Toll like Receptor signalling by Prevotella histicola activates alternative NF-κB signalling in Cystic Fibrosis bronchial epithelial cells compared to P. aeruginosa

Abstract

Cystic Fibrosis (CF), caused by mutations affecting the CFTR gene, is characterised by viscid secretions in multiple organ systems. CF airways contain thick mucus, creating a gradient of hypoxia, which promotes the establishment of polymicrobial infection. Such inflammation predisposes to further infection, a self-perpetuating cycle in mediated by NF-κB. Anaerobic Gram-negative Prevotella spp. are found in sputum from healthy volunteers and CF patients and in CF lungs correlate with reduced levels of inflammation. Prevotella histicola (P. histicola) can suppress murine lung inflammation, however, no studies have examined the role of P. histicola in modulating infection and inflammation in the CF airways. We investigated innate immune signalling and NF-kB activation in CF epithelial cells CFBE41o-in response to clinical stains of P. histicola and Pseudomonas aeruginosa (P. aeruginosa). Toll-Like Receptor (TLR) expressing HEK-293 cells and siRNA assays for TLRs and IKKα were used to confirm signalling pathways. We show that P. histicola infection activated the alternative NF-kB signalling pathway in CF bronchial epithelial cells inducing HIF-1α protein. TLR5 signalling was responsible for the induction of the alternative NF-kB pathway through phosphorylation of IKKα. The induction of transcription factor HIF-1α was inversely associated with the induction of the alternative NF-kB pathway and knockdown of IKKα partially restored canonical NF-kB activation in response to P. histicola. This study demonstrates that different bacterial species in the respiratory microbiome can contribute differently to inflammation, either by activating inflammatory cascades (P. aeruginosa) or by muting the inflammatory response by modulating similar or related pathways (P. histicola). Further work is required to assess the complex interactions of the lung microbiome in response to mixed bacterial infections and their effects in people with CF.

Genomic diversity and antimicrobial resistance of Prevotella species isolated from chronic lung disease airways

“…However, their precise role in airway disease is poorly understood due to conflicting reports about their beneficial vs. pathogenic potential. For instance, P. melaninogenica, P. salivae and P. nanceiensis may inhibit non-typeable Haemophilus influenzae-induced pro-inflammatory cytokine production by lung dendritic cells [53], increased P. melaninogenica prevalence in CF airways has been associated with improved lung function [15], and P. histicola can mute pro-inflammatory pathways triggered by P. aeruginosa in CF airways [57].…”

Genomic diversity and antimicrobial resistance of Prevotella spp. isolated from chronic lung disease airways

“…However, their precise role in airway disease is poorly understood due to conflicting reports about their beneficial vs. pathogenic potential. For instance, P. melaninogenica, P. salivae , and P. nanceiensis may inhibit non-typeable Haemophilus influenzae -induced pro-inflammatory cytokine production by lung dendritic cells 52 , increased P. melaninogenica prevalence in CF airways has been associated with improved lung function 15 , and P. histicola can mute pro-inflammatory pathways triggered by P. aeruginosa in CF airways 55 . In contrast, P. melaninogenica produced the greatest variety of short-chain fatty acids (associated with inflammation) amongst five tested anaerobes 18 , and P. denticola shares similarities in its lipid A moieties to P. intermedia (a known pathogenic species), pointing towards pro-inflammatory traits 56 .…”

Commensal Bacteria in the Cystic Fibrosis Airway Microbiome Reduce P. aeruginosa Induced Inflammation

“…It is known that airway epithelial cells have innate sensor functions and may detect microbial signals (Weitnauer et al, 2016). It was shown in a recent report that Rothia mucilaginosa could inhibit host NF-kB pathway activation in a human lung epithelial cell line (Rigauts et al, 2021), and another commensal Prevotella histicola was reported to be able to induce alternative NF-kB activation in CF bronchial epithelial cells (Bertelsen et al, 2020). Earlier work from us has shown that airway epithelial cells adjust their expression pattern of innate immune receptors with less TLR2 and CD36 being expressed, resulting in low reactivity towards microbes that preferentially activate through TLR1/2 or TLR2/6.…”