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ANIMAL MODELS OF MOLD ALLERGY
Citation:
Ward, MDW. ANIMAL MODELS OF MOLD ALLERGY. Presented at Mold Related Health Effects: Cliinical, Remediation Worker Protection and Biomedical Research Issues, Washington, DC, June 28-29, 2004.
Description:
The concept of molds as causative agents for allergy/asthma is not new. In fact many fungal genera have been associated with allergic lung disease, but only a few fungi are well studied and even fewer fungal allergens well characterized. The complexity and variety of fungal propagules in environmental samples, as well as the variety of assessment methods (recently reviewed Dillion et al., 1999; Burge, 2002; Levetin and Horner, 2002) has made identification and characterization of fungal allergens challenging. Furthermore, exposure assessment has focused largely on fungal spores, while exposure to mycelium or component fragments is not only possible, but may be sufficient to induce allergic responsiveness.
A number of epidemiological studies have shown an association between positive skin tests to molds, basidiospores (Lehrer et al., 1994; Chavasco et al., 1997), Alternaria and Penicillium (Eggleston et al., 1998) and asthma. Jacob et al. (2002) found that Cladosporium and Apergillus spore counts were associated with increased risk of allergic sensitization. Cooley et al. (1998) found a strong association between the presence of Penicillium species (especially P. chrysogenum) and Stachybotrys species and sick building syndrome. However, such species-specific associations generally are not made or are complicated by positive results to multiple molds and/or other agents. Therefore, the role that specific molds may play in the induction and/or exacerbation of allergic lung disease can be elucidated in animal studies.
Animal studies related to the assessment of allergic airway disease are somewhat limited. These studies address mold induced nonspecific inflammation and adjuvant activity as well as IgE mediated allergy.
Rylander's (1999) review of field/epidemiology studies relating (1?3)?-D glucan as a marker of biomass to increased extent of symptoms and inflammation suggested that (1?3)?-D glucan may be a useful measure of fungal contamination/exposure in assessing the risk of adverse health effects. Additionally, several animal studies indicate that (1?3)?-D glucan induces nonspecific inflammatory responses. Vassallo et al. (2000) determined that exposures to a (1?3)?-D glucan rich Pneumocystis carinii cell wall fraction resulted in increased pro-inflammatory cytokines (TNF ??) and neutrophilic infiltrate following intratracheal instillation of mice. Additionally, in vitro exposure of primary mouse alveolar macrophages resulted in increased levels of TNF ? and macrophage-inflammatory protein-2 (MIP-2). Guinea pigs, exposed for five weeks to daily inhalations of (1?3)?-D glucan, had increased eosinophils in lung lavage, lung interstitium, and airway epithelium, as well as an interstitial increase in lymphocytes (Fogelmark et al. 2001). Endotoxin did not cause these effects and in fact modulated the effects of (1?3)?-D glucan.
Other studies have shown that (1?3)?-D glucan can enhance allergic responses to known antigens. These studies have demonstrated that mice exposed to aerosolized (1?3)?-D glucan, in an ovalbumin (OVA) allergy model, had higher levels of OVA specific IgE and IgG1 but not IgG2a then mice exposed to OVA alone (Ormstad et al., 2000; Wan et al., 1999). Furthermore, Wan et al. (1999) found that (1?3)?-D glucan increased eosinophils in lung lavage, while, increasing IL-10 mRNA and decreasing IL-12 mRNA in mouse lung cells. These studies suggest that (1?3)?-D glucan may act as an adjuvant during the initiation (induction) phase of allergic responses.
(1?3)?-D glucan may not be the only source of fungal-induced respiratory irritation. Acute exposures to S. chartarum (Korpi et al., 2002) and Aspergillus versicolor (Korpi et al., 2003), fungi frequently found in water damaged buildings, provoked a dose-dependent upper respiratory tact irritation in mouse airways. Korpi et al. (2003) found that pure -glucan exposure resulted in lower sensory irritation responses than did A. versicolor extract. They concluded that -glucan was unlikely to be the major source of the irritation but could not rule out ergosterol since its biological effects are unclear. In their S. chartarum study, they found that the extract caused sensory irritation in both immunized and non-immunized mice. Additionally, repeated exposures resulted in significantly increased IgE.
An apparently unique hypersensitivity disorder resulting from the frank colonization of the lungs by Aspergillus fumigatus is allergic bronchopulmonary aspergillosis (ABPA). ABPA is most common in asthmatic and cystic fibrosis patients. Diagnostic criteria includes A. fumigatus-specific responses such as immediate cutaneous reactivity, precipitating IgG antibodies, and elevated IgE, as well as, peripheral eosinophilia coincident with chest radiographic infiltrates and the presence of A. fumigatus in sputum (Leonard et al., 2001). In a murine model of ABPA, Kurup et al. (1999), concluded that A. fumigatus exposure resulted in a predominately Th2 type response in an immunologically unaltered host. However, upon depletion of Th2 cytokines a similar lung inflammation occurs but with a Th1 type response. Kurup et al. (1999) concluded that ABPA pathogenesis results from multiple induction pathways.
A number of animal studies have addressed the cause-effect relationship between molds and allergic asthma including Aspergillus fumigatus (Kurup et al., 2001), Pencillium chrysogenum (Cooley et al., 2000; Schwab et al. 2003; Chung et al., unpublished data), Metarhizium anisopliae (Ward et al., 2000a, 2000b), and Stachybotrys chartarum (Viana et al., 2002; Korpi, 2002).
Kurup et al. (2001) investigated the role of individual A. fumigatus allergens in the development of allergic asthma in a murine model, using four recombinant A. fumigatus allergens (Asp f1, f3, f4, and f6). Mice exposed to the crude extract developed higher levels of serum total IgE, as well as, peripheral blood and lung eosinophils then those exposed to the recombinant allergens. However, airway hyperreactivity was significantly increased upon challenge of sensitized mice with either the crude extract or the recombinant allergens. This study suggests that the respiratory allergic responses observed are the result of the cumulative effects of all of the allergens.
Studies of P. chrysogenum exposed mice demonstrated that six weekly intranasal instillations of viable (averaging 25% viability) P. chrysogenum conidia induced allergic asthma-like responses including elevated total and specific IgE and IgG1 and eosinophilia compared to controls. However, instillation of non-viable conidia resulted in a significant increase in total serum IgG2a suggesting a Th1 mediated immune response (Cooley et al., 2000). Subsequent studies found that mice did not develop allergic responses with intranasal low-level P. chrysogenum conidia (102 viable conidia) exposures over 11 weeks (Schwab et al., 2003). Additionally, Schwab et al. (2003) found that intraperitoneal sensitization, with an aqueous protease extract from viable P. chrysogenum spores, resulted in significant increases in serum IgE and IgG1 in mice intranasally challenged with either the protease extract or viable spores, but not non-viable spores. These studies clearly suggest that viable P. chrysogenum spores can play a role in allergy development, and that the level of exposure is a critical factor in allergic sensitization.
Studies from our laboratory (Chung et al., unpublished data; Ward et al., 2000a, 2000b; Viana et al., 2002) have demonstrated that fungal extracts of P. chrysogenum, M. anisopliae, and S. chartarum induce antigen non-specific lung injury through increased permeability (BALF total protein) and cell damage (BALF LDH activity), as well as neutrophil influx. Additionally, multiple exposures to these extracts induced responses indicative of an antigen-specific immune response including elevated BALF IL-5, serum total and antigen-specific IgE, and immediate airway reactivity, following fungal extract challenge, and hyperresponsiveness to non-specific methacholine challenge. The conclusion from this work is that S. chartarum, P. chrysogenum, and M. anisopliae have demonstrated the ability to induce responses in mouse that are characteristic of allergic lung disease in humans. Additionally, the P. chrysogenum extract was shown to induce these responses in a dose-dependent manner. Furthermore, a higher dose of P. chrysogenum extract was required to obtain a level of responsiveness similar to that demonstrated in M. anisopliae extract exposed mice. These results indicate that P. chrysogenum may be less potent at allergy induction.
Research needs are the following: 1) identify and characterize mold allergens and 2) extrapolation from animal models to humans. Our laboratory is currently in the process of characterizing the IgE binding proteins identified in the mold extracts. This information will aid in addressing the question of "What makes a protein an allergen?" Sera collected in a pilot study of mild to moderate asthmatics in North Carolina will be used to assess exposure (i.e. IgG), atopy (i.e. IgE) and more recent exposures (i.e. IgM) to a panel of mold extracts. IgE binding patterns from human and mouse sera will be compared to determine if these two species responsed to the same fungal proteins for IgE induction.
In conclusion, animal studies demonstrate the potential for mold exposures to induce responses characteristic of allergic lung disease and therefore have the potential to induce allergic asthma in humans. In addition, some mold/fungi components can function as adjuvants in the induction of allergic responses while acute exposure animal studies indicate that molds can induce pro-inflammatory responses. Furthermore, these animal models provide the means to address issues such as the potency of mold allergens relative to other indoor allergens, as well as, to identify and characterize mold allergens. It is clear that there are still a number of questions, both qualitative and quantitative, that need to be answered before the risks associated with indoor exposure to fungi can be adequately assessed.