摘要
目的:构建髓细胞选择性表达人类趋化因子受体1(chemokine receptor-like 1,CMKLR1或human chemerinreceptor 23,hChemR23)的转基因小鼠,检测其在小鼠腹膜炎和牙周炎的炎症反应中的抑制作用。方法:将全长hChemR23的cDNA和启动子hCD11b克隆到pcDNA3质粒,转基因片段(2.9 kb)经纯化后由波士顿大学转基因/基因敲除小鼠中心构建转基因小鼠。通过实时定量PCR检测亲代hChemR23转基因的拷贝数和mRNA水平。hChemR23转基因雄性小鼠和野生型FVB雌性小鼠杂交繁殖产生杂合型转基因小鼠后代。采用PCR方法检测子代小鼠基因组DNA明确基因型。转基因小鼠经腹腔注射1 mL酵母多糖A溶液(1 g/L)构建急性腹膜炎模型,灌洗细胞经PE标记的抗鼠F4/80和FITC标记的抗鼠Ly6G染色后,采用FACSort方法检测。在小鼠上颌第一磨牙结扎9-0丝线诱导转基因小鼠牙周炎模型,测量釉牙骨质界距牙槽骨骨嵴顶的距离(cementoenamel junction to thealveolar bone crest,CEJ-ABC)检测牙槽骨的丧失。所有实验数据采用配对t检验和方差分析方法进行统计学分析。结果:4个F1亲代小鼠携带hChemR23基因。hChemR23转基因小鼠腹膜炎腹腔灌洗液中中性粒细胞的含量比野生型减少了56%。在丝线结扎诱导的小鼠牙周炎模型中,hChemR23转基因小鼠比野生型小鼠的牙槽骨骨吸收显著降低(P<0.05)。结论:hChemR23转基因小鼠过表达hChemR23 mRNA水平,在急性腹膜炎和丝线结扎诱导的牙周炎模型中抑制炎症反应。
Objective ： To prepare a transgenic mouse model with myeloid-selective expression of human chemokine-like receptor 1 （ CMKLR1 or hChemR23 ） and investigate the in vivo inflammatory response of the particular transgenic mice in induced-peritonitis and periodontitis. Methods： The full-length hChemR23 cDNA and hCDllb promoters were cloned into pcDNA3 plasmid. Purified transgene （2.9 kb） was used to make transgenic mice at Boston University Transgenic/Knoek out Mouse Core Facility. Copy numbers and mRNA level of hChemR23 transgene in the founders were verified by Custom TaqMan Gene Expression Real-time PCR assay. Hemizygous colonies were amplified by out-breeding hChemR23 transgenic males with wide-type FVB females. Transgenic mice were injected intraperitoneally with 1 mL of zymosan A solution （1 g/L in PBS）. Peritoneal lavage cells stained with PE-conjugated anti-mouse F4/80 and FITC-conjugated anti-mouse Ly6G were analyzed by FACSort. Then 9-0 silk ligature was tied around the first molar to induce a periotontitis mouse model. The distance of cementoenamel junction to the alveolar bone crest （CEJ-ABC） was calculated to evaluate the alveolar bone loss. Data were analyzed with student＇ s t-test and variance analysis. Results： One out of the four F1 founders carried the hChemR23 transgene, hChemR23 transgenic mice had reduced 56% fewer positive labeled polymorpho- nuclear neutrophil （PMNs） in peritonitis abdominal lavage fluid as compared with wide-type littermates. In a ligature-induced peirodontitis mouse model, hChemR23 transgenic mice showed decreased alveolar bone loss as compared with WT littermates （P 〈 0.05 ）. Conclusion： The hChemR23 transgenic mice over-expressed the hChemR23 at mRNA level and had reduced inflammation in the acute peritonitis and ligature-induced periodontitis model.
出处
《北京大学学报（医学版）》
CAS
CSCD
北大核心
2012年第3期469-474,共6页
Journal of Peking University:Health Sciences
基金
国家自然科学基金(81000440)
教育部留学回国启动基金(2009-1342)资助~~
