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白立鼎,陈银潇,李兆栋,李佳颖,樊亚东,张晓雨,张翟轶,边育红,王相玲,刘建卫,赵舒武.肠上皮细胞 Tlr4 特异特性征敲观除察基因鼠鉴定及免疫学[J].中国实验动物学报,2022,30(6):733~741.
肠上皮细胞 Tlr4 特异特性征敲观除察基因鼠鉴定及免疫学
Identification and immunological characterization of intestinal epithelial cell-specific Tlr4 gene knockout mice
投稿时间:2022-03-15  
DOI:10. 3969 / j.issn.1005-4847. 2022. 06. 001
中文关键词:  Tlr4 基因  肠黏膜  CRISPR/ Cas9  Cre / Loxp  基因敲除
英文关键词:Tlr4 gene  intestinal mucosal  CRISPR/ Cas9  Cre / Loxp  gene knockout
基金项目:
作者单位
白立鼎 天津中医药大学,天津 301617 
陈银潇 天津中医药大学,天津 301617 
李兆栋 天津中医药大学,天津 301617 
李佳颖 天津中医药大学,天津 301617 
樊亚东 天津中医药大学,天津 301617 
张晓雨 天津中医药大学,天津 301617 
张翟轶 天津中医药大学,天津 301617 
边育红 天津中医药大学,天津 301617 
王相玲 天津中医药大学,天津 301617 
刘建卫 天津中医药大学,天津 301617 
赵舒武 天津中医药大学,天津 301617 
Author NameAffiliation
BAI Liding Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
CHEN Yinxiao Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
LI Zhaodong Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
LI Jiaying Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
FAN Yadong Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
ZHANG Xiaoyu Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
ZHANG Zhaiyi Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
BIAN Yuhong Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
WANG Xiangling Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
LIU Jianwei Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
ZHAO Shuwu Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China 
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中文摘要:
       目的 鉴定肠上皮 Tlr4 特异性敲除(Tlr4f/ f cre T)鼠,评价其免疫学特征。 方法 应用 CRISPR/ Cas9 技术构建 Tlr4f/ f cre T 基因鼠,PCR 和免疫荧光鉴定 Tlr4f/ f cre T 基因鼠基因型,观察基因鼠的一般生物学特征、繁殖能力和子代存活率。 HE 染色、流式细胞术及 ELISA 比较基因鼠和野生型小鼠免疫器官结构、肠黏膜免疫细胞比例及细胞因子分泌水平差异。 结果 从基因和蛋白水平验证 Tlr4f/ fcre T 基因鼠的建立。 与野生型鼠比,Tlr4f/ f cre T 基因鼠的一般生物学特征无明显差异、子代存活率 > 90%;胸腺、脾及肝生理结构无显著性差异;脾淋巴细胞增殖能力、血清 及肠黏膜细胞因子分泌水平无显著性差异;但 CD4+T 和 γδT 细胞显著减少。 结论 成功构建了肠上皮 Tlr4 特异性敲除小鼠(Tlr4f/ f cre T),为研究肠上皮 Tlr4 基因在肠道疾病、肿瘤及代谢性疾病中的作用提供实验手段。
英文摘要:
       Objective To study the role of intestinal epithelial Tlr4 in systemic immune effects by establishing a mouse model of conditional Tlr4 knockout in intestinal epithelia. Methods Intestinal epithelial cell-specific Tlr4 gene knockout mice ( Tlr4f/ f Cre T) were established by CRISPR/ Cas9 technology. The genotype of Tlr4f/ f Cre T mice was identified by PCR and immunofluorescence. The general biological characteristics, reproductive capacity, and offspring survival rate were also investigated. Differences in immune organ structures, the intestinal mucosal immune cell ratio, and cytokine secretion levels between wildtype and Tlr4f/ f Cre T mice were compared by HE staining, flow cytometry, and ELISAs. Results Establishment of Tlr4f/ f Cre T mice was verified at gene and protein levels. Compared with wildtype mice, Tlr4f/ f Cre T mice had no significant difference in general biological characteristics and offspring survival rate of > 90%. No significant differences in the physiological structure of the thymus, spleen, or liver, proliferation of splenic lymphocytes, and serum cytokine level, and intestinal mucosa were found between the two groups. However, the number of CD4+ T and γδT cells appeared to be significantly low in Tlr4f/ f Cre T mice. Conclusions The intestinal epithelial cell- specific Tlr4 gene knockout mouse model was successfully established, which provides the experimental means to study the regulatory roles of Tlr4 expression of intestinal epithelial cells in diseases.
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