March 22, 2023

have also been epidemiologically and serologically linked to HP and a more complex immune response that includes TH2 cell activation in more severe and chronic forms of HP [51, 52]

have also been epidemiologically and serologically linked to HP and a more complex immune response that includes TH2 cell activation in more severe and chronic forms of HP [51, 52]. to additional inflammatory agents such as lipopolysaccharide (LPS), with uncertain relevance to disease manifestation. To determine how high dose inhalation of LPS influences the manifestation of allergic airway disease induced from the allergenic mold (with and without 1?g of LPS over two weeks. Changes in airway hyperreactivity, airway and lung inflammatory cell recruitment, antigen-specific immunoglobulins, and histopathology were determined. Results In comparison to mice challenged only with abrogated airway hyperresponsiveness and strongly attenuated airway eosinophilia, PAS+?goblet cells and TH2 reactions while enhancing TH1 and TH17 cell recruitment to lung. Addition of LPS AFN-1252 resulted in more severe, diffuse lung swelling with spread, loosely-formed parenchymal granulomas, but failed to alter fungus-induced IgE and IgG antibodies. Conclusions In AFN-1252 contrast to the strongly allergic lung phenotype induced by fungal spores only, addition of a relatively high dose of LPS abrogates asthma-like features, replacing them with a phenotype more consistent with acute hypersensitivity pneumonitis (HP). These findings extend the already established link between airway mycosis and asthma to HP and describe a powerful model for further dissecting the pathophysiology of HP. Supplementary Info The online version contains supplementary material available at 10.1186/s12931-021-01850-5. (4??105 spores) in the presence or absence of 1?g LPS (O127:B8, Sigma) 3 times per week AFN-1252 for a total of 8 difficulties, allowed to rest, and euthanized the following day while described in the Fig.?1A. Open in a separate windowpane Fig. 1 LPS attenuates fungal-induced asthma to promote neutrophilia (A) Mice were challenged intranasally with 4??105 (AN) conidia and/or 1?g LPS for 3?weeks while diagramed. The effects on airway hyperreactivity (B) and inflammatory cells in bronchoalveolar lavage fluid (BALF) (C, D) were determined. Results are offered as the mean??SEM (n??3 in each group). *p? ?0.05 compared with PBS administration; #p? ?0.05 between the indicated organizations. Data are from one of two representative and self-employed experiments Airway physiology Bronchial responsiveness was determined by assessing the increase in respiratory system resistance (RRS) to 5 increasing intravenous acetylcholine doses, measured by whole body, semi-invasive plethysmography as previously explained [14]. Sample collection and solitary cell suspensions Mice were intubated having a 20G ? catheter (Smith Medical) and lavaged with 1.6?mL PBS. Cells were isolated from bronchoalveolar lavage fluid (BALF) by centrifugation and supernatant stored Rabbit Polyclonal to MMP17 (Cleaved-Gln129) at ??80? for cytokine analysis. Lung homogenates were prepared from PBS perfused lungs by incubating mechanically processed lung cells in digestion buffer with 2?mg/mL collagenase type II (Worthington), AFN-1252 0.04?mg/mL DNase I (Sigma-Aldrich), 20% FBS (Gibco) in 1X HBSS (Gibco) for 30?min at 37? and approved through a 70um mesh basket (Fisher Scientific). Lung homogenates were further processed for circulation cytometric analysis or cultured in total RPMI 1640 medium (GenClone) at 37? with 5% CO2 immediately. Lung cell tradition supernatants were collected the next day and stored at ??80 for cytokine analysis by multiplex. Cytokine and chemokine analysis The indicated cytokines and chemokines were measured in BALF or lung supernatant by using mouse cytokine/chemokine magnetic bead and mouse Th17 magnetic bead panel kits (Millipore) on a Luminex-based multianalyte platform (Bio-Plex; Bio-Rad, Hercules, CA) following a manufactures recommended methods. Flow cytometric analysis For circulation cytometric analysis of inflammatory cells, single-cell suspensions of BALF cells or lung single-cell suspensions were preincubated having a obstructing cocktail of anti-mouse CD16/32 (clone 93, Biolegend) for 10?min at 4?. Differential staining of inflammatory cells.