Presentation Type

Poster

Keywords

Inflammasome, Carbon Black, nanoparticles, macrophage, cell death

Department

Biology

Major

Biology

Abstract

Carbon black (CB) is the primary nanoparticulate component of air pollution from fossil fuel combustion. This work examines the cellular impact of ultrafine carbon (carbon black, CB) nanoparticles, that range in size down to 30 nm, upon murine macrophages. The size analysis of the carbon black nanoparticles was performed using atomic force microscopy (AFM) and transmission electron microscopy (TEM) techniques. RAW246.7 macrophage cells were exposed to CB doses ranging from 50 – 200 ug/ml in complete media. Analysis of cell survival over time revealed elevated rates of significant nuclear degradation and cell lifting after 48 hours of exposure, and in a dose dependent pattern. Live cell imaging of cells exposed to nanoparticles revealed a visible uptake of nanoparticles with accumulation over time. To assess inflammasome signaling, both caspase-1 activation and IL-1b production were observed in whole cell lysates. Caspase-1 activation was measured as the appearance of the active (cleaved) form of the protease appearing in immunoblot analysis. The analysis revealed significant activation of caspase-1 with 48-hour CB exposures at doses of 100-200 ug/ml. Similarly, levels of IL-1b were significantly induced by CB exposure, with maximal induction observed after a 48 hour exposure. Macrophage cells were assessed for accumulation of LC3, a marker for autophagosome vesicle accumulation. Immunoblot analysis revealed a significant accumulation of LC3 in response to CB exposure and in response to chloroquine, which inhibits autophagosome/lysosome fusion. Taken together, these results support a model in which CB exposure activates the inflammasome and disrupts autophagy in macrophages.

Faculty Mentor

Jay Brewster

Funding Source or Research Program

Summer Undergraduate Research in Biology

Location

Waves Cafeteria

Start Date

29-3-2019 2:00 PM

End Date

29-3-2019 3:00 PM

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Mar 29th, 2:00 PM Mar 29th, 3:00 PM

Ultrafine carbon nanoparticles activate inflammasome signaling and cell death in murine macrophages

Waves Cafeteria

Carbon black (CB) is the primary nanoparticulate component of air pollution from fossil fuel combustion. This work examines the cellular impact of ultrafine carbon (carbon black, CB) nanoparticles, that range in size down to 30 nm, upon murine macrophages. The size analysis of the carbon black nanoparticles was performed using atomic force microscopy (AFM) and transmission electron microscopy (TEM) techniques. RAW246.7 macrophage cells were exposed to CB doses ranging from 50 – 200 ug/ml in complete media. Analysis of cell survival over time revealed elevated rates of significant nuclear degradation and cell lifting after 48 hours of exposure, and in a dose dependent pattern. Live cell imaging of cells exposed to nanoparticles revealed a visible uptake of nanoparticles with accumulation over time. To assess inflammasome signaling, both caspase-1 activation and IL-1b production were observed in whole cell lysates. Caspase-1 activation was measured as the appearance of the active (cleaved) form of the protease appearing in immunoblot analysis. The analysis revealed significant activation of caspase-1 with 48-hour CB exposures at doses of 100-200 ug/ml. Similarly, levels of IL-1b were significantly induced by CB exposure, with maximal induction observed after a 48 hour exposure. Macrophage cells were assessed for accumulation of LC3, a marker for autophagosome vesicle accumulation. Immunoblot analysis revealed a significant accumulation of LC3 in response to CB exposure and in response to chloroquine, which inhibits autophagosome/lysosome fusion. Taken together, these results support a model in which CB exposure activates the inflammasome and disrupts autophagy in macrophages.