Dataset Information

MUC5B induces in vitro neutrophil extracellular trap formation: Implication in otitis media.

ABSTRACT:

Background

Chronic otitis media (COM) is characterized by middle ear fluid predominantly containing cytokines, Nontypeable haemophilus influenzae (NTHi), the mucin MUC5B, and neutrophil extracellular traps (NETs). NETs consist of extracellular DNA coated with antibacterial proteins such as myeloperoxidase (MPO) and citrullinated histone 3 (CitH3). NETs can damage tissues and sustain inflammation. Our study aimed to develop an in vitro model of NETosis, testing COM inductors.

Methods

NETosis was evaluated in fresh blood human neutrophils attached to collagen-coated plates and in suspension exposed to phorbol myristate acetate (PMA) as a control, and COM relevant mediators. Confocal microscopy, DNA fluorescence assay and flow cytometry were used to quantify NETosis.

Results

PMA exposure induced DNA, MPO, and CitH3 by immunofluorescence (IF) most significantly at 3 hours (3.8-fold for DAPI, 7.6-fold for MPO, and 6.9-fold for CitH3, all P?P?ConclusionWe developed a reliable in vitro assay for NETosis which demonstrated that salivary MUC5B is a potent inductor of NETs whereas IL-8, TNF-?, live and lyzed NTHi demonstrated minimal to no NETosis.

Level of evidence

NA.

SUBMITTER: Val S

PROVIDER: S-EPMC7314476 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

MUC5B induces in vitro neutrophil extracellular trap formation: Implication in otitis media.

Val Stéphanie S Krueger Anna A Hussain Arman A Tomney Amarel A Chen Yajun Y Lazarski Christopher C Preciado Diego D

Laryngoscope investigative otolaryngology 20200523 3

<h4>Background</h4>Chronic otitis media (COM) is characterized by middle ear fluid predominantly containing cytokines, <i>Nontypeable haemophilus influenzae</i> (NTHi), the mucin MUC5B, and neutrophil extracellular traps (NETs). NETs consist of extracellular DNA coated with antibacterial proteins such as myeloperoxidase (MPO) and citrullinated histone 3 (CitH3). NETs can damage tissues and sustain inflammation. Our study aimed to develop an in vitro model of NETosis, testing COM inductors.<h4>Me ...[more]

PMID: 32596498

Similar Datasets

Project description:Neutrophils undergo a unique form of cell death to generate neutrophil extracellular traps (NETs). It is well established that citrullination of histones (e.g., CitH3) facilitates chromatin decondensation during NET formation (NETosis), particularly during calcium-induced NETosis that is independent of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activation. However, the importance of other forms of histone modifications in NETosis has not been established. We considered that acetylation of histones would also facilitate NETosis. To test this hypothesis, we induced NOX-dependent NETosis in human neutrophils with phorbol myristate acetate or lipopolysaccharide (from Escherichia coli 0128), and NOX-independent NETosis with calcium ionophores A23187 or ionomycin (from Streptomycesconglobatus) in the presence or absence of two pan histone deacetylase inhibitors (HDACis), belinostat and panobinostat (within their half maximal inhibitory concentration (IC50) range). The presence of these inhibitors increased histone acetylation (e.g., AcH4) in neutrophils. Histone acetylation was sufficient to cause a significant increase (~20%) in NETosis in resting neutrophils above baseline values. When acetylation was promoted during NOX-dependent or -independent NETosis, the degree of NETosis additively increased (~15⁻30%). Reactive oxygen species (ROS) production is essential for baseline NETosis (mediated either by NOX or mitochondria); however, HDACis did not promote ROS production. The chromatin decondensation step requires promoter melting and transcriptional firing in both types of NETosis; consistent with this point, suppression of transcription prevented the NETosis induced by the acetylation of histones. Collectively, this study establishes that histone acetylation (e.g., AcH4) promotes NETosis at baseline, and when induced by both NOX-dependent or -independent pathway agonists, in human neutrophils. Therefore, we propose that acetylation of histone is a key component of NETosis.

Dataset Information

MUC5B induces in vitro neutrophil extracellular trap formation: Implication in otitis media.

Background

Methods

Results

Level of evidence

Publications

MUC5B induces in vitro neutrophil extracellular trap formation: Implication in otitis media.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets