Formation of a stable mimic of ambient particulate matter containing viable infectious respiratory syncytial virus and its dry-deposition directly onto cell cultures.

Epidemiological associations of worse respiratory outcomes from combined exposure to ambient particulate matter (PM) and respiratory viral infection suggest possible interactions between PM and viruses. To characterize outcomes of such exposures, we developed an in vitro mimic of the in vivo event of exposure to PM contaminated with respiratory syncytial virus (RSV). Concentration of infectious RSV stocks and a particle levitation apparatus were the foundations of the methodology developed to generate specific numbers of PM mimics (PM(Mimics)) of known composition for dry, direct deposition onto airway epithelial cell cultures. Three types of PM(Mimics) were generated for this study: (i) carbon alone (P(C)), (ii) carbon and infectious RSV (P(C+RSV)), and (iii) aerosols consisting of RSV (A(RSV)). P(C+RSV) were stable in solution and harbored infectious RSV for up to 6 months. Unlike A(RSV) infection, P(C+RSV) infection was found to be dynamin dependent and to cause lysosomal rupture. Cells dosed with PM(Mimics) comprised of RSV (A(RSV)), carbon (P(C)), or RSV and carbon (P(C+RSV)) responded differentially as exemplified by the secretion patterns of IL-6 and IL-8. Upon infection, and prior to lung cell death due to viral infection, regression analysis of these two mediators in response to incubation with A(RSV), P(C), or P(C+RSV) yielded higher concentrations upon infection with the latter and at earlier time points than the other PM(Mimics). In conclusion, this experimental platform provides an approach to study the combined effects of PM-viral interactions and airway epithelial exposures in the pathogenesis of respiratory diseases involving inhalation of environmental agents.

Dose-response studies involving controlled deposition of less than 100 particles generated and levitated in an ac trap onto lung cells, in vitro, and quantitation of ICAM-1 differential expression.

A developing area of interest regarding the relationship between the adverse health effects associated with particles suspended in the troposphere is an understanding of how particle chemical composition influences different biological outcomes. Described is the development and application of an apparatus and methodology wherein a known number of particles of tropospherically relevant chemical composition can be designed and levitated in an alternating current (ac) trap followed by their controlled deposition directly from the ac trap onto air-liquid interface cultured lung cells. A downstream biological response, differential upregulation of intercellular adhesion molecule (ICAM)-1, was measurable using fluorescence microscopy in the air-liquid interface human lung cell cultures even though the dose per culture was 0-100 lipopolysaccharide (LPS)-containing elemental carbon particles (52 pg LPS per 6.3 microm diameter particle). Fluorescence emission intensity data measured from a 1 mm2 area centered over the site of particle deposition were fitted using a least squares linear regression line. Because the total mass of each different compound comprising each of the particles delivered to the culture was known, the data generated with this methodology can be expressed as a pro-inflammation potential (in this case ICAM-1 expression) per particle number and composition. Also described is how this methodology affords opportunities to quantitatively study pro-inflammatory intercellular signaling leading to ICAM-1 expression at sites distal to the site of particle deposition.