-5p microRNA is a key regulator of the G1/S phase cell cycle transition. Genome Biol. 2008;9(8):R127.

    Article  PubMed Central  PubMed  Google Scholar 

  66. Ma Y, Zhang P, Wang F, Zhang H, Yang Y, Shi C, et al. Elevated oncofoetal miR-17-5p expression regulates colorectal cancer progression by repressing its target gene P130. Nat Commun. 2012;3:1291.

    Article  PubMed  Google Scholar 

  67. Fang L, Li H, Wang L, Hu J, Jin T, Wang J, et al. MicroRNA-17-5p promotes chemotherapeutic drug resistance and tumour metastasis of colorectal cancer by repressing PTEN expression. Oncotarget. 2014;5(10):2974–87.

    Article  PubMed Central  PubMed  Google Scholar 

  68. Yang X, Du WW, Li H, Liu F, Khorshidi A, Rutnam ZJ, et al. Both mature miR-17-5p and passenger strand miR-17-3p target TIMP3 and induce prostate tumor growth and invasion. Nucleic Acids Res. 2013;41(21):9688–704.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  69. Hossain A, Kuo MT, Saunders GF. Mir-17-5p regulates breast cancer cell proliferation by inhibiting translation of AIB1 mRNA. Mol Cell Biol. 2006;26(21):8191–201.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This investigation was supported by grants from CNPq-Brazil (N° 473485/2012-5), FAPESP-Brazil (N° 2011/21967-1) and FONDECYT-Chile (N°1130675). T. Z. and A. C. were recipients of fellowships from CONICYT-Chile. M.H.H. and R.D.C.H. are recipients of fellowships from CNPq-Brazil.

Author information

Authors and Affiliations

  1. Center of Molecular Biology and Pharmacogenetics, Scientific and Technological Bioresource Nucleus (BIOREN), Universidad de La Frontera, Temuco, Chile

    Tomás Zambrano, Álvaro Cerda & Luis A. Salazar

  2. School of Pharmaceutical Sciences, University of Sao Paulo, São Paulo, Brazil

    Tomás Zambrano, Rosario D.C. Hirata, Mario H. Hirata & Álvaro Cerda

  3. Centro de Biología Molecular & Farmacogenética, Departamento de Ciencias Básicas, Facultad de Medicina, Universidad de La Frontera, Av. Francisco Salazar 01145, Casilla 54-D, Temuco, Chile

    Luis A. Salazar

Authors
  1. Tomás Zambrano
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rosario D.C. Hirata
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mario H. Hirata
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Álvaro Cerda
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Luis A. Salazar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Luis A. Salazar.

Electronic supplementary material

Supplementary Figure 1

Relative expression of genes involved in cholesterol synthesis and cholesterol efflux following 10 μM atorvastatin treatment in HepG2 cells. (ABCG1, P < 0.05; ABCA1, P < 0.05; HMGCR, P = NS; LDLR, P = NS; SREBF-1, P = NS; SREBF-2, P = NS) (DOCX 94 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zambrano, T., Hirata, R.D., Hirata, M.H. et al. Altered microRNome Profiling in Statin-Induced HepG2 Cells: A Pilot Study Identifying Potential new Biomarkers Involved in Lipid-Lowering Treatment. Cardiovasc Drugs Ther 29, 509–518 (2015). https://doi.org/10.1007/s10557-015-6627-0

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10557-015-6627-0

Keywords

Search

Navigation