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.

Intracellular Chloride Concentration Changes Modulate IL-1β Expression and Secretion in Human Bronchial Epithelial Cultured Cells

“…We have previously reported that the expression IL‐1β was increased approximately 40% in IB3‐1 CF cells compared to CFTR‐corrected S9 cells [Clauzure et al, ]. These effects of CFTR over IL‐1β expression could be due to the CFTR Cl − transport activity or to the presence/absence of the CFTR at the cell membrane, independently of its transport activity, as appears to occur with RANTES [Estell et al, ].…”

“…Several genes are differentially expressed in cells with impaired CFTR activity [Gonzalez‐Guerrico et al, ; Valdivieso et al, , ; Taminelli et al, ; Valdivieso and Santa‐Coloma, ; Clauzure et al, ; Massip Copiz and Santa‐Coloma, ], including IL‐1β [Clauzure et al, ; Grassme et al, ]. Thus, by using the IL‐1β expression levels of IB3‐1 CF epithelial cells as a model system (these cells over‐express basal IL‐1β levels [Clauzure et al, ]), here, we show that changes in the intracellular Cl − concentrations modulate the steady‐state levels of IL‐1β mRNA, with similar effects on the secreted protein. The effect of Cl − on the IL‐1β mRNA steady state levels disappear in the presence of the IL‐1β receptor antagonist IL1RN or in the presence of IL‐1β blocking antibody.…”

An unexpected effect of TNF-α on F508del-CFTR maturation and function

“…TNFα induces IL-8 and IL-1β synthesis and secretion from adenocarcinoma lung cancer cells, Calu-3 cells 1 . IL-1β has recently been reported to stimulate the expression of CFTR in T84 colon carcinoma cells, through NF-κB signalling 23 . For these reasons, possible involvement of other inflammatory mediators in the F508delCFTR response to TNFα could not be excluded.…”

The chloride anion as a signalling effector

“…Nunes et al (2015) reported direct effects of Cl − on mitochondrial membrane potential and ATP production. Recent work has shown that IL-1β, which is also produced by a Cl − -dependent gene, is involved in the reduction of mitochondrial Complex I and increased ROS activity in cells with impaired CFTR activity, cells that have an increased [Cl − ] i since their anion extrusion is affected (Valdivieso et al, 2012;Valdivieso & Santa-Coloma, 2013;Clauzure et al, 2014Clauzure et al, , 2017Massip-Copiz et al, 2017).…”