Daven N. Jackson-Humbles

Enhanced allergic airway disease in old mice is associated with a Th17 response

The prevalence of asthma in the elderly is increasing and associated with higher mortality than in children or young adults. However, the effects of age on the development and character of allergic asthma have been understudied. It has been suggested that mixed Th2/Th17 responses cause more severe forms of asthma, but the role of Th17 response in allergic airway disease and aging is not well understood.

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.

Ozone-Induced Type 2 Immunity in Nasal Airways. Development and Lymphoid Cell Dependence in Mice

Inhalation exposures to ozone commonly encountered in photochemical smog cause airway injury and inflammation. Elevated ambient ozone concentrations have been epidemiologically associated with nasal airway activation of neutrophils and eosinophils. In the present study, we elucidated the temporal onset and lymphoid cell dependency of eosinophilic rhinitis and associated epithelial changes in mice repeatedly exposed to ozone. Lymphoid cell-sufficient C57BL/6 mice were exposed to 0 or 0.5 parts per million (ppm) ozone for 1, 2, 4, or 9 consecutive weekdays (4 h/d). Lymphoid cell-deficient, Rag2(-/-)Il2rg(-/-) mice were similarly exposed for 9 weekdays. Nasal tissues were taken at 2 or 24 hours after exposure for morphometric and gene expression analyses. C57BL/6 mice exposed to ozone for 1 day had acute neutrophilic rhinitis, with airway epithelial necrosis and overexpression of mucosal Ccl2 (MCP-1), Ccl11 (eotaxin), Cxcl1 (KC), Cxcl2 (MIP-2), Hmox1, Il1b, Il5, Il6, Il13, and Tnf mRNA. In contrast, 9-day ozone exposure elicited type 2 immune responses in C57BL/6 mice, with mucosal mRNA overexpression of Arg1, Ccl8 (MCP-2), Ccl11, Chil4 (Ym2), Clca1 (Gob5), Il5, Il10, and Il13; increased density of mucosal eosinophils; and nasal epithelial remodeling (e.g., hyperplasia/hypertrophy, mucous cell metaplasia, hyalinosis, and increased YM1/YM2 proteins). Rag2(-/-)Il2rg(-/-) mice exposed to ozone for 9 days, however, had no nasal pathology or overexpression of transcripts related to type 2 immunity. These results provide a plausible paradigm for the activation of eosinophilic inflammation and type 2 immunity found in the nasal airways of nonatopic individuals subjected to episodic exposures to high ambient ozone.

Ozone-Induced Nasal Type 2 Immunity in Mice Is Dependent on Innate Lymphoid Cells

Epidemiological studies suggest that elevated ambient concentrations of ozone are associated with activation of eosinophils in the nasal airways of atopic and nonatopic children. Mice repeatedly exposed to ozone develop eosinophilic rhinitis and type 2 immune responses. In this study, we determined the role of innate lymphoid cells (ILCs) in the pathogenesis of ozone-induced eosinophilic rhinitis by using lymphoid-sufficient C57BL/6 mice, Rag2(-/-) mice that are devoid of T cells and B cells, and Rag2(-/-)Il2rg(-/-) mice that are depleted of all lymphoid cells including ILCs. The animals were exposed to 0 or 0.8 ppm ozone for 9 consecutive weekdays (4 h/d). Mice were killed 24 hours after exposure, and nasal tissues were selected for histopathology and gene expression analysis. ILC-sufficient C57BL/6 and Rag2(-/-) mice exposed to ozone developed marked eosinophilic rhinitis and epithelial remodeling (e.g., epithelial hyperplasia and mucous cell metaplasia). Chitinase-like proteins and alarmins (IL-33, IL-25, and thymic stromal lymphopoietin) were also increased morphometrically in the nasal epithelium of ozone-exposed C57BL/6 and Rag2(-/-) mice. Ozone exposure elicited increased expression of Il4, Il5, Il13, St2, eotaxin, MCP-2, Gob5, Arg1, Fizz1, and Ym2 mRNA in C57BL/6 and Rag2(-/-) mice. In contrast, ozone-exposed ILC-deficient Rag2(-/-)Il2rg(-/-) mice had no nasal lesions or overexpression of Th2- or ILC2-related transcripts. These results indicate that ozone-induced eosinophilic rhinitis, nasal epithelial remodeling, and type 2 immune activation are dependent on ILCs. To the best of our knowledge, this is the first study to demonstrate that ILCs play an important role in the nasal pathology induced by repeated ozone exposure.

Neonatal epithelial hypoxia inducible factor-1α expression regulates the response of the lung to experimental asthma

Allergic airway disease is characterized by a T helper type 2 cell-mediated airway inflammation and airway hyperresponsiveness. Little is known about the role of hypoxia-mediated signaling in the progression of the disease. To address this knowledge gap, a mouse model was created in which doxycycline exposure induces the functional deletion of hypoxia inducible factor-1α from alveolar type II and Clara cells of the lung. When hypoxia inducible factor-1α deletion was induced during the early postnatal development period of the lung, the mice displayed an enhanced response to the ovalbumin model of allergic airway disease. These hypoxia inducible factor-1α-deficient mice exhibit increased cellular infiltrates, eosinophilia in the lavage fluid and parenchyma, and T helper type 2 cytokines, as compared with ovalbumin-treated control mice. Moreover, these hypoxia inducible factor-1α-deficient mice display increased airway resistance when compared with their control counterparts. Interestingly, if the loss of hypoxia inducible factor-1α was induced in early adulthood, the exacerbated phenotype was not observed. Taken together, these results suggest that epithelial hypoxia inducible factor-1α plays an important role in establishing the innate immunity of the lung and epithelial-specific deficiency in the transcription factor, during early postnatal development, increases the severity of inflammation and functional airway resistance, following ovalbumin challenge. Finally, these results might explain some of the chronic respiratory pathology observed in premature infants, especially those that receive supplemental oxygen. This early hyperoxic exposure, from normal ambient and supplemental oxygen, would presumably inhibit normal hypoxia inducible factor-1α signaling, mimicking the functional deletion described.

Strain Differences in a Murine Model of Air Pollutant–induced Nonatopic Asthma and Rhinitis

Ozone is an irritating gas found in photochemical smog. Epidemiological associations have been made between the onset of asthma and childhood exposures to increasing levels of ambient ozone (i.e., air pollutant–induced nonatopic asthma). Individuals, however, vary in their susceptibility to this outdoor air pollutant, which may be due, in part, to their genetic makeup. The present study was designed to test the hypothesis that there are murine strain-dependent differences in pulmonary and nasal pathologic responses to repeated ozone exposures. C57BL/6NTac and BALB/cNTac mice were exposed to 0 or 0.8 ppm ozone, 4 hr/day, for 9 consecutive weekdays. In both strains of mice, ozone induced eosinophilic inflammation and mucous cell metaplasia in the nasal and pulmonary airways. Lungs of ozone-exposed C57BL/6NTac mice, however, had greater eosinophilic inflammation, mucous cell metaplasia, and expression of genes related to type 2 immunity and airway mucus hypersecretion, as compared to similarly exposed BALB/cNTac mice. Ozone-exposed C57BL/6NTac mice also had greater eosinophilic rhinitis but a similar degree of mucous cell metaplasia in nasal epithelium, as ozone-exposed BALB/cNTac mice. These findings suggest that nonatopic individuals may differ in their inflammatory and epithelial responses to repeated ozone exposures that are due, in part, to genetic factors.

Innate Lymphoid Cells Mediate Pulmonary Eosinophilic Inflammation, Airway Mucous Cell Metaplasia, and Type 2 Immunity in Mice Exposed to Ozone:

Exposure to elevated levels of ambient ozone in photochemical smog is associated with eosinophilic airway inflammation and nonatopic asthma in children. In the present study, we determined the role of innate lymphoid cells (ILCs) in the pathogenesis of ozone-induced nonatopic asthma by using lymphoid cell-sufficient C57BL/6 mice, ILC-sufficient Rag2−/− mice (devoid of T and B cells), and ILC-deficient Rag2−/−Il2rg−/− mice (depleted of all lymphoid cells including ILCs). Mice were exposed to 0 or 0.8 parts per million ozone for 1 day or 9 consecutive weekdays (4 hr/day). A single exposure to ozone caused neutrophilic inflammation, airway epithelial injury, and reparative DNA synthesis in all strains of mice, irrespective of the presence or absence of ILCs. In contrast, 9-day exposures induced eosinophilic inflammation and mucous cell metaplasia only in the lungs of ILC-sufficient mice. Repeated ozone exposures also elicited increased messenger RNA expression of transcripts associated with type 2 immunity and airway mucus production in ILC-sufficient mice. ILC-deficient mice repeatedly exposed to ozone had no pulmonary pathology or increased gene expression related to type 2 immunity. These results suggest a new paradigm for the biologic mechanisms underlying the development of a phenotype of childhood nonatopic asthma that has been linked to ambient ozone exposures.

Dietary Docosahexaenoic Acid Prevents Silica-Induced Development of Pulmonary Ectopic Germinal Centers and Glomerulonephritis in the Lupus-Prone NZBWF1 Mouse

Ectopic lymphoid structures (ELS) consist of B-cell and T-cell aggregates that are initiated de novo in inflamed tissues outside of secondary lymphoid organs. When organized within follicular dendritic cell (FDC) networks, ELS contain functional germinal centers that can yield autoantibody-secreting plasma cells and promote autoimmune disease. Intranasal instillation of lupus-prone mice with crystalline silica (cSiO2), a respirable particle linked to human lupus, triggers ELS formation in the lung, systemic autoantibodies, and early onset of glomerulonephritis. Here we tested the hypothesis that consumption of docosahexaenoic acid (DHA), an ω-3 polyunsaturated fatty acid with anti-inflammatory properties, influences the temporal profile of cSiO2-induced pulmonary ectopic germinal center formation and development of glomerulonephritis. Female NZBWF1 mice (6-wk old) were fed purified isocaloric diets supplemented with 0, 4, or 10 g/kg DHA - calorically equivalent to 0, 2, or 5 g DHA per day consumption by humans, respectively. Beginning at age 8 wk, mice were intranasally instilled with 1 mg cSiO2, or saline vehicle alone, once per wk, for 4 wk. Cohorts were sacrificed 1, 5, 9, or 13 wk post-instillation (PI) of the last cSiO2 dose, and lung and kidney lesions were investigated by histopathology. Tissue fatty acid analyses confirmed uniform dose-dependent DHA incorporation across all cohorts. As early as 1 wk PI, inflammation comprising of B (CD45R+) and T (CD3+) cell accumulation was observed in lungs of cSiO2-treated mice compared to vehicle controls; these responses intensified over time. Marked follicular dendritic cell (FDC; CD21+/CD35+) networking appeared at 9 and 13 wk PI. IgG+ plasma cells suggestive of mature germinal centers were evident at 13 wk. DHA supplementation dramatically suppressed cSiO2-triggered B-cell, T-cell, FDC, and IgG+ plasma cell appearance in the lungs as well as anti-dsDNA IgG in bronchial lavage fluid and plasma over the course of the experiment. cSiO2 induced glomerulonephritis with concomitant B-cell accumulation in the renal cortex at 13 wk PI but this response was abrogated by DHA feeding. Taken together, realistic dietary DHA supplementation prevented initiation and/or progression of ectopic lymphoid neogenesis, germinal center development, systemic autoantibody elevation, and resultant glomerulonephritis in this unique preclinical model of environment-triggered lupus.