5-methyl-1-phenyl-2-(1H)-pyridone reveals immunmodulatory anti-fibrotic effects associated to antioxidant system activation in primary culture of human Hepatic Stellate Cells
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1
Universidad de Guadalajara, Laboratorio de Investigación en Ciencias Biomédicas y Biotecnología. CUValles, Mexico
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2
Universidad de Guadalajara, Instituto de Biología Molecular en Medicina y Terapia Génica. CUCS, Mexico
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3
Università degli Studi di Firenze, Center for Research, High Education and Transfer DENOThe, Italy
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4
Università degli Studi di Torino, Dipartimento di Medicina e Oncologia Sperimentale, Italy
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5
University Collage London, Institute for Liver and Digestive Health, United Kingdom
Background. Hepatic stellate cells (HSC) profibrogenic cytokines are key targets of anti-fibrotic therapies. 5-methyl-1-phenyl-2-(1H)-pyridone or pirfenidone (PFD) is a small molecule indicated for treatment of chronic inflammation and fibrogenesis. Oxidative stress is directly involved in the onset of hepatic fibrosis by HSC activation. Aim. In order to identify whether anti-inflammatory and anti-fibrogenic effects of PFD are related to activation of the endogenus antioxidant system, HSC were incubated with PDGF or 2-methyl-1,4-naphthoquinone (MEN) a ROS-inducer. Methods and Results. PFD was able to inhibit PDGF or MEN-induced pro-fibrogenic actions, including cell proliferation, cell motility and de novo synthesis of Collagen type I, TGFβ, TIMP-1, IL-1 and TNFα. These effects were associated with an increase of nuclear Nrf2 assessed by western blotting and confocal microscopy. Because PFD activates JNK, which stimulates Nrf2 transcriptional factor, through siRNA-mediated silencing we examined downstream antioxidant targets as antioxidant enzimes. JNK blockade by siRNA and SP600125 down-regulates Nrf2 activation. Also PFD induced a dose- and time-dependent activation of several antioxidant genes, (Glutamyl cysteine synthetase catalytic subunit, Glutamyl cysteine synthetase regulatory subunit and Heme oxygenase 1) and increase glutathione content, whose activity may contribute to the down-regulation of ROS-induced pro-fibrogenic and pro-inflammatory effects. Conclusion. These results provide molecular insights in anti-fibrogenic immunmodulatory action of PFD by counteracting ROS-induced pro-fibrogenic signalling, and by regulation of the biosynthesis of antioxidant proteins. This study indicates that activation of the antioxidant system plays an essential role in the modulation of inflamatory and fibrogenic cytokines in HSC.
Keywords:
Hepatic Stellate Cells,
TGF-β,
Glutathione,
Pirfenidone,
Inflammation,
Antioxidant Enzymes
Conference:
15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013.
Presentation Type:
Abstract
Topic:
Immune-mediated disease pathogenesis
Citation:
Macias-Barragan
J,
Caligiuri
A,
Novo
E,
Parola
M,
Vera-Cruz
J,
García-Bañuelos
J,
Pinzani
M and
Armendáriz-Borunda
J
(2013). 5-methyl-1-phenyl-2-(1H)-pyridone reveals immunmodulatory anti-fibrotic effects associated to antioxidant system activation in primary culture of human Hepatic Stellate Cells.
Front. Immunol.
Conference Abstract:
15th International Congress of Immunology (ICI).
doi: 10.3389/conf.fimmu.2013.02.00087
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Received:
08 Mar 2013;
Published Online:
22 Aug 2013.
*
Correspondence:
Dr. Jose Macias-Barragan, Universidad de Guadalajara, Laboratorio de Investigación en Ciencias Biomédicas y Biotecnología. CUValles, Ameca, Jalisco, 46600, Mexico, josemacias@libero.it