RESUMEN
The presence of fungi in tracheal wash (TW) of horses was recently linked to mild-moderate equine asthma, indicating a possible causal role; however, increased numbers of fungi may also stem from asthma-related alteration of tracheal mucus clearance or from environmental exposure. Our objective was to elucidate the association between the presence of fungi in TW and asthma status while controlling for relevant confounders. We conducted a retrospective case-control study involving 73 horses, including 34 controls and 39 asthmatic cases. Each asthmatic horse was matched with a control from the same barn to account for the influence of environmental exposure. All horses underwent respiratory clinical scoring, endoscopy, TW, and bronchoalveolar lavage (BAL). The association between asthma status and presence of TW fungi was tested with multivariable logistic regression modelling, accounting for selected management factors, tracheal mucus accumulation, and selected TW and BAL cytological characteristics, including multinucleated giant cells (MGCs) in the TW. Given the variability in MGC definitions in the literature, particularly concerning their morphology and number of nuclei, we constructed two distinct models for each outcome (asthma status or presence of fungi in TW): one considering MGCs as cells with ≥ 3 nuclei, and another using a criterion of ≥ 10 nuclei. Horses with a tracheal mucus score ≥ 2 exhibited 3.6 to 4.3 higher odds of being asthmatic, depending on the MGC definition. None of the other variables examined were associated with either asthma status or TW fungi detection. Notably, the presence of fungal elements in the TW was not associated with equine asthma.
Asunto(s)
Asma , Hongos , Enfermedades de los Caballos , Tráquea , Animales , Caballos/microbiología , Asma/microbiología , Tráquea/microbiología , Estudios de Casos y Controles , Enfermedades de los Caballos/microbiología , Hongos/aislamiento & purificación , Estudios Retrospectivos , Masculino , Femenino , Líquido del Lavado Bronquioalveolar/microbiologíaRESUMEN
Severe equine asthma (SEA) is a complex respiratory condition characterized by chronic airway inflammation. It shares many clinical and pathological features with human neutrophilic asthma, making it a valuable model for studying this condition. However, the immune mechanisms driving SEA have remained elusive. Although SEA has been primarily associated with a Th2 response, there have also been reports of Th1, Th17, or mixed-mediated responses. To uncover the elusive immune mechanisms driving SEA, we performed single-cell mRNA sequencing (scRNA-seq) on cryopreserved bronchoalveolar cells from 11 Warmblood horses, 5 controls and 6 with SEA. We identified six major cell types, including B cells, T cells, monocytes-macrophages, dendritic cells, neutrophils, and mast cells. All cell types exhibited significant heterogeneity, with previously identified and novel cell subtypes. Notably, we observed monocyte-lymphocyte complexes and detected a robust Th17 signature in SEA, with CXCL13 upregulation in intermediate monocytes. Asthmatic horses exhibited expansion of the B-cell population, Th17 polarization of the T-cell populations, and dysregulation of genes associated with T-cell function. Neutrophils demonstrated enhanced migratory capacity and heightened aptitude for neutrophil extracellular trap formation. These findings provide compelling evidence for a predominant Th17 immune response in neutrophilic SEA, driven by dysregulation of monocyte and T-cell genes. The dysregulated genes identified through scRNA-seq have potential as biomarkers and therapeutic targets for SEA and provide insights into human neutrophilic asthma.
Asunto(s)
Asma , Enfermedad Pulmonar Obstructiva Crónica , Animales , Caballos , Humanos , Neutrófilos , Células Th17 , Inflamación , Líquido del Lavado BronquioalveolarRESUMEN
The transcriptomic profile of a cell population can now be studied at the cellular level using single-cell mRNA sequencing (scRNA-seq). This novel technique provides the unprecedented opportunity to explore the cellular composition of the bronchoalveolar lavage fluid (BALF) of the horse, a species for which cell type markers are poorly described. Here, scRNA-seq technology was applied to cryopreserved equine BALF cells. Analysis of 4,631 cells isolated from three asthmatic horses in remission identified 16 cell clusters belonging to six major cell types: monocytes/macrophages, T cells, B/plasma cells, dendritic cells, neutrophils and mast cells. Higher resolution analysis of the constituents of the major immune cell populations allowed deep annotation of monocytes/macrophages, T cells and B/plasma cells. A significantly higher lymphocyte/macrophage ratio was detected with scRNA-seq compared to conventional cytological differential cell count. For the first time in horses, we detected a transcriptomic signature consistent with monocyte-lymphocyte complexes. Our findings indicate that scRNA-seq technology is applicable to cryopreserved equine BALF cells, allowing the identification of its major (cytologically differentiated) populations as well as previously unexplored T cell and macrophage subpopulations. Single-cell gene expression analysis has the potential to facilitate understanding of the immunological mechanisms at play in respiratory disorders of the horse, such as equine asthma.
Asunto(s)
Asma , Enfermedades de los Caballos , Animales , Líquido del Lavado Bronquioalveolar , Caballos , Análisis de la Célula Individual , TranscriptomaRESUMEN
OBJECTIVES: To determine the impact of age on survival in horses with colitis and to elucidate whether a lower type-1/type-2 cytokine ratio or an exaggerated inflammatory state contribute to reduced survival in aged horses. DESIGN: Part 1: Retrospective cohort analysis. Part 2: Analytic observational study. ANIMALS: Part 1: One hundred twenty-four adult horses with colitis. Part 2: Twenty-nine adult horses with new diarrhea onset while hospitalized. MEASUREMENTS AND MAIN RESULTS: Part 1: Patient signalment, select clinicopathological data, diagnoses, treatment, hospitalization length, and invoice were compared between survivors (n = 101) and nonsurvivors (n = 23). Only age and plasma transfusion retained statistical significance in the final multivariate outcome model, with 8.5 times lower odds of survival in transfused horses (95% confidence interval [CI], 2.6-27.2%). Additionally, the likelihood of nonsurvival increased by 11.8% (95% CI, 4-20.2%) for every year the horse aged (P = 0.002). Similarly, geriatric horses (≥20 years) were 15.2 times more likely to die than young-adults (2-12 years, P = 0.03), independent of financial investment, documented comorbidities, and duration of hospitalization. Part 2: Select cytokine analyses were performed on serum collected from hospitalized horses within 1 hour of diarrhea onset (T0) and 6 hours later. At T0, all recorded clinicopathological variables were comparable between geriatric and young-adult horses, suggesting a similar degree of systemic illness. The median concentration of type-2 cytokines interleukin-4 and interleukin-10, and type-1 cytokine interferon-γ did not differ between age groups. Inflammatory cytokines interleukin-6 and tumor necrosis factor-α were significantly higher in geriatric compared to young-adult horses at both sampling time points. CONCLUSIONS: Outcome of colitis was less favorable in aging horses and patients receiving a plasma transfusion. Although an exaggerated inflammatory state, based on increased interleukin-6 and tumor necrosis factor-α concentrations, in geriatric horses may contribute to reduced survival, a lower type-1/type-2 cytokines ratio was not identified in our geriatric population.