Institute of Immunology, Faculty of Veterinary Medicine, Leipzig University, Leipzig, Germany.
Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY, United States.
Front Immunol. 2022 Jul 6;13:896255. doi: 10.3389/fimmu.2022.896255. eCollection 2022.
Equine asthma (EA) is a highly relevant disease, estimated to affect up to 20% of all horses, and compares to human asthma. The pathogenesis of EA is most likely immune-mediated, yet incompletely understood. To study the immune response in the affected lower airways, mixed leukocytes were acquired through bronchoalveolar lavage (BAL) and the cell populations were analyzed on a single-cell basis by flow cytometry (FC). Samples of 38 horses grouped as respiratory healthy or affected by mild to moderate (mEA) or severe EA (sEA) according to their history, clinical signs, and BAL cytology were analyzed. Using FC, BAL cells and PBMC were comprehensively characterized by cell surface markers . An increased percentage of DH24A polymorphonuclear cells, and decreased percentages of CD14 macrophages were detected in BAL from horses with sEA compared to healthy horses or horses with mEA, while lymphocyte proportions were similar between all groups. Independently of EA, macrophages in BAL were CD14CD16, which contrasts the majority of CD14CD16 classical monocytes in PBMC. Percentages of CD16-expressing BAL macrophages were reduced in BAL from horses with sEA compared to healthy horses. While PBMC lymphocytes predominantly contain CD4 T cells, B cells and few CD8 T cells, BAL lymphocytes comprised mainly CD8 T cells, fewer CD4 T cells and hardly any B cells. These lymphocyte subsets' distributions were similar between all groups. After PMA/ionomycin stimulation , lymphocyte activation (CD154 and T helper cell cytokine expression) was analyzed in BAL cells of 26 of the horses and group differences were observed (p=0.01-0.11). Compared to healthy horses' BAL, CD154 lymphocytes from horses with mEA, and CD4IL-17A lymphocytes from horses with sEA were increased in frequency. Activated CD4 T helper cells were more frequent in asthmatics' (mEA, sEA) compared to healthy horses' PBMC lymphocytes. In summary, FC analysis of BAL cells identified increased polymorphonuclear cells frequencies in sEA as established, while macrophage percentages were mildly reduced, and lymphocyte populations remained unaffected by EA. Cytokine production differences of BAL lymphocytes from horses with sEA compared to healthy horses' cells point towards a functional difference, namely increased local type 3 responses in sEA.
马气喘病（EA）是一种高度相关的疾病，据估计，多达 20%的马都会受到影响，与人类气喘病类似。EA 的发病机制极有可能是免疫介导的，但尚未完全了解。为了研究受影响的下呼吸道的免疫反应，通过支气管肺泡灌洗（BAL）获得混合白细胞，并通过流式细胞术（FC）在单细胞基础上分析细胞群体。根据病史、临床症状和 BAL 细胞学，将 38 匹马分为呼吸健康或轻度至中度（mEA）或严重 EA（sEA）组。使用 FC，全面描述了 BAL 细胞和 PBMC 的细胞表面标记物特征。与健康马或 mEA 马相比，sEA 马的 BAL 中 DH24A 多形核细胞的百分比增加，CD14 巨噬细胞的百分比降低，而所有组之间的淋巴细胞比例相似。独立于 EA，BAL 中的巨噬细胞是 CD14CD16，与大多数 PBMC 中的 CD14CD16 经典单核细胞相反。与健康马相比，sEA 马的 BAL 中表达 CD16 的 BAL 巨噬细胞的百分比降低。虽然 PBMC 淋巴细胞主要包含 CD4 T 细胞、B 细胞和少数 CD8 T 细胞，但 BAL 淋巴细胞主要包含 CD8 T 细胞、较少的 CD4 T 细胞和几乎没有 B 细胞。所有组之间这些淋巴细胞亚群的分布相似。在用 PMA/离子霉素刺激后，分析了 26 匹马的 BAL 细胞中的淋巴细胞激活（CD154 和 T 辅助细胞细胞因子表达），观察到组间差异（p=0.01-0.11）。与健康马的 BAL 相比，mEA 马的 BAL 中 CD154 淋巴细胞和 sEA 马的 CD4IL-17A 淋巴细胞的频率增加。与健康马的 PBMC 淋巴细胞相比，哮喘马（mEA、sEA）中的活化 CD4 T 辅助细胞更为频繁。总之，BAL 细胞的 FC 分析确定了 sEA 中增加的多形核细胞频率，而巨噬细胞百分比略有降低，EA 对淋巴细胞群体没有影响。与健康马的细胞相比，sEA 马的 BAL 淋巴细胞的细胞因子产生差异表明存在功能差异，即在 sEA 中局部 3 型反应增加。
