Klier John, May Anna, Fuchs Sebastian, Schillinger Ulrike, Plank Christian, Winter Gerhard, Gehlen Heidrun, Coester Conrad
Ludwig Maximilians University, Department of Veterinary Medicine, Equine Clinic, Veterinärstr. 13, 80539 Munich, Germany.
Vet Immunol Immunopathol. 2011 Nov 15;144(1-2):79-87. doi: 10.1016/j.vetimm.2011.07.009. Epub 2011 Jul 20.
Recurrent airway obstruction (RAO) in horses has become a common problem in stabled horses in industrialized countries and deserves new therapeutic strategies. CpG-oligodeoxynucleotides (CpG-ODNs) were developed as effective immunostimulating agents to induce a Th2/Th1 shift. These agents showed a beneficial therapeutic effect in allergic diseases with predominant Th2 immunoresponse. CpG-ODN delivery by gelatin nanoparticles (GNPs) resulted in enhanced cellular uptake in murine and human in vitro studies and was a starting point for the present trial. The aim of this study was to identify an optimal stimulating CpG motif in horses with regard to species specificity on equine bronchoalveolar lavage (BAL) cells, in terms of a possible specific immunomodulation effect (Th2/Th1 shift) by used CpG-ODN. Accordingly, GNPs were evaluated as a delivery system to improve CpG-ODN immunostimulation in equine BAL cells. BAL fluid (BALF) was obtained from seven horses with moderate RAO and from four healthy horses and was subsequently incubated with five different CpG-ODN sequences (from A-, B- and C-class) and one ODN without any CpG motif. Release of three key cytokines (IL-4, IL-10 and IFN-γ) was quantified by ELISA to detect an allergy mediated Th2 immunoresponse (IL-4) as well as a proinflammatory Th1 response (IFN-γ). Due to its specific anti-inflammatory and anti-allergic effects, IL-10 was considered as a beneficial agent in pathophysiology of RAO. Results showed a significant upregulation of IL-10 and IFN-γ on the one hand and a downregulation of IL-4 on the other hand in RAO affected horses. Cell cultures from healthy horses had a significantly stronger response in cytokine release to all the applied stimuli in contrast to RAO derived cells. Comparing all five CpG sequences, A-class 2216 significantly showed the highest immunomodulatory effects on equine BALF cells and, hence, was chosen for follow-up preliminary clinical studies.
复发性气道阻塞（RAO）在工业化国家的圈养马匹中已成为一个常见问题，需要新的治疗策略。CpG-寡脱氧核苷酸（CpG-ODNs）被开发为有效的免疫刺激剂，以诱导Th2/Th1偏移。这些药物在以Th2免疫反应为主的过敏性疾病中显示出有益的治疗效果。在小鼠和人类的体外研究中，通过明胶纳米颗粒（GNPs）递送CpG-ODN可增强细胞摄取，这是本试验的起点。本研究的目的是在马支气管肺泡灌洗（BAL）细胞上，就物种特异性而言，确定一种对马具有最佳刺激作用的CpG基序，从使用的CpG-ODN可能产生的特异性免疫调节作用（Th2/Th1偏移）方面进行研究。因此，评估了GNPs作为一种递送系统，以改善马BAL细胞中CpG-ODN的免疫刺激作用。从7匹患有中度RAO的马和4匹健康马中获取支气管肺泡灌洗液（BALF），随后将其与5种不同的CpG-ODN序列（来自A、B和C类）以及1种无任何CpG基序的ODN一起孵育。通过酶联免疫吸附测定（ELISA）对三种关键细胞因子（IL-4、IL-10和IFN-γ）的释放进行定量，以检测过敏介导的Th2免疫反应（IL-4）以及促炎性Th1反应（IFN-γ）。由于其特定的抗炎和抗过敏作用，IL-10被认为是RAO病理生理学中的一种有益因子。结果显示，一方面，RAO患病马中IL-10和IFN-γ显著上调，另一方面，IL-4下调。与源自RAO的细胞相比，健康马的细胞培养物对所有施加刺激的细胞因子释放反应显著更强。比较所有5种CpG序列，A类2216对马BALF细胞显示出最高的免疫调节作用，因此被选用于后续的初步临床研究。
