Reactive oxygen species via redox signaling to PI3K/AKT pathway contribute to the malignant growth of 4-hydroxy estradiol-transformed mammary epithelial cells

PLoS One. 2013;8(2):e54206. doi: 10.1371/journal.pone.0054206. Epub 2013 Feb 21.

Abstract

The purpose of this study was to investigate the effects of 17-β-estradiol (E2)-induced reactive oxygen species (ROS) on the induction of mammary tumorigenesis. We found that ROS-induced by repeated exposures to 4-hydroxy-estradiol (4-OH-E2), a predominant catechol metabolite of E2, caused transformation of normal human mammary epithelial MCF-10A cells with malignant growth in nude mice. This was evident from inhibition of estrogen-induced breast tumor formation in the xenograft model by both overexpression of catalase as well as by co-treatment with Ebselen. To understand how 4-OH-E2 induces this malignant phenotype through ROS, we investigated the effects of 4-OH-E2 on redox-sensitive signal transduction pathways. During the malignant transformation process we observed that 4-OH-E2 treatment increased AKT phosphorylation through PI3K activation. The PI3K-mediated phosphorylation of AKT in 4-OH-E2-treated cells was inhibited by ROS modifiers as well as by silencing of AKT expression. RNA interference of AKT markedly inhibited 4-OH-E2-induced in vitro tumor formation. The expression of cell cycle genes, cdc2, PRC1 and PCNA and one of transcription factors that control the expression of these genes - nuclear respiratory factor-1 (NRF-1) was significantly up-regulated during the 4-OH-E2-mediated malignant transformation process. The increased expression of these genes was inhibited by ROS modifiers as well as by silencing of AKT expression. These results indicate that 4-OH-E2-induced cell transformation may be mediated, in part, through redox-sensitive AKT signal transduction pathways by up-regulating the expression of cell cycle genes cdc2, PRC1 and PCNA, and the transcription factor - NRF-1. In summary, our study has demonstrated that: (i) 4-OH-E2 is one of the main estrogen metabolites that induce mammary tumorigenesis and (ii) ROS-mediated signaling leading to the activation of PI3K/AKT pathway plays an important role in the generation of 4-OH-E2-induced malignant phenotype of breast epithelial cells. In conclusion, ROS are important signaling molecules in the development of estrogen-induced malignant breast lesions.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Azoles / pharmacology
  • Catalase / metabolism
  • Catechols / metabolism
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • Cell Proliferation / drug effects
  • Cell Transformation, Neoplastic / drug effects
  • Cell Transformation, Neoplastic / pathology*
  • Collagen / pharmacology
  • Colony-Forming Units Assay
  • Dose-Response Relationship, Drug
  • Epithelial Cells / enzymology*
  • Epithelial Cells / pathology
  • Estradiol / analogs & derivatives
  • Estradiol / pharmacology
  • Estrogens, Catechol / pharmacology*
  • Fulvestrant
  • Gene Expression Regulation, Neoplastic / drug effects
  • Humans
  • Mammary Glands, Human / drug effects
  • Mammary Glands, Human / enzymology
  • Mammary Glands, Human / pathology
  • Mice
  • Models, Biological
  • Neoplasm Invasiveness
  • Organoselenium Compounds / pharmacology
  • Oxidation-Reduction / drug effects
  • Phenotype
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Proto-Oncogene Proteins c-akt / metabolism*
  • Reactive Oxygen Species / metabolism*
  • Signal Transduction / drug effects*
  • Signal Transduction / genetics
  • Spheroids, Cellular / drug effects
  • Spheroids, Cellular / metabolism
  • Spheroids, Cellular / pathology

Substances

  • Azoles
  • Catechols
  • Cell Cycle Proteins
  • Estrogens, Catechol
  • Organoselenium Compounds
  • Reactive Oxygen Species
  • Fulvestrant
  • ebselen
  • Estradiol
  • Collagen
  • 4-hydroxyestradiol
  • Catalase
  • Phosphatidylinositol 3-Kinases
  • Proto-Oncogene Proteins c-akt