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Immunohistochemical Expression of Neurokinin-A and Interleukin-8 in the Bronchial Epithelium of Horses with Severe Equine Asthma Syndrome during Asymptomatic, Exacerbation, and Remission Phase.

ABSTRACT: Severe equine asthma (EA) syndrome is a chronic obstructive disease characterized by exaggerated contraction, inflammation, and structural alteration of the airways in adult horses, when exposed to airborne molds and particulate material. However, little is known about the relationship between the degree and type of inflammation on one hand, and the severity of the disease and the response to treatment on the other. Furthermore, to date, very few studies evaluate the diagnostic value of histology and immunohistochemical features of endoscopic biopsies on subjects with severe equine asthma. To investigate the expression of two inflammatory markers (NKA and IL-8) before, during, and after the exacerbation of severe EA, a histological and immunohistochemical study was carried out on a series of biopsy samples collected by bronchoscopy from six EA-affected horses subjected to process exacerbation through environmental stimuli and then to pharmacological treatment. The application of a histological biopsy scoring system revealed a significant difference between control cases and the EA-affected horses in all experimental phases (asymptomatic, early exacerbation phase, late exacerbation phase, and remission phase). For immunohistochemistry (IHC), only the intensity of NKA positivity increases significantly between control horses and the EA horses at late exacerbation and remission phases. In EA-affected horses, a difference was detected by comparing histology between asymptomatic and remission phase, meanwhile, NKA and IL-8 showed no differences between the experimental phases. Based on these results we can assert that: (1) The endoscopic biopsies generate reliable and homogeneous samples in the entire bronchial tree; (2) the clinical improvement associated with treatment is characterized by a significant worsening of the histological findings; and (3) the NKA immunopositivity seems to increase significantly rather than decrease, as one would have expected, after pharmacological treatment. Further studies are necessary both to implement the number of samples and to use other markers of inflammation to characterize the potential role of cytokines in the diagnosis and therapeutic approach of severe equine asthma.

SUBMITTER: Morini M

PROVIDER: S-EPMC8151432 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The olfactory epithelium (OE) is the only body site where neurons contact directly the environment and are therefore exposed to a broad variation of substances and insults. It can serve as portal of entry for neurotropic viruses which spread via the olfactory pathway to the central nervous system. For horses, it has been proposed and concluded mainly from rodent studies that different viruses, e.g., Borna disease virus, equine herpesvirus 1 (EHV-1), hendra virus, influenza virus, rabies virus, vesicular stomatitis virus can use this route. However, little is yet known about cytoarchitecture, protein expression and the intranasal location of the equine OE. Revealing differences in cytoarchitecture or protein expression pattern in comparison to rodents, canines, or humans might help to explain varying susceptibility to certain intranasal virus infections. On the other hand, disclosing similarities especially between rodents and other species, e.g., horses would help to underscore transferability of rodent models. Analysis of the complete noses of five adult horses revealed that in the equine OE two epithelial subtypes with distinct marker expression exist, designated as types a and b which resemble those previously described in dogs. Detailed statistical analysis was carried out to confirm the results obtained on the descriptive level. The equine OE was predominantly located in caudodorsal areas of the nasal turbinates with a significant decline in rostroventral direction, especially for type a. Immunohistochemically, olfactory marker protein and doublecortin (DCX) expression was found in more cells of OE type a, whereas expression of proliferating cell nuclear antigen and tropomyosin receptor kinase A was present in more cells of type b. Accordingly, type a resembles the mature epithelium, in contrast to the more juvenile type b. Protein expression profile was comparable to canine and rodent OE but equine types a and b were located differently within the nose and revealed differences in its cytoarchitecture when compared to canine OE. Equine OE type a closely resembles rat OE. Whether the observed differences contribute to species-specific susceptibility to intranasal insults such as virus infections has to be further investigated.

Project description:BackgroundSevere equine asthma is a naturally occurring lung inflammatory disease of mature animals characterized by neutrophilic inflammation, bronchoconstriction, mucus hypersecretion and airway remodeling. Exacerbations are triggered by inhalation of dust and microbial components. Affected animals eventually are unable of aerobic performance. In this study transcriptomic differences between asthmatic and non-asthmatic animals in the response of the bronchial epithelium to an inhaled challenge were determined.ResultsPaired endobronchial biopsies were obtained pre- and post-challenge from asthmatic and non-asthmatic animals. The transcriptome, determined by RNA-seq and analyzed with edgeR, contained 111 genes differentially expressed (DE) after challenge between horses with and without asthma, and 81 of these were upregulated. Genes involved in neutrophil migration and activation were in central location in interaction networks, and related gene ontology terms were significantly overrepresented. Relative abundance of specific gene products as determined by immunohistochemistry was correlated with differential gene expression. Gene sets involved in neutrophil chemotaxis, immune and inflammatory response, secretion, blood coagulation and apoptosis were overrepresented among up-regulated genes, while the rhythmic process gene set was overrepresented among down-regulated genes. MMP1, IL8, TLR4 and MMP9 appeared to be the most important proteins in connecting the STRING protein network of DE genes.ConclusionsSeveral differentially expressed genes and networks in horses with asthma also contribute to human asthma, highlighting similarities between severe human adult and equine asthma. Neutrophil activation by the bronchial epithelium is suggested as the trigger of the inflammatory cascade in equine asthma, followed by epithelial injury and impaired repair and differentiation. Circadian rhythm dysregulation and the sonic Hedgehog pathway were identified as potential novel contributory factors in equine asthma.

Project description:The Salivary Scavenger and Agglutinin (SALSA) protein is an innate immune protein with various alleged functions, including the regulation of inflammation and tissue remodeling. Transcriptomic studies of severe equine asthma (SEA) showed downregulation of the gene encoding SALSA in bronchial epithelium of asthmatic compared to non-asthmatic horses. This study aimed to characterize expression of SALSA in equine tissues by immunohistochemistry (IHC), corroborate potential differences in epithelial gene expression between asthmatic and non-asthmatic horses, and assess the structure of equine SALSA. An antibody against SALSA was validated through immunoprecipitation followed by mass spectrometry and Western blotting to recognize the equine protein. This antibody was applied to tissue microarrays (TMAs) containing 22 tissues each from four horses. A quantitative PCR assay was designed to compare gene expression for SALSA between six asthmatic and six non-asthmatic horses, before and after an asthmatic challenge, using cDNA from endoscopic bronchial biopsies as source material. The SALSA gene from bronchial cDNA samples of 10 horses, was amplified and sequenced, and translated to characterize the protein structure. Immunostaining for SALSA was detected in the mucosal surfaces of the trachea, bronchi, bronchioles, stomach, small intestine and bladder, in pancreatic and salivary gland ducts, and in uterine gland epithelium. Staining was strongest in the duodenum, and the intercalated ducts and Demilune cells of the salivary gland. SALSA was concentrated in the apical regions of the epithelial cell cytoplasm, suggestive of a secreted protein. Gene expression was significantly lower (p = 0.031) in asthmatic compared to non-asthmatic horses. Equine SALSA consisted of three to five scavenger receptor cysteine-rich (SRCR) domains, two CUB (C1r/C1s, uegf, bmp-1) domains and one Zona Pellucida domain. These domains mediate the binding of ligands involved in innate immunity. Varying numbers of SRCR domains were identified in different horses, indicating different isoforms. In summary, equine SALSA has a predilection for mucosal sites, has multiple isoforms, and has decreased expression in asthmatic horses, suggesting alterations in innate immunity in equine asthma.

Dataset Information

Immunohistochemical Expression of Neurokinin-A and Interleukin-8 in the Bronchial Epithelium of Horses with Severe Equine Asthma Syndrome during Asymptomatic, Exacerbation, and Remission Phase.

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