title: Integrin-linked kinase mediates the hydrogen peroxide-dependent transforming growth factor-β1 up-regulation.
abstract: Transforming growth factor type-β1 (TGF-β1) has been recognized as a central mediator in many pathological events related to extracellular matrix (ECM) proteins accumulation, where their locally increased expression has been implicated in the fibrosis process of numerous organs, including glomerular fibrosis in the kidney. We and others have reported the TGF-β1 synthesis regulation by reactive oxygen species (ROS), and moreover we also described the implication of integrin-linked kinase (ILK) in the AP-1-dependent TGF-β1 up-regulation. Thus, we propose here that hydrogen peroxide (H2O2)-dependent TGF-β1 regulation may be mediated by ILK activation. First we confirmed the increase in TGF-β1 expression in human mesangial cells (HMC) after treatment with H2O2 or with an alternative H2O2-generating system such as the glucose-oxidase enzyme (GOX). By using immunoblotting, immunofluorescence, and ELISA techniques, we demonstrate that extracellular H2O2 up-regulates TGF-β1 transcription, as well as increases TGF-β1 promoter activity. Furthermore, catalase-decreased intracellular H2O2 abolished TGF-β1 up-regulation. The use of pharmacological inhibitors as well as knockdown of ILK with small interfering RNA (siRNA) demonstrated the implication of a PI3K/ILK/AKT/ERK MAPK signaling pathway axis in the H2O2-induced TGF-β1 overexpression. Finally, we explored the physiological relevance of these findings by treating HMC with angiotensin II, a known stimuli of H2O2 synthesis. Our results confirm the relevance of previous findings after a more physiological stimulus. In summary, our results provide evidence that ILK activity changes may act as a mechanism in response to different stimuli such as H2O2 in the induced TGF-β1 up-regulation in pathological or even physiological conditions.
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Gonzalez-Ramos
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain.
authors.forename: S
authors.initials: S
authors.lastname: de Frutos
authors.affiliation: IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain; Biomedical Research Unit Foundation, Hospital Universitario Príncipe de Asturias, Alcalá de Henares, Madrid, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Griera
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Luengo
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain.
authors.forename: G
authors.initials: G
authors.lastname: Olmos
authors.affiliation: IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain; Department of Medicine, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; Nephrology Section, Hospital Universitario Príncipe de Asturias, Alcalá de Henares, Madrid, Spain.
authors.forename: D
authors.initials: D
authors.lastname: Rodriguez-Puyol
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain. Electronic address: laura.calleros@uah.es.
authors.forename: L
authors.initials: L
authors.lastname: Calleros
authors.affiliation: Department of Physiology, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain; IRSIN, Madrid, Spain; RedInRen (Instituto de Salud Carlos III), Madrid, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Rodriguez-Puyol
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology; D002478:Cells, Cultured; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology; D005949:Glucose Oxidase / Q000502:physiology; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis*; D015854:Up-Regulation; D000099200:Scaffold Protein ILK
chemical_list: D000804:Angiotensin II; D048049:Extracellular Signal-Regulated MAP Kinases; D005949:Glucose Oxidase; D006861:Hydrogen Peroxide; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D053773:Transforming Growth Factor beta1; D000099200:Scaffold Protein ILK
keywords: AKT; Angiotensin II; ERK; ILK; ROS; TGF-β1