Abstract: Objective　To establish animal models of impaired upper respiratory tract mucosal immune function induced by different methods, providing experimental evidence for the study of disease mechanisms and prevention strategies.Methods　Fifty male BALB/c mice were randomly divided into 10 groups, control groups (A0, B0) and model group 1 (A1, B1), model group 2 (A2, B2), model group 3 (A3, B3), and model group 4 (A4, B4), with five mice in each group. Except for the control groups, all other groups underwent modeling, with group A receiving continuous modeling for 7 d and group B for 14 d. The specific modeling method were as follows: model group 1 (-20 ℃), model group 2 (4 ℃), model group 3 (-20 ℃ + cyclophosphamide), and model group 4 (4 ℃ + cyclophosphamide). After continuous modeling for 7 d and 14 d, changes in mouse behavior, body weight, and immune organ (thymus, spleen) indices were observed. Hematoxylin-eosin (HE) staining was used to examine pathological changes in the upper respiratory tract mucosa (nose, oral cavity, throat). Enzyme-linked immunosorbent assay (ELISA) was employed to measure immune factor levels in saliva and serum. Immunofluorescence was used to localize polymeric immunoglobulin receptor (PIgR) and secretory component (SC) proteins.Results　After 7 d of modeling, compared with control group A0, mice in model groups A1, A3, and A4 showed reduced spontaneous activity, poor mental state, and significant decreases in body weight, thymus index, and spleen index (P<0.05). Hemorrhage, edema, and inflammatory cell infiltration were observed in the upper respiratory tract mucosa. Model group A2 showed a significantly decreased thymus index (P<0.05), but no significant difference in spleen index. The detection results of immune factors in saliva and serum showed that the levels of lysozyme (LZM), secretory immunoglobulin A (sIgA), and immunoglobulin A (IgA) in the saliva of groups A1, A3, and A4 were significantly decreased (P<0.05). In serum, LZM and IgA were significantly decreased in group A1 (P<0.05), while sIgA showed no significant difference. Serum LZM, sIgA, and IgA were all significantly decreased in group A3 (P<0.05). Serum LZM and IgA were significantly decreased in group A4 (P<0.05), while sIgA showed no significant difference. In model group A2, only the salivary sIgA content was significantly decreased (P<0.05), and there were no significant differences in other indicators compared with the control group A0. After 14 d of modeling, compared with the control group B0, mice in model groups B1, B2, B3, and B4 all showed reduced spontaneous activity, listlessness, significantly decreased body mass, thymus index, and spleen index (P<0. 05), and persistent pathological changes in the upper respiratory mucosa. The detection results of immune factors showed that salivary and serum levels of LZM, sIgA, and IgA in groups B1, B3, and B4 were significantly lower than those in the control group (P<0.05). In group B2, salivary LZM, sIgA, and IgA were significantly decreased (P<0.05), and serum LZM and sIgA were significantly decreased (P<0.05), while serum IgA showed no significant difference.Conclusion　Exposure to -20 ℃ environmental stimulation for 20 min, twice daily, for 7 consecutive days can induce impaired upper respiratory tract mucosal immunity in mice. In contrast, crating a corresponding model under 4 ℃ conditions requires either combined treatment with cyclophosphamide or extended modeling duration.

Key words: upper respiratory tract mucosa, cold-stress, cyclophosphamide, low mucosal immunity, secretory immunoglobulin A

摘要： 目的　建立不同方法诱导的上呼吸道黏膜免疫功能低下动物模型,为相关疾病机制研究与防治提供实验依 据。方法　50只雄性BALB/c小鼠随机分为10组,对照组(A0、B0)、模型组1(A1、B1)、模型组2(A2、B3)、模型组 3(A3、B3)和模型组4(A4、B4),每组5只。除对照组外,其余各组均进行造模,其中A组连续造模7 d,B组连续造 模14 d。造模具体方法分别为模型组1(-20 ℃)、模型组2(4℃)、模型组3(-20 ℃+环磷酰胺)、模型组4(4℃+环 磷酰胺)。连续造模7 d和14 d后,观察小鼠状态、体质量及免疫器官(胸腺、脾)指数的变化;采用苏木精-伊红 (HE)染色观察上呼吸道黏膜(鼻、口腔、咽喉)病理变化;酶联免疫吸附测定(ELISA)法检测唾液和血清中免疫因 子水平;免疫荧光观察多聚免疫球蛋白受体(PIgR)和分泌成分(SC)蛋白定位情况。结果　造模7 d后,与对照组 A0相比,模型组A1、A3、A4小鼠自主活动减少、精神状态欠佳,体质量、胸腺指数及脾指数均显著降低(P<0.05), 上呼吸道黏膜出现出血、水肿及炎性细胞浸润;模型组A2胸腺指数显著降低(P<0.05),脾指数无显著差异。唾液 和血清中免疫因子检测结果显示,A1、A3、A4组唾液中免疫因子溶菌酶(LZM)、分泌型免疫球蛋白 A(sIgA)、免疫 球蛋白A(IgA)含量均显著下降(P<0.05);A1组血清中LZM和IgA显著降低(P<0.05),sIgA无显著差异;A3组血 清中LZM、sIgA和IgA均显著降低(P<0.05);A4组血清中LZM和IgA显著降低(P<0.05),sIgA无显著差异;模型 组A2仅唾液中sIgA含量显著降低(P<0.05),其余各项指标与对照组A0相比无显著差异。造模14 d后,与对照 组B0相比,模型组B1、B2、B3、B4小鼠均表现出自主活动减少、精神萎靡,体质量、胸腺指数和脾指数均显著降低 (P<0.05),上呼吸道黏膜病理改变持续存在。免疫因子检测结果显示,B1、B3、B4组唾液和血清中LZM、sIgA和 IgA含量均显著低于对照组(P<0.05);B2组唾液中LZM、sIgA和IgA含量均显著降低(P<0.05),血清中LZM和 sIgA显著降低(P<0.05),IgA无显著差异。结论　-20 ℃环境刺激 20 min,2次/d,连续7 d可造成小鼠上呼吸道 黏膜免疫低下,而4℃环境条件制作相应模型需要联合环磷酰胺或者延长造模时间。

关键词: 上呼吸道黏膜, 寒冷刺激, 环磷酰胺, 黏膜免疫功能低下, 分泌型免疫球蛋白A