title: Disruption of interleukin-1β autocrine signaling rescues complex I activity and improves ROS levels in immortalized epithelial cells with impaired cystic fibrosis transmembrane conductance regulator (CFTR) function.
abstract: Patients with cystic fibrosis (CF) have elevated concentration of cytokines in sputum and a general inflammatory condition. In addition, CF cells in culture produce diverse cytokines in excess, including IL-1β. We have previously shown that IL-1β, at low doses (∼30 pM), can stimulate the expression of CFTR in T84 colon carcinoma cells, through NF-κB signaling. However, at higher doses (>2.5 ng/ml, ∼150 pM), IL-1β inhibit CFTR mRNA expression. On the other hand, by using differential display, we found two genes with reduced expression in CF cells, corresponding to the mitochondrial proteins CISD1 and MTND4. The last is a key subunit for the activity of mitochondrial Complex I (mCx-I); accordingly, we later found a reduced mCx-I activity in CF cells. Here we found that IB3-1 cells (CF cells), cultured in serum-free media, secrete 323±5 pg/ml of IL-1β in 24 h vs 127±3 pg/ml for S9 cells (CFTR-corrected IB3-1 cells). Externally added IL-1β (5 ng/ml) reduces the mCx-I activity and increases the mitochondrial (MitoSOX probe) and cellular (DCFH-DA probe) ROS levels of S9 (CFTR-corrected IB3-1 CF cells) or Caco-2/pRSctrl cells (shRNA control cells) to values comparable to those of IB3-1 or Caco-2/pRS26 cells (shRNA specific for CFTR). Treatments of IB3-1 or Caco-2/pRS26 cells with either IL-1β blocking antibody, IL-1 receptor antagonist, IKK inhibitor III (NF-κB pathway) or SB203580 (p38 MAPK pathway), restored the mCx-I activity. In addition, in IB3-1 or Caco-2/pRS26 cells, IL-1β blocking antibody, IKK inhibitor III or SB203580 reduced the mitochondrial ROS levels by ∼50% and the cellular ROS levels near to basal values. The AP-1 inhibitors U0126 (MEK1/2) or SP600125 (JNK1/2/3 inhibitor) had no effects. The results suggest that in these cells IL-1β, through an autocrine effect, acts as a bridge connecting the CFTR with the mCx-I activity and the ROS levels.
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: Mariángeles
authors.initials: M
authors.lastname: Clauzure
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: Angel G
authors.initials: AG
authors.lastname: Valdivieso
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: María M
authors.initials: MM
authors.lastname: Massip Copiz
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Schulman
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: María Luz
authors.initials: ML
authors.lastname: Teiber
authors.affiliation: Laboratory of Cellular and Molecular Biology, Institute for Biomedical Research (BIOMED), School of Medical Sciences, Pontifical Catholic University of Argentina (UCA), and the National Scientific and Technical Research Council (CONICET), Buenos Aires, Argentina.
authors.forename: Tomás A
authors.initials: TA
authors.lastname: Santa-Coloma
mesh_terms: D000906:Antibodies / Q000276:immunology; D019898:Autocrine Communication / Q000187:drug effects; D018938:Caco-2 Cells; D002460:Cell Line; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D042967:Electron Transport Complex I / Q000378:metabolism*; D014450:Electron Transport Complex III / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D053583:Interleukin-1beta / Q000276:immunology / Q000378:metabolism*; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D017472:Receptors, Interleukin-1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D000906:Antibodies; D016895:Culture Media, Serum-Free; D007093:Imidazoles; D053583:Interleukin-1beta; D016328:NF-kappa B; D011725:Pyridines; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D017472:Receptors, Interleukin-1; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D042967:Electron Transport Complex I; D014450:Electron Transport Complex III; C093642:SB 203580