芳香烃受体敲除小鼠的培育和生物学特征及其在严重创伤动物模型中的应用

目的探讨芳香烃受体敲除（AhRKO，AhR-／-）小鼠的饲养、繁育、基因分型和生物学特征，并初步应用于严重创伤研究。方法引进C57BU6背景的AhR杂合子（＋／-）小鼠；配对繁殖后提取仔鼠脚趾基因组脱氧核糖核酸（DNA），通过聚合酶链反应对仔鼠基因型进行鉴定；观察不同基因型仔鼠的生活状态、存活率和体重变化差异等生物学特征；制备AhR-／-和AhR野生型（WT，＋／＋）小鼠“骨折佚血”模型并收集骨髓细胞，采用粒细胞巨噬细胞集落刺激因子（GM—CSF）体外诱导培养骨髓来源树突状细胞（BMDC）7d，脂多糖（LPS）刺激24h后ELISA检测不同品系来源BMDC炎性因子分泌水平的差异。结果AhR-／-小鼠的饲养和繁殖均获得成功，子代小鼠基因型分布符合孟德尔分离定律；相对AhR＋／＋小鼠而言，AhR-／-小鼠发育迟缓，体型小，体重轻（P〈0．05），生活状态差，存活率偏低（P〈0．05）；严重创伤后AhR-／-小鼠来源的成熟BMDC分泌α型肿瘤坏死因子（TNFa）和白细胞介素（IL）-6的能力高于AhR＋／＋，（P〈0．05），而IL-10的分泌水平则低于AhR＋／＋（P〈0．05）。结论通过AhR＋／-繁育和适当的基因分型技术是获得表型稳定的AhR-／-的有效方法；严重创伤后AhR-／-来源的BMDC分泌炎性细胞因子的水平与AhR＋／＋来源BMDC存在显著差异，提示AhR与严重创伤后机体的炎性反应有关。

Breeding and biocharacteristics of aryl hydrocarbon receptor knockout mice and application in severe traumatic animal model

Objective To explore the breeding, genotyping and phenotype of aryl hydrocarbon receptor knockout （AhR KO, AhR-/-） mice and their secretion function of bone marrow-derived dendritic cell （BMDC） in severe trauma. Methods AhR heterozygous （AhR ＋/-） mice of C57BL/6 background were obtained from Jackson Laboratory, U.S.A. and bred based on the genetic rules. The genomie deoxyribonucleic acid （DNA） was extracted from the toes of weaning mice of the second generation, and genotyping was carried out by polymerase chain reaction. The bioeharacteristics of pups such as animation, survival rate and body weight changes were observed. A fracture/hemorrhage model in vivo was established in mice of AhR ＋/＋ and AhR-/-, bone marrow cells-derived dendritic cells （BMDC） in different strains were collected and induced by granuloeyte-macrophage colony-stimulating factor （GM-CSF） in vitro. 7 days later, fipopolysaccharide, （LPS） was added so as to stimulate them to mature. After 24 hours, the secretion levels of tumor necrosis factor α （TNFα）, interleukin 6 （IL-6） and IL-10 in each strain were detected by enzyme linked immunosorbent assay （ELISA）. Results The breeding of AhR-/- mice was carried out successfully and the genotypes ratio of the second generation was consistent with the law of segregation by Mendel. The status of pups in AhR-/- mice was worse than that of the AhR＋/＋ mice, including body weight （P〈 0.05） survival rate （P〈 0.05） and so on. After trauma/hemorrhage, the expression levels of TNFα and IL-6 of the mature BMDC＇s of the AhR-/- strain were significantly higher than that of the AhR＋/＋ strain （P〈 0.05） while the secretion level of IL-10 of the AhR-/- mice was significantly lower than that of the AhR-/- mice （P〈 0.05）.Conclusion Breeding and genotypin of AhR＋/c mice are effective methoda to obtain AhR-/- mice with stable phenotype. After severe trauma, the expression levels of inflammatory cytokines reflect quite different between Ah