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Am J Physiol Lung Cell Mol Physiol. 2014 Nov 15;307(10):L791-9. doi: 10.1152/ajplung.00167.2014. Epub 2014 Sep 19.

Impaired TLR4 and HIF expression in cystic fibrosis bronchial epithelial cells downregulates hemeoxygenase-1 and alters iron homeostasis in vitro.

Author information

1
Department of Medicine, Pulmonary Critical Care Philipps University, Marburg, Germany; Institute for Lung Research, Philipps-University, Marburg, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany; Member of the German Center for Lung Research (DZL);
2
Department of Internal Medicine, Justus-Liebig-University, Giessen, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany; Member of the German Center for Lung Research (DZL);
3
Clinical Research Group 'Molecular Pathology of Cystic Fibrosis and Pseudomonas Genomics', Clinic for Pediatric Pneumology, Allergology and Neonatology, Hannover Medical School, Hannover, Germany; Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Germany;
4
Department of Chemistry-Biochemistry, Philipps University, Marburg, Germany;
5
Department of Medicine, Pulmonary Critical Care Philipps University, Marburg, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany;
6
Department of Internal Medicine, Justus-Liebig-University, Giessen, Germany; Lung Clinic Waldhof-Elgershausen, Greifenstein, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany; Member of the German Center for Lung Research (DZL);
7
Institute for Lung Research, Philipps-University, Marburg, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany; Member of the German Center for Lung Research (DZL);
8
Department of Medicine, Pulmonary Critical Care Philipps University, Marburg, Germany; Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, Germany; Member of the German Center for Lung Research (DZL); Pneumology, Asklepios Fachkliniken München-Gauting, Germany; Comprehensive Pneumology Center (CPC), Helmholtz Zentrum, Munich, Germany m.henke@asklepios.com.

Abstract

Hemeoxygenase-1 (HO-1), an inducible heat shock protein, is upregulated in response to multiple cellular insults via oxidative stress, lipopolysaccharides (LPS), and hypoxia. In this study, we investigated in vitro the role of Toll-like receptor 4 (TLR4), hypoxia-inducible factor 1α (HIF-1α), and iron on HO-1 expression in cystic fibrosis (CF). Immunohistochemical analysis of TLR4, HO-1, ferritin, and HIF-1α were performed on lung sections of CFTR-/- and wild-type mice. CFBE41o- and 16HBE14o- cell lines were employed for in vitro analysis via immunoblotting, immunofluorescence, real-time PCR, luciferase reporter gene analysis, and iron quantification. We observed a reduced TLR4, HIF-1α, HO-1, and ferritin in CFBE41o- cell line and CF mice. Knockdown studies using TLR4-siRNA in 16HBE14o- revealed significant decrease of HO-1, confirming the role of TLR4 in HO-1 downregulation. Inhibition of HO-1 using tin protoporphyrin in 16HBE14o- cells resulted in increased iron levels, suggesting a probable role of HO-1 in iron accumulation. Additionally, sequestration of excess iron using iron chelators resulted in increased hypoxia response element response in CFBE41o- and 16HBE14o-, implicating a role of iron in HIF-1α stabilization and HO-1. To conclude, our in vitro results demonstrate that multiple regulatory factors, such as impaired TLR4 surface expression, increased intracellular iron, and decreased HIF-1α, downregulate HO-1 expression in CFBE41o- cells.

KEYWORDS:

Toll-like receptor 4; cystic fibrosis; hypoxia-inducible factor 1α; iron

PMID:
25239913
DOI:
10.1152/ajplung.00167.2014
[Indexed for MEDLINE]
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