Small RNA and transcriptome sequencing reveal the role of miR‐199a‐3p in inflammatory processes in cystic fibrosis airways

Abstract

Cystic fibrosis (CF) is the most common lethal genetic disease, caused by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations. CF is characterized by an ionic imbalance and thickened mucus, which impair mucociliary clearance and promote bacterial colonization and the establishment of infection/inflammation cycles. However, the origin of this inflammation remains unclear, although microRNAs (miRNAs) are suspected to be involved. MiRNAs are small non-coding RNAs that bind to the 3'-untranslated regions (UTRs) of target gene mRNA, thereby repressing their translation and/or inducing their degradation. The goal of this study was to investigate the role of microRNAs associated with pulmonary inflammation in CF patients. Through the analysis of all miRNAs (miRNome) in human primary air-liquid interface cultures, we demonstrated that miR-199a-3p is the only miRNA downregulated in CF patients compared to controls. Moreover, through RNA sequencing (transcriptome) analysis, we showed that 50% of all deregulated mRNAs are linked directly or indirectly to the NF-κB pathway. To identify a specific target, we used bioinformatics analysis to predict whether miR-199a-3p targets the 3'-UTR of IKBKB, which encodes IKKβ, a major protein in the NF-κB pathway. Subsequently, we used bronchial explants from CF patients to show that miR-199a-3p expression is downregulated compared to controls and inversely correlated with increases in expression of IKKβ and IL-8. Through functional studies, we showed that miR-199a-3p modulates the expression of IKBKB through a direct interaction at its 3'-UTR in bronchial epithelial cells from CF patients. In miR-199a-3p overexpression experiments, we demonstrated that for CF cells, miR-199a-3p reduced IKKβ protein expression, NF-κB activity, and IL-8 secretion. Taken together, our findings show that miR-199a-3p plays a negative regulatory role in the NF-κB signalling pathway and that its low expression in CF patients contributes to chronic pulmonary inflammation. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Interstitial microRNA miR-214 attenuates inflammation and polycystic kidney disease progression

“…Nevertheless, the full scope of the biological pathways regulated by miR-214 in ADPKD is not known. Finally, miR-199a, which is cotranscribed with miR-214, and the host Dnm3os lncRNA itself, are implicated in inflammation (20,45). To what extent they modulate the ADPKD phenotype is unknown.…”

Emerging microRNA Therapeutic Approaches for Cystic Fibrosis

“…The miR-199a-3p expression in the airway, which is decreased in CF context, is associated with an NF-κB hyperactivation. In fact, miR-199a-3p directly regulates IKKβ, which acts on the NF-κB signaling pathway and therefore on IL-8 secretion, the main cytokine dysregulated in the airways of CF patients ( Bardin et al, 2018 ). Moreover, miR-509-3p, miR-494, and miR-126 directly target NF-κB and have shown that CFTR expression and function decreases when NF-κB is functional ( McKiernan et al, 2013 ; Ramachandran et al, 2013 ).…”

Plasma microRNA levels in male and female children with cystic fibrosis

“…miR-17 targets IL-8, and we previously reported decreased levels of miR-17 in CF bronchial brushings 34 , and later suggested how manipulation of miR-17 in CF bronchial epithelial cells could have therapeutic benefit 35 . A recent report by Bardin et al pointed to a role for miR-199a-3p in the chronic inflammation of the CF airway 36 . miR-199a-3p was found downregulated in primary air-liquid interface cultures and bronchial explants from CF versus non-CF controls, and was demonstrated to regulate the expression of inhibitor of nuclear factor kappa-B kinase subunit beta (IKK-β), a signalling intermediary of the NF-κB pathway 36 .…”

“…miR-199a-3p was found downregulated in primary air-liquid interface cultures and bronchial explants from CF versus non-CF controls, and was demonstrated to regulate the expression of inhibitor of nuclear factor kappa-B kinase subunit beta (IKK-β), a signalling intermediary of the NF-κB pathway 36 . No significant difference in expression of miR-199a-3p was found in CF versus non-CF plasma, however the authors have not stated the sex of the patients in each group 36 . Larger cohort studies could confirm whether these miRNAs represent useful clinical biomarkers.…”