Combustion-Derived Hydrocarbons Localize to Lipid Droplets in Respiratory Cells

Abstract

Combustion-generated radicals interact to form polynuclear aromatic hydrocarbons (PAHs), including carcinogens. PAHs aggregate into 20- to 50-nm particles, which extend into branched-chain structures (soots). Incomplete combustion yields black soot particles and black smoke. Many PAHs, including those in soots, fluoresce upon excitation. We have reported that butadiene soot (BDS), generated during combustion of the high-volume petrochemical 1,3-butadiene, serves as a reproducible example of combustion-derived fine and ultrafine particles, with the potential for acute or delayed health effects. Human bronchoepithelial cells (BEAS-2B) display time- and concentration-dependent responses to BDS exposure, culminating in concentration of fluorescent PAHs within discrete cytoplasmic bodies. Here we identify the cytoplasmic compartment(s) in which combustion-derived PAHs concentrate and assess the metabolic responses associated with this compartmentalization. BDS-associated fluorescence colocalized with a red fluorescent cholesterol analog and a transfected plasmid coding for a fluorescent lipid droplet surface protein within BEAS-2B cells. After BDS exposure, murine alveolar macrophages (MH-S) and adipocytes (3T3-L1) also develop fluorescence. These findings, especially within adipocytes, support the accumulation of PAHs within lipid droplets. Microarray data revealed up-regulation of aryl hydrocarbon receptor-induced Phase I biotransformation enzymes and nuclear erythroid-2 related factor 2-mediated oxidative stress responses in BEAS-2B cells. Quantitative RT-PCR results confirmed a time-dependent up-regulation of Phase I biotransformation enzymes (CYP1A1, CYP1B1, and ALDH3A1) in BDS-exposed BEAS-2B and MH-S cells. Thus, respiratory cell lipid droplets concentrate PAHs delivered by combustion-derived ultrafine particles. These PAHs, including several found in BDS and in cigarette smoke, activate xenobiotic metabolism pathways and thereby potentiate their toxicity.

Live cell imaging of the intracellular compartmentalization of the contaminate benzo[a]pyrene

“…To determine the localization of B[a]P in lysosomes of Hepa1c1c7 cells LysoTracker ¾ was used (see Figure 9). Although it was reported earlier that there is [18,19] no significant accumulation of B[a]P in lysosomes of Clone 9 cells and BEAS-2B cells respectively, our results show substantial colocalization of B[a]P fluorescence with the lysosomal fluorescence in Hepa1c1c7 cells.…”

“…Stored contaminants might get released in the cell cytoplasm at a later time point in a controlled manner to prevent any immediate damage to cellular processes due to their toxic nature. Miller and Whitlock showed [18] that two thirds of the original B[a]P recovered adhered supposedly to LDs in lung cells more than 5 months after exposure.…”

“…The delocalized pi bonds of B[a]P and its metabolites make them highly fluorescent. This property has been used previously for experimental purposes in techniques like flow cytometry [16], fluorescence microscopy [17,18], laser cytometry [19][20][21], and multiphoton imaging [22][23][24].…”

Effects of atmospheric pollutants on the Nrf2 survival pathway

“…BDS is rich in particles that can absorb PAHs. Microarray analysis of human bronchoepithelial cells (BEAS-2B) exposed to BDS showed upregulation of Nrf2-associated genes (Murphy et al 2008). The authors suggested that an increase in cellular oxidative stress was the result of exposure to BDS.…”

Metabolism of benzo(a)pyrene by aortic subcellular fractions in the setting of abdominal aortic aneurysms

“…Subsequent to exposure, PAHs cross through cell membranes and being lipophilic, undergo intracellular accumulation by concentrating in lipid droplets [5]. Studies from our research group have shown that PAHs through a & A.…”