ABSTRACT
BACKGROUND: Epidemiologic studies report an association between pneumonia and urban particulate matter (PM) less than 10 microns (µm) in aerodynamic diameter (PM(10)). Streptococcus pneumoniae is a common cause of bacterial pneumonia worldwide. To date, the mechanism whereby urban PM enhances vulnerability to S pneumoniae infection is unclear. Adhesion of S pneumoniae to host cells is a prerequisite for infection. Host-expressed proteins, including the receptor for platelet-activating factor (PAFR), are co-opted by S pneumoniae to adhere to lower airway epithelial cells. OBJECTIVES: To define whether inhalable urban PM enhances the adhesion of S pneumoniae to airway epithelial cells. METHODS: A549 cells were cultured with PM(10) from Leicester (United Kingdom [UK]) and PM(10) and PM less than 2.5 µm in aerodynamic diameter (PM(2.5)) from Accra (Ghana), then infected with S pneumoniae strain D39. Pneumococcal adhesion to human primary bronchial epithelial cells was also assessed. Bacterial adhesion was determined by quantitative culture and confocal microscopy. The role of oxidative stress was assessed by N-acetyl cysteine, and the role of PAFR was assessed by mRNA transcript level, receptor expression, and receptor blocking. RESULTS: PM(10) (UK) increased S pneumoniae adhesion to both A549 airway epithelial cells and human primary bronchial epithelial cells. PM(10) (Ghana) and PM(2.5) (Ghana) also increased adhesion. Culture of A549 cells by PM(10) (UK) increased PAFR mRNA transcript level and PAFR expression. PM(10) (UK)-stimulated adhesion to A549 cells was attenuated by a PAFR blocker and N-acetyl cysteine. CONCLUSION: Urban PM increases adhesion of S pneumoniae to human airway epithelial cells. PM-stimulated adhesion is mediated by oxidative stress and PAFR.
Subject(s)
Bacterial Adhesion/drug effects , Bacterial Adhesion/physiology , Epithelial Cells/microbiology , Particulate Matter/pharmacology , Platelet Membrane Glycoproteins/metabolism , Receptors, G-Protein-Coupled/metabolism , Respiratory System/cytology , Streptococcus pneumoniae/physiology , Cell Line , Epithelial Cells/cytology , Epithelial Cells/drug effects , Ghana , Humans , Oxidative Stress , Particulate Matter/metabolism , Platelet Activating Factor/genetics , Platelet Activating Factor/metabolism , Platelet Membrane Glycoproteins/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, G-Protein-Coupled/genetics , Respiratory System/drug effects , Respiratory System/microbiology , United KingdomABSTRACT
The study of biological processes has become increasingly reliant on obtaining high-resolution spatial and temporal data through imaging techniques. As researchers demand molecular resolution of cellular events in the context of whole organisms, correlation of non-invasive live-organism imaging with electron microscopy in complex three-dimensional samples becomes critical. The developing blood vessels of vertebrates form a highly complex network which cannot be imaged at high resolution using traditional methods. Here we show that the point of fusion between growing blood vessels of transgenic zebrafish, identified in live confocal microscopy, can subsequently be traced through the structure of the organism using Focused Ion Beam/Scanning Electron Microscopy (FIB/SEM) and Serial Block Face/Scanning Electron Microscopy (SBF/SEM). The resulting data give unprecedented microanatomical detail of the zebrafish and, for the first time, allow visualization of the ultrastructure of a time-limited biological event within the context of a whole organism.
