Maternal Antiasthma Simplified Herbal Medicine Intervention therapy prevents airway inflammation and modulates pulmonary innate immune responses in young offspring mice.

BACKGROUND: Maternal asthma is a risk factor for asthma in offspring; however, transmission of the risk for allergic asthma without direct offspring sensitization has not been explored. OBJECTIVE: To determine whether offspring from mothers with ovalbumin (OVA)-sensitized asthma would develop airway disease at first-ever exposure to OVA and whether preconception maternal treatment with the Antiasthma Simplified Herbal Medicine Intervention (ASHMI) or dexamethasone (DEX) could modify this risk in offspring. METHODS: Female BALB/c mice (F0) with OVA-induced asthma were generated using established protocols. Mice with asthma were treated with ASHMI, DEX, or water for 6 to 7 weeks. Naive mice served as controls. Subsequently, mice were mated. Twelve-day-old F1 offspring received 3 consecutive intranasal low- or high-dose OVA exposures without sensitization. Forty-eight hours later, airway inflammation, mucus hypersecretion, serum antibodies, and cytokines were evaluated. RESULTS: Offspring from OVA-sensitized mothers, but not naive mothers, showed eosinophilic and neutrophilic airway inflammation, and mucus hyperplasia after OVA exposure and he presence of OVA-specific IgG1 and IgG2a. Offspring of ASHMI- and DEX-treated mothers showed decreased airway inflammation and mucus hypersecretion after low-dose OVA (P < .05-.001 for the 2 comparisons vs offspring of OVA/Sham mothers). Offspring of ASHMI-treated, but not DEX-treated, mothers were protected after the high-dose OVA challenge (P < .05-.01 vs offspring OVA/Sham). Maternal ASHMI therapy was associated with increased IgG2a (P < .01 vs offspring of OVA/Sham mothers) and decreased bronchoalveolar lavage fluid CXCL-1 and eotaxin-1 levels (P < .01 and P < .05, respectively, vs offspring of OVA/Sham mothers). CONCLUSION: Offspring of mothers with OVA-induced asthma developed airway inflammation and mucus to first-ever OVA exposure without prior sensitization. Maternal therapy with ASHMI was superior to DEX in decreasing offspring susceptibility to airway disease and could be a strategy to lower asthma prevalence.

Supplementation with γ-tocopherol attenuates endotoxin-induced airway neutrophil and mucous cell responses in rats.

Neutrophil-mediated tissue injury is a shared pathogenesis of both chronic pulmonary diseases and acute responses to pathogens, allergens, and airborne pollutants. Interventions to minimize toxic effects of neutrophil-derived oxidants and proteases are usually limited to corticosteroids, which can have adverse side effects. We used a rodent model of endotoxin-induced lung injury to test the hypothesis that the dietary supplement γ-tocopherol (γT), a natural form of vitamin E with antioxidant and novel anti-inflammatory properties, will protect from adverse nasal and pulmonary inflammatory responses induced by endotoxin (lipopolysaccharide; LPS). Male Fisher F344 rats were intranasally (i.n.) instilled with LPS for 2 consecutive days. Beginning 2 days before i.n. LPS, the rats were gavaged daily with 30mg/kg γT. Twenty-four hours after the last i.n. LPS, bronchoalveolar lavage fluid (BALF) was collected, and pulmonary and nasal tissues were analyzed for gene expression and morphometric analyses of neutrophils and intraepithelial mucosubstances (IM). LPS caused increased BALF total cells (70% increase), neutrophils (300%), protein (35%), PGE2 (500%), and secreted mucins (75%). Robust increases in neutrophils and IM were detected in conducting airways. Pulmonary expression of MUC5AC, MIP-2, CINC-1, and MCP-1 was elevated three- to eightfold by LPS. Treatment with γT inhibited LPS-induced increases in BALF total cells, neutrophils, protein, PGE2, and secreted mucins, as well as IM and tissue neutrophil influx. Furthermore γT induced the expression of the regulatory cytokines IL-10 and IFN-γ while decreasing MUC5AC, MIP-2, CINC-1, and MCP-1. These data demonstrate novel therapeutic effects of the dietary vitamin E γT promoting anti-inflammatory pathways to protect from neutrophil-mediated lung injury.

sPLA2-IIa amplifies ocular surface inflammation in the experimental dry eye (DE) BALB/c mouse model.

PURPOSE: sPLA2-IIa is a biomarker for many inflammatory diseases in humans and is found at high levels in human tears. However, its role in ocular surface inflammation remains unclear. An experimentally induced BALB/c mouse dry eye (DE) model was used to elucidate the role of sPLA2-IIa in ocular surface inflammation. METHODS: BALB/c mice were subcutaneously injected with scopolamine and placed in a daytime air-drying device for 5 to 10 days. Control mice received no treatment. DE status was evaluated with tear production with a phenol-red thread method. Tear inflammatory cytokines were quantified by multiplex immunoassays. Ocular surface inflammation and sPLA2-IIa expression were examined by immune-staining and quantitative (q)RT(2)-PCR. Conjunctiva (CNJ) of the mice was cultured for prostaglandin E2 production induced by sPLA2-IIa with various amount of sPLA2-IIa inhibitor, S-3319. RESULTS: Treated mice produced fewer tears and heavier corneal (CN) fluorescein staining than the untreated controls (P < 0.001). They also revealed lower goblet cell density (P < 0.001) with greater inflammatory cell infiltration within the conjunctiva, and higher concentration of tear inflammatory cytokines than the controls. Moreover, treated mice showed heavier sPLA2-IIa immune staining than the controls in the CNJ epithelium, but not in the CN epithelium or the lacrimal gland. Treated mice exhibited upregulated sPLA2-IIa and cytokine gene transcription. Furthermore, CNJ cultures treated with sPLA2-IIa inhibitor showed significantly reduced sPLA2-IIa-induced inflammation. CONCLUSIONS: This is the first report regarding sPLA2-IIa in the regulation of ocular surface inflammation. The findings may therefore lead to new therapeutic strategies for ocular surface inflammation, such as DE disease.

The traditional Chinese herbal formula ASHMI inhibits allergic lung inflammation in antigen-sensitized and antigen-challenged aged mice.

BACKGROUND: Although asthma is typically characterized as a childhood disease, it can develop later in life. Older asthmatic patients may be at increased risk for corticosteroid adverse effects. We developed a novel traditional Chinese medicine to treat asthma called antiasthma simplified herbal medicine intervention (ASHMI). Herbal products may offer safer adjunctive treatment for older asthmatic patients. OBJECTIVE: To investigate the effects of ASHMI on characteristics of allergic asthma in an aged mouse model of asthma. METHODS: BALB/c mice (6 weeks old [young] and 6, 12, and 18 months old [aged]) received ASHMI treatment before and during intraperitoneal ovalbumin sensitization and intratracheal challenges. The control groups were untreated, age-matched, ovalbumin-sensitized and ovalbumin-challenged mice (ovalbumin mice) and naive mice. After the final antigen challenge, airway pressure (defined as the time-integrated change in peak airway pressure) after acetylcholine provocation was measured, representing airway hyperresponsiveness, and bronchoalveolar lavage fluid, sera, lung tissues for histologic analysis, messenger RNA, and collagen were collected. RESULTS: Mean time-integrated change in peak airway pressure values in 6-week-old and 6-, 12-, and 18-month-old ASHMI ovalbumin mice were significantly reduced compared with those of age-matched, nontreated ovalbumin mice. Bronchoalveolar lavage fluid eosinophil numbers were significantly lower in all ASHMI ovalbumin mice. Treatment with ASHMI of young and aged ovalbumin mice resulted in significantly decreased lung inflammation, detected via hematoxylin-eosin staining; airway mucous cell metaplasia, determined by means of periodic acid-Schiff staining; and messenger RNA copy numbers of the mucin gene MUC5AC. Levels of ovalbumin specific IgE and the T(H)2 cytokines interleukin 4 (IL-4), IL-5, and IL-13 in lung and splenocyte cultures were reduced. Interferon gamma secretion was increased. Treatment with ASHMI reduced collagen production. CONCLUSION: Treatment with ASHMI reduces several features of asthma in aged antigen-sensitized and antigen-challenged mice.

The effects of targeted deletion of cannabinoid receptors CB1 and CB2 on intranasal sensitization and challenge with adjuvant-free ovalbumin.

The mechanisms by which cannabinoid receptors CB(1) and CB(2) modulate immune function are not fully elucidated. Critical tools for the determination of the role of both receptors in the immune system are CB(1)/CB(2) double null mice (CB(1)/CB(2) null), and previous studies have shown that CB(1)/CB(2) null mice exhibit exaggerated responses to various immunological stimuli. The objective of these studies was to determine the magnitude to which CB(1)/CB(2) null mice responded to the respiratory allergen ovalbumin (OVA) as compared with wild-type C57BL/6 mice. The authors determined that in the absence of adjuvant, both wild-type and CB(1)/CB(2) null mice mounted a marked response to intranasally instilled OVA as assessed by inflammatory cell infiltrate in the bronchoalveolar lavage fluid (BALF), eosinophilia, induction of mucous cell metaplasia, and IgE production. Many of the endpoints measured in response to OVA were similar in wild-type versus CB(1)/CB(2) null mice, with exceptions being modest reductions in OVA-induced IgE and attenuation of BALF neutrophilia in CB(1)/CB(2) null mice as compared with wild-type mice. These results suggest that T-cell responses are not universally exaggerated in CB(1)/CB(2) null mice.

Acute exposure to ozone exacerbates acetaminophen-induced liver injury in mice.

Ozone (O(3)), an oxidant air pollutant in photochemical smog, principally targets epithelial cells lining the respiratory tract. However, changes in gene expression have also been reported in livers of O(3)-exposed mice. The principal aim of the present study was to determine if acute exposure to environmentally relevant concentrations of O(3) could cause exacerbation of drug-induced liver injury in mice. Overdose with acetaminophen (APAP) is the most common cause of drug-induced liver injury in developed countries. In the present study, we examined the hepatic effects of acute O(3) exposure in mice pretreated with a hepatotoxic dose of APAP. C57BL/6 male mice were fasted overnight and then given APAP (300 mg/kg ip) or saline vehicle (0 mg/kg APAP). Two hours later, mice were exposed to 0, 0.25, or 0.5 ppm O(3) for 6 h and then sacrificed 9 or 32 h after APAP administration (1 or 24 h after O(3) exposure, respectively). Animals euthanized at 32 h were given 5-bromo-2-deoxyuridine 2 h before sacrifice to identify hepatocytes undergoing reparative DNA synthesis. Saline-treated mice exposed to either air or O(3) had no liver injury. All APAP-treated mice developed marked centrilobular hepatocellular necrosis that increased in severity with time after APAP exposure. O(3) exposure increased the severity of APAP-induced liver injury as indicated by an increase in necrotic hepatic tissue and plasma alanine aminotransferase activity. O(3) also caused an increase in neutrophil accumulation in livers of APAP-treated animals. APAP induced a 10-fold increase in the number of bromodeoxyuridine-labeled hepatocytes that was markedly attenuated by O(3) exposure. Gene expression analysis 9 h after APAP revealed differential expression of genes involved in inflammation, oxidative stress, and cellular regeneration in mice treated with APAP and O(3) compared to APAP or O(3) alone, providing some indications of the mechanisms behind the APAP and O(3) potentiation. These results suggest that acute exposure to near ambient concentrations of this oxidant air pollutant may exacerbate drug-induced liver injury by delaying hepatic repair.

Pharmacology and immunological actions of a herbal medicine ASHMI on allergic asthma.

Allergic asthma is a chronic and progressive inflammatory disease for which there is no satisfactory treatment. Studies reported tolerability and efficacy of an anti-asthma herbal medicine intervention (ASHMI) for asthma patients, developed from traditional Chinese medicine. To investigate the pharmacological actions of ASHMI on early- and late-phase airway responses (EAR and LAR), Ovalbumin (OVA)-sensitized mice received 6 weeks of ASHMI treatment beginning 24 h following the first intratracheal OVA challenge. EAR were determined 30 min following the fourth challenge and LAR 48 h following the last challenge. ASHMI effects on cytokine secretion, murine tracheal ring contraction and human bronchial smooth muscle cell prostaglandin (PG) production were also determined.ASHMI abolished EAR, which was associated with significantly reduced histamine, leukotriene C4, and OVA-specific IgE levels, as well as LAR, which was associated with significantly reduced bronchoalveolar lavage fluid (BALF) eosinophils, decreased airway remodeling, and lower Th2 cytokine levels in BALF and splenocyte cultures. Furthermore, ASHMI inhibited contraction of murine tracheal rings and increased production of the potent smooth muscle relaxer PGI(2). ASHMI abrogation of allergic airway responses is associated with broad effects on asthma pathological mechanisms.

An adjuvant-free mouse model of tree nut allergy using hazelnut as a model tree nut.

BACKGROUND: Tree nut allergy, a major group of food allergy, is often linked to fatal or near-fatal systemic anaphylaxis. Currently, an adjuvant-free mouse model to study tree nut hypersensitivity is unavailable. Here we tested the hypothesis that transdermal exposure to hazelnut, a model tree nut, without the use of an adjuvant is sufficient to sensitize mice for immediate hypersensitivity reaction to oral hazelnut challenge. METHODS: BALB/c mice were repeatedly exposed to hazelnut protein via the transdermal route and systemic allergic and anaphylactic responses were studied. RESULTS: Transdermal exposure to hazelnut protein elicited robust systemic IgE response in a dose-dependent manner with immunological memory. Oral challenge of transdermally sensitized mice with hazelnut protein resulted in immediate (30 min after the challenge) clinical signs of systemic anaphylaxis as measured by significant clinical scores and drop in rectal temperature. Clinical hypersensitivity reaction was associated with severe pathological changes in the small intestine. Hazelnut-allergic but not control mice exhibited in vivo activation of GATA-3 and hazelnut-driven recall IL-4, IL-5 and IL-13 response by splenocytes, thus elucidating the underlying mechanism of hazelnut allergy development in this model. CONCLUSIONS: These data suggest that (1) transdermal exposure to hazelnut protein is sufficient to activate the key immune pathways necessary for sensitizing mice for clinical immediate hypersensitivity reactions and (2) this mouse model may be useful for further basic and applied studies on tree nut allergy, especially because it does not depend on an adjuvant for eliciting immediate hypersensitivity reactions to nut protein.

Allergic and anaphylactic response to sesame seeds in mice: identification of Ses i 3 and basic subunit of 11s globulins as allergens.

BACKGROUND: Allergy to sesame seeds is an emerging food allergy of a serious nature due to a high risk of systemic anaphylaxis. Although a mouse model to study sesame anaphylaxis is desirable, currently it is not available. Here, using a transdermal exposure model system, we tested the hypothesis that sesame seed elicits IL-4-associated IgE antibody response with consequent clinical sensitization in mice. METHODS: Groups of BALB/c mice were exposed to sesame seed extract or saline or a control food (vanilla bean extract) by transdermal applications. Systemic IgE, IgG1 and IgG2a antibody responses were examined using preoptimized ELISA. Type 2 and type 1 cytokine responses were evaluated by ex vivo antigen-mediated activation of spleen cells. Clinical response to oral sesame challenge was studied. Western blot and N-terminal amino acid sequence analyses were performed to identify the sesame allergens. RESULTS: Transdermal exposure to sesame elicited robust IgE and IgG1 but very little IgG2a antibody responses. IgE response to transdermal exposure in two high-IgE responder mice strains with disparate MHC confirmed the intrinsic allergenicity of sesame seed. Transdermal sensitization was associated with activation of IL-4 but not IFN-gamma. Furthermore, oral exposure to sesame resulted in clinical signs of systemic anaphylaxis. Western blot and sequence analysis identified four allergens including Ses i 3 and the basic subunit of 11s globulins. CONCLUSION: These data argue that transdermal exposure to sesame seed can result in IL-4 activation, IgE response and clinical sensitization for systemic anaphylaxis.

Hazelnut Allergy: evidence that hazelnut can directly elicit specific IgE antibody response via activating type 2 cytokines in mice.

BACKGROUND: Hazelnut is one of the major tree nuts that causes potentially fatal food allergy, with underlying mechanisms that are unclear at present. One suggestion is that hazelnut allergy results from immune crossreactivity of IgE antibodies produced against certain aeroallergens. We tested the hypothesis that hazelnut is intrinsically capable of eliciting an allergic response using a mouse model. METHODS: Groups of mice were injected intraperitoneally with hazelnut/filbert protein extract with or without alum as an adjuvant, and hazelnut-specific antibody (IgE, IgG1) responses were examined using optimized enzyme-linked immunosorbent assay. Hazelnut-specific type 2 and type 1 cytokine responses were evaluated by ex vivo antigen-mediated activation of spleen cells. RESULTS: Hazelnut elicited robust IgE and IgG1 antibody responses. Timecourse and dose-response analyses further provided evidence for memory type 2-dependent antibody responses to hazelnuts. Hazelnut-specific IgE response in two strains of mice with different MHC haplotypes and IgE response to hazelnut without the use of alum adjuvant asserted that hazelnut is intrinsically an allergenic food. The type 2 cytokine analyses revealed that hazelnut sensitization results from activation of IL-4 and IL-5, thus providing a mechanistic basis for hazelnut-specific IgE response. CONCLUSION: Our data argue that hazelnut - a widely consumed food - is intrinsically an allergenic food capable of directly eliciting hazelnut-binding specific IgE antibodies viaactivation of type 2 cytokines in mice.

CCR3 and CXCR3 as drug targets for allergy: principles and potential.

Atopic disorders include a range of conditions such as allergic asthma, -rhinitis, -conjunctivitis, -dermatitis, food and drug allergies and anaphylaxis. Induction of T helper (Th)-2 immune response with consequent IgE dependent eosinophil, basophil and mast cell mediated tissue damage is the characteristic feature of allergies. The mechanism underlying this unique and long appreciated feature of allergy is being elucidated at the molecular level with advances in our knowledge of the chemokine system. Thus, chemokines that target CCR3 in concert with Th2 cytokines appear to play a pathogenic role in allergy. In contrast, chemokines that target CXCR3 in concert with Th1 cytokine appear to play a beneficial role. Accordingly, inhibiting CCR3/Th2 pathway using CCR3 antagonists is viewed as a potentially useful strategy for anti-allergy drug development. In contrast, the idea of using CXCR3 agonists to inhibiting allergic response by promoting CXCR3/Th1 pathway faces serious concerns of their potential pro-inflammatory activities in vivo. In this article we have critically evaluated the literature examining the principle and potential of this anti-allergy drug development strategy including a summary of various compounds that are under investigation.

An ELISA-based method for measurement of food-specific IgE antibody in mouse serum: an alternative to the passive cutaneous anaphylaxis assay.

Passive cutaneous anaphylaxis (PCA) assay has been a gold standard method to measure allergen-specific IgE antibody (ASIgE Ab) levels in allergy mouse models. Many factors including stringent guidelines for laboratory animal use make PCA a difficult choice. Therefore, alternative methods are needed that can be readily applied for measurement of specific IgE antibody levels in mouse serum. Herein we describe a novel ELISA-based method that is more sensitive in comparison to PCA, IgE isotype-specific (because it has little cross-reactivity with IgG1 or IgG2a isotype) and highly reproducible (<10% inter- or intra-assay variation). Furthermore, we demonstrate the utility of this assay to measure specific IgE Ab against a variety of food extracts including chicken egg, peanut, almond, filbert/hazelnut and sweet potato. These findings are of particular interest to those who are seeking (i) to measure food-extract-specific IgE antibody in animal models and (ii) an alternative to the animal-based PCA method to measure mouse IgE antibodies.

Relative immunogenicity of commonly allergenic foods versus rarely allergenic and nonallergenic foods in mice.

Food allergies affect 6 to 8% of children and 2% of adults in the United States. For reasons that are not clear, eight types of food account for a vast majority (approximately 90%) of food-induced hypersensitivity reactions. In this study, C57Bl/6 mice were used to test the hypothesis that commonly allergenic foods are intrinsically more immunogenic than rarely allergenic or nonallergenic foods in allergy-susceptible hosts. Groups of mice (n = 4 to 5) were injected intraperitoneally with the protein extracts (plus alum as an adjuvant) from chicken eggs, peanuts, almonds, filberts-hazelnuts, walnuts, soybeans, and wheat (commonly allergenic foods) and coffee, sweet potatoes, carrots, white potatoes, cherries, lettuce, and spinach (rarely allergenic and nonallergenic foods). Primary and secondary immune responses (as measured by specific IgG1 antibody serum levels) were measured by an enzyme-linked immunosorbent assay. Proteins from peanuts, almonds, filberts, sweet potatoes, cherries, and spinach elicited robust primary and/or secondary immune responses. Proteins from eggs, walnuts, and lettuce elicited poor primary responses but significant secondary responses. In contrast, wheat, soybeans, coffee, carrots, and white potatoes elicited barely detectable to poor primary and secondary immune responses. The order of the immunogenicity levels of these foods in mice is as follows: almonds = filberts > spinach (Rubisco) > peanuts > or = sweet potatoes > cherries > lettuce > walnuts > chicken eggs > carrots > or = white potatoes > wheat = coffee = soybeans. In summary, these data demonstrate for the first time that: (i) foods vary widely with regard to their relative immunogenicity in allergy-susceptible hosts and (ii) intrinsic immunogenicity in mice does not distinguish commonly allergenic foods from rarely allergenic or nonallergenic foods.

Chemokines in health and disease.

Chemokines belong to a large family of structurally related proteins that play a pivotal role in immune system development and deployment. While a large number of chemokines (approximately 50) and their receptors (approximately 20) have been identified from humans or mice, only a few are known in domestic veterinary species. Recent data implicate CXCL8 (old name, IL-8), CXCL10 (old name, IP-10) (both CXC chemokines) and CCL2 (old name, MCP-1) (a CC chemokine) in veterinary infections, inflammatory diseases or reproduction. There is compelling evidence for neutrophil targeting chemokines such as CXCL8, in ovine bacterial mastitis, bovine pneumonic pasturellosis and equine chronic obstructive pulmonary disease (COPD). Monocyte and lymphocyte targeting chemokines appear to play a role in caprine arthritis encephalitis (CCL2) and canine endotoxemia (CXCL10). Interestingly CCL2 is considered a missing link between hormonal and cellular control of luteolysis. On the other hand, canine cardiovascular conditions are associated with overexpression of CCL2 and CXCL8. Furthermore, a number of veterinary viral pathogens encode chemokine/chemokine receptor like molecules or chemokine binding proteins that may help viruses to evade the immune system. Here, we provide an overview of the chemokine system and critically evaluate the current literature implicating chemokines in veterinary pathophysiology. Furthermore, we highlight promising areas for further research and discuss how and why chemokine antagonists are viewed as next generation anti-inflammatory drugs for the 21st century.