Exogenous Reactive Oxygen Species Augments SMAD4 Expression And TGF-β Paradox in Human Breast Cancer
Abstract
Background: The Transforming growth factor-β (TGF-β) functions to induce apoptosis, cell cycle arrest, and differentiation is central to sustaining tissue homeostasis and maintaining genomic stability. TGF-β normally, an effective tumor-suppressor that restricts the uncontrolled division of cells augments the development and progression of human malignancies when cytostatic activities of TGF-β are resisted by genetic and epigenetic events caused by tumorigenesis. This dichotomic nature of TGF-β during oncogenesis termed as “TGF-β Paradox,” persists to be the most crucial and puzzling query regarding its physio-pathological function and the role of cellular antioxidant status is highly interrelated which warrants more studies on the role of endogenous reactive oxygen species (ROS) in deciding epithelial-mesenchymal transition (EMT) process. The objective of the study was to check whether enhanced ROS augments the TGF-β pathway facilitating EMT.
Methods: In vitro toxicity assay was performed to assess the appropriate concentration of hydrogen peroxide (H2O2) imparting oxidative stress. Comet assay and 8-OHdG (8-hydroxy-2’-deoxyguanosine) enzyme-linked immunosorbent assay (ELISA) were performed to check the extent of DNA damage and adduct production respectively. Mitogen-activated protein kinase (MAPK) p38 ELISA and mRNA gene expression analysis of TGF-β and SMAD were done to verify the effect of H2O2 on these signaling.
Results: The objective of the study was to check whether enhanced ROS augments the TGF-β pathway facilitating EMT. Along with morphological alterations, a dose-dependent decrease in cell viability was seen at 300µM of H2O2 compared to 75µM. DCFDA labeling discovered the dose-dependent gradation of intracellular ROS generation and this was correlated to increased cellular DNA damage and DNA adduct production which was increased linearly with increasing H2O2 as evident with comet test and 8-OHdG ELISA. Significantly reduced MAPK p38 activity revealed by indirect ELISA analysis suggests lessened suppression of cell growth.
Conclusions: The study establishes that higher intracellular ROS will facilitate the TGF-β paradox leading to epithelial mesenchymal transition which can adversely affect therapeutic strategies targeting EMT
Keywords
DOI: 10.33371/ijoc.v18i3.1179
Article Metrics
Abstract View: 149, 
PDF Download: 87
References
Akram M, Iqbal M, Daniyal M, Khan AU. Awareness and current knowledge of breast cancer. Biol Res [Internet]. 2017 [cited 2024 Jan 11];50. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5625777/
Sun YS, Zhao Z, Yang ZN, Xu F, Lu HJ, Zhu ZY, et al. Risk Factors and Preventions of Breast Cancer. Int J Biol Sci. 2017;13(11):1387.
Wendt MK, Tian M, Schiemann WP. Deconstructing the mechanisms and consequences of TGF-β-induced EMT during cancer progression. Cell Tissue Res. 2012 Jan 1;347(1):85–101.
Cui W, Fowlis DJ, Bryson S, Duffie E, Ireland H, Balmain A, et al. TGFβ1 Inhibits the Formation of Benign Skin Tumors, but Enhances Progression to Invasive Spindle Carcinomas in Transgenic Mice. Cell. 1996 Aug 23;86(4):531–42.
Gj I. Switching TGFβ from a tumor suppressor to a tumor promoter. Curr Opin Genet Dev [Internet]. 2011 Feb [cited 2024 Jan 11];21(1). Available from: https://pubmed.ncbi.nlm.nih.gov/21251810/
Lawrence DA. Transforming growth factor-beta: an overview. Kidney Int Suppl. 1995 Jun 1;49:S19-23.
Liu RM, Gaston Pravia KA. Oxidative stress and glutathione in TGF-β-mediated fibrogenesis. Free Radic Biol Med. 2010 Jan 1;48(1):1–15.
Ramundo V, Giribaldi G, Aldieri E. Transforming Growth Factor-β and Oxidative Stress in Cancer: A Crosstalk in Driving Tumor Transformation. Cancers. 2021 Jan;13(12):3093.
Y H, D B, P TD. TGF-β-Mediated Epithelial-Mesenchymal Transition and Cancer Metastasis. Int J Mol Sci [Internet]. 2019 Jun 5 [cited 2023 Aug 17];20(11). Available from: https://pubmed.ncbi.nlm.nih.gov/31195692/
Roberts AB, Wakefield LM. The two faces of transforming growth factor β in carcinogenesis. Proc Natl Acad Sci U S A. 2003 Jul 7;100(15):8621.
Jain M, Rivera S, Monclus EA, Synenki L, Zirk A, Eisenbart J, et al. Mitochondrial Reactive Oxygen Species Regulate Transforming Growth Factor-β Signaling. J Biol Chem. 2013 Jan 11;288(2):770–7.
Chung J, Huda MN, Shin Y, Han S, Akter S, Kang I, et al. Correlation between Oxidative Stress and Transforming Growth Factor-Beta in Cancers. Int J Mol Sci [Internet]. 2021 Dec [cited 2024 Jan 12];22(24). Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8707703/
Liu RM, Desai LP. Reciprocal regulation of TGF-β and reactive oxygen species: A perverse cycle for fibrosis. Redox Biol. 2015 Oct 10;6:565–77.
Hua W, ten Dijke P, Kostidis S, Giera M, Hornsveld M. TGFβ-induced metabolic reprogramming during epithelial-to-mesenchymal transition in cancer. Cell Mol Life Sci. 2020 Jun 1;77(11):2103–23.
Chang CH, Pauklin S. ROS and TGFβ: from pancreatic tumour growth to metastasis. J Exp Clin Cancer Res. 2021 May 3;40(1):152.
Mei S, Gu H, Ward A, Yang X, Guo H, He K, et al. p38 Mitogen-activated Protein Kinase (MAPK) Promotes Cholesterol Ester Accumulation in Macrophages through Inhibition of Macroautophagy. J Biol Chem. 2012 Apr 4;287(15):11761.
B K, S K, J P, L K, Js H, Rb M, et al. p38 mitogen-activated protein kinase-driven MAPKAPK2 regulates invasion of bladder cancer by modulation of MMP-2 and MMP-9 activity. Cancer Res [Internet]. 2010 Jan 15 [cited 2023 Aug 18];70(2). Available from: https://pubmed.ncbi.nlm.nih.gov/20068172/
A B, Jl MC, G SJ, C C, Mu M, J D, et al. A synthetic peptide from transforming growth factor-β₁ type III receptor inhibits NADPH oxidase and prevents oxidative stress in the kidney of spontaneously hypertensive rats. Antioxid Redox Signal [Internet]. 2013 Nov 10 [cited 2023 Aug 18];19(14). Available from: https://pubmed.ncbi.nlm.nih.gov/23350688/
Ramachandran R, Saraswathy M. Up-regulation of nuclear related factor 2 (NRF2) and antioxidant responsive elements by metformin protects hepatocytes against the acetaminophen toxicity. Toxicol Res. 2014 Jun 24;3(5):350.
Anuf AR, Ramachandran R, Krishnasamy R, Gandhi PSS, Periyasamy S. Antiproliferative effects of Plumbago rosea and its purified constituent plumbagin on SK-MEL 28 melanoma cell lines. Pharmacogn Res. 2014 Dec;6(4):312.
R R, M S. Postconditioning with metformin attenuates apoptotic events in cardiomyoblasts associated with ischemic reperfusion injury. Cardiovasc Ther [Internet]. 2017 Dec [cited 2024 Jan 12];35(6). Available from:
https://pubmed.ncbi.nlm.nih.gov/28643448/
M T, Wp S. The TGF-beta paradox in human cancer: an update. Future Oncol Lond Engl [Internet]. 2009 Mar [cited 2023 Sep 1];5(2). Available from: https://pubmed.ncbi.nlm.nih.gov/19284383/
Wu F, Weigel KJ, Zhou H, Wang XJ. Paradoxical roles of TGF-β signaling in suppressing and promoting squamous cell carcinoma. Acta Biochim Biophys Sin. 2018 Jan;50(1):98.
Krstić J, Trivanović D, Mojsilović S, Santibanez JF. Transforming Growth Factor-Beta and Oxidative Stress Interplay: Implications in Tumorigenesis and Cancer Progression. Oxid Med Cell Longev [Internet]. 2015 [cited 2023 Sep 2];2015. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4452864/
G VE, A A, M G, Ri AZ, J C. Molecular and Cellular Effects of Hydrogen Peroxide on Human Lung Cancer Cells: Potential Therapeutic Implications. Oxid Med Cell Longev [Internet]. 2016 [cited 2023 Sep 2];2016. Available from: https://pubmed.ncbi.nlm.nih.gov/27375834/
C F, Qm C, S Z, Jp M, J R, O T. Subcytotoxic H2O2 stress triggers a release of transforming growth factor-beta 1, which induces biomarkers of cellular senescence of human diploid fibroblasts. J Biol Chem [Internet]. 2001 Jan 26 [cited 2023 Sep 2];276(4). Available from: http
Refbacks
- There are currently no refbacks.
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
Email this article 
