Characterization and pro-inflammatory potential of indoor mold particles.

A number of epidemiological studies find an association between indoor air dampness and respiratory health effects. This is often suggested to be linked to enhanced mold growth. However, the role of mold is obviously difficult to disentangle from other dampness-related exposure including microbes as well as non-biological particles and chemical pollutants. The association may partly be due to visible mycelial growth and a characteristic musty smell of mold. Thus, the potential role of mold exposure should be further explored by evaluating information from experimental studies elucidating possible mechanistic links. Such studies show that exposure to spores and hyphal fragments may act as allergens and pro-inflammatory mediators and that they may damage airways by the production of toxins, enzymes, and volatile organic compounds. In the present review, we hypothesize that continuous exposure to mold particles may result in chronic low-grade pro-inflammatory responses contributing to respiratory diseases. We summarize some of the main methods for detection and characterization of fungal aerosols and highlight in vitro research elucidating how molds may induce toxicity and pro-inflammatory reactions in human cell models relevant for airway exposure. Data suggest that the fraction of fungal hyphal fragments in indoor air is much higher than that of airborne spores, and the hyphal fragments often have a higher pro-inflammatory potential. Thus, hyphal fragments of prevalent mold species with strong pro-inflammatory potential may be particularly relevant candidates for respiratory diseases associated with damp/mold-contaminated indoor air. Future studies linking of indoor air dampness with health effects should assess the toxicity and pro-inflammatory potential of indoor air particulate matter and combined this information with a better characterization of biological components including hyphal fragments from both pathogenic and non-pathogenic mold species. Such studies may increase our understanding of the potential role of mold exposure.

Pro-inflammatory responses induced by A. fumigatus and A. versicolor in various human macrophage models.

Exposure to mold-contaminated indoor air has been associated with various respiratory diseases, and there is a need for experimental data to confirm these associations. The pro-inflammatory properties of well-characterized aerosolized spores and hyphal fragments from Aspergillus fumigatus and Aspergillus versicolor were examined and compared using various human macrophage cell models including phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophages (THP-1 Ma), primary peripheral blood monocyte-derived macrophages (MDM), and primary airway macrophages (AM) from induced sputum. X-ray treated samples of the two mold species induced different responses with A. fumigatus displaying the most potent induction of pro-inflammatory responses. While hyphal fragments from A. fumigatus were more potent than spores, similar responses were produced by the two growth stages of A. versicolor. THP-1 Ma was the most sensitive model releasing a broad range of cytokines/chemokines. MDM exhibited a similar cytokine/chemokine profile as THP-1 Ma, except for a low-quantity release of interleukin-1ß (IL-1ß). In contrast, AM appeared to be nonresponsive and yielded a different pattern of pro-inflammatory markers. Toll-like receptor (TLR)4, but also to a certain degree TLR2, was involved in several responses induced by spores and aerosolized hyphal fragments of A. fumigatus in MDM. Taken together, MDM seems to be the most promising experimental macrophage model. Abbreviations: AF: A. fumigatus, Aspergillus fumigatus; AV: A. versicolor, Aspergillus versicolor; AM: Airway Macrophage; CBA: Cytometric Bead Array; CD: Cluster of Differentiation; DTT: dithiothreitol; ELISA: Enzyme Linked Immunosorbent Assay; FBS: fetal bovine serum; GM-CSF: Granulocyte macrophage colony-stimulating factor; IL-1ß: Interleukin-1beta; MDM: Monocyte-Derived Macrophages; NF-κB: Nuclear Factor kappa light chain enhancer of activated B cells; NLR: NOD-like Receptor; PAMP: Pathogen Associated Molecular Pattern; PMA: Phorbol 12-myristate 13-acetate; PRR: Pattern Recognition Receptor; THP-1: Human leukemia monocyte cell line; TLR: Toll-like Receptor; TNF-α: Tumor Necrosis Factor- alpha.

Pro-Inflammatory Responses in Human Bronchial Epithelial Cells Induced by Spores and Hyphal Fragments of Common Damp Indoor Molds.

Damp indoor environments contaminated with different mold species may contribute to the development and exacerbation of respiratory illnesses. Human bronchial epithelial BEAS-2B cells were exposed to X-ray treated spores and hyphal fragments from pure cultures of Aspergillus fumigatus, Penicillum chrysogenum, Aspergillus versicolor and Stachybotrys chartarum. Hyphal fragments of A. fumigatus and P. chrysogenum induced expression and release of the pro-inflammatory cytokine interleukin (IL)-6 and the chemokine IL-8, while none of the other hyphal preparations had effects. Hyphal fragments from A. fumigatus and P. chrysogenum also increased the expression of IL-1α, IL-1ß and tumor necrosis factor (TNF)-α, but these cytokines were not released. X-ray treated spores had little or no inflammatory potential. Attenuating Toll-like receptor (TLR)-2 by blocking antibodies strongly reduced the A. fumigatus and P. chrysogenum hyphae-induced IL-6 and IL-8 release, whereas TLR4 antagonist treatment was without effects. Untreated A. fumigatus spores formed hyphae and triggered expression of pro-inflammatory genes with similarities to the effects of hyphal fragments. In conclusion, while X-ray treated spores induced no pro-inflammatory responses, hyphal fragments of A. fumigatus and P. chrysogenum enhanced a TLR2-dependent expression and release of IL-6 and IL-8.

DNA damage and DNA damage response in human bronchial epithelial BEAS-2B cells following exposure to 2-nitrobenzanthrone and 3-nitrobenzanthrone: role in apoptosis.

Nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) are mutagenic and carcinogenic environmental pollutants found in diesel exhaust and on urban air pollution particles. In the present study, human bronchial epithelial BEAS-2B cells were exposed to 2-nitrobenzanthrone (2-NBA) and 3-nitrobenzanthrone (3-NBA). DNA damage responses were compared to those observed after exposure to 1-nitropyrene (1-NP) and benzo[a]pyrene (B[a]P). Examination by microscopy revealed that 3-NBA was the most potent toxic compound while weaker responses were observed with 1-NP and B[a]P. Most interestingly, 2-NBA did not induce cell death or any other stress-related responses. 3-NBA induced a typical apoptotic cell death judged by nuclear condensation and little plasma membrane damage as well as cleavage of caspase 3 and poly-(ADP-ribose) polymerase (PARP). Exposure to 3-NBA resulted in an accumulation of cells in S-phase, and further analysis by Western blotting, immunocytochemistry and flow cytometry revealed that 3-NBA induced a DNA damage response characterized by phosphorylation of ATM (ataxia-telangiectasia mutated), checkpoint kinase (Chk) 2/Chk1, H2AX and p53. The p53 inhibitor pifithrin-α inhibited 3-NBA-induced apoptosis while small effects were seen using pifithrin-µ, suggesting that 3-NBA-induced cell death is a result of transcriptional activation of p53. In conclusion, 3-NBA is a potent inducer of apoptosis, which seemed to be triggered by the DNA damage response. Furthermore, a change of the nitro-group to the second position (i.e. 2-NBA) dramatically changed the cellular reactivity of the compound.