奶牛趾间皮肤外植体模型的构建及效果评价  

Construction and effect evaluation of skin eplant model between toes of dairy cows

作　　者：张鹤飞 周志新 孙悦 杨春雪 郑家三[1] ZHANG Hefei;ZHOU Zhixin;SUN Yue;YANG Chunxue;ZHENG Jiasan(College of Animal Science and Technology,Heilongjiang Bayi Agricultural University,Daqing 163319,China)

机构地区：[1]黑龙江八一农垦大学动物科技学院,黑龙江大庆163319

出　　处：《中国农业大学学报》2023年第1期172-179,共8页Journal of China Agricultural University

基　　金：国家自然科学基金项目(32172935)。

摘　　要：为建立一种可用于奶牛蹄部感染性疾病研究的皮肤外植体模型,本研究利用气液交界面皮肤组织培养模式离体培养健康奶牛后蹄趾间皮肤,分别于培养0、12、24、36、48和72 h收集皮肤组织并采用HE染色法、原位末端标记(TUNEL)法及免疫组织化学方法对外植体的病理组织变化、组织细胞凋亡及凋亡相关蛋白Caspase-3表达进行检测。结果表明:1)与0 h对照组模型相比,培养12、24和36 h的皮肤组织表皮结构完整,真皮层发生不同程度变性;36 h皮肤组织上皮细胞出现变性迹象;48 h皮肤组织上皮细胞发生不同程度变性、坏死,并伴有核固缩;72 h皮肤组织表皮脱落、缺失,皮肤附属器官发生不同程度坏死,无法保持正常的组织结构和功能。2)皮肤外植体细胞发生不同程度凋亡,Caspase-3蛋白呈阳性表达,且表皮中凋亡细胞数量及Caspase-3蛋白阳性表达情况高于真皮。与对照组相比,24 h皮肤外植体细胞凋亡情况出现显著差异(P<0.05),36 h皮肤外植体Caspase-3蛋白阳性率出现显著差异(P<0.05)。本研究构建了一种稳定的奶牛趾间皮肤外植体模型,离体培养24 h仍保持正常的组织结构、形态功能及活性,为奶牛蹄部感染性疾病体外研究提供一种新方法。In order to establish a skin explant model for the study of infectious diseases of dairy cows’hoof,the skin between the hoof and toe of healthy dairy cows was cultured in vitro by using the air-liquid interface skin tissue culture mode.The skin tissues were collected at 0,12,24,36,48 and 72 h.Histological changes,apoptosis and caspase-3 expression of explants were detected by HE staining,in situ terminal labeling(TUNEL)and immunohistochemistry.The results showed as follows:1)Compared with the 0 h control model,the epidermal structure of skin tissue was intact at 12,24 and 36 h,and the dermis was denatured to varying degrees.The epithelial cells of skin tissue showed signs of degeneration at 36 h.At 48 h,the epithelial cells of skin tissue showed different degrees of degeneration and necrosis,accompanied by nuclear pyknosis.After 72 h,the epidermis of skin tissue was exfoliated and lost,and the accessory organs of skin were necrotic to varying degrees,and the normal tissue structure and function could not be maintained.2)The number of apoptotic cells and caspase-3 protein expression in epidermis were higher than that in dermis.Compared with the control group,the apoptosis of skin explants at 24 h was significantly different(P<0.05),and the positive rate of caspase-3 protein in skin explants at 36 h was significantly different(P<0.05).In this experiment,one stable intertoe skin explants model of dairy cows was established,and the normal tissue structure,morphological function and activity were still maintained after 24 h in vitro culture,which provided a new method for the in vitro study of dairy hoof infectious diseases.

关 键 词：奶牛肢蹄病 动物模型 皮肤外植体 病理变化 细胞凋亡 

分 类 号：S857.5[农业科学—临床兽医学]