诱导新生小鼠对Graves病免疫耐受的研究  

Study on induction of immunotolerance against Graves＇ disease in neonatal mice

作　　者：伍丽萍[1] 旬利茹[1] 施秉银[1] 杨靖[1] 徐利[1] 田竹芳[1] 高珊[1] 张煜[1] 

机构地区：[1]西安交通大学医学院第一附属医院内分泌科,71006l

出　　处：《中华内分泌代谢杂志》2012年第9期744-749,共6页Chinese Journal of Endocrinology and Metabolism

基　　金：国家自然科学基金项日（30371335）

摘　　要：目的研究TSH受体（TSHR）289基因诱导Graves病新生免疫耐受的可行性，并初步探索其可能机制。方法将出牛0～24hBALB／c雌性小鼠分为腹腔注射组、肌肉注射组、造模组及正常对照组，腹腔注射组或肌肉注射组分别设低剂量、中剂量、高剂量耐受组及相应对照组，将腹腔注射、肌肉注射不同剂量耐受组及各自对照组给予1×10。particles（低）、1×10^8particles（中）、1×10^10particles（高）的Ad—TSHR289或Ad—lacz进行预处理。6～7周后．除正常对照组肌肉注射Ad—lacz外，其余各组小鼠均给予Ad—TSHR289免疫，每3周1次．共3次，首次免疫后10d检测血清促甲状腺素受体抗体（TRAb），末次免疫后4周测血清TRAb、TT4、脾CD4＋CD25＋Foxp3／CD4＋比例，并行甲状腺组织学检查。结果首次注射后10d，腹腔注射及肌肉注射高剂量耐受组均未诱导Ⅲ针对TSHR的抗体反应，而对照组TRAb滴度明显升高（P〈0．05）。术次免疫后4周，腹腔及肌肉注射高剂量耐受组均只有1／10小鼠出现阳性的抗体反应，腹腔注射高剂量耐受组没有小鼠发生甲状腺功能亢进，而肌肉注射高剂耐受量组2／10小鼠出现轻微的TT4升高，其相应对照组却出现了强烈的抗体反应（P〈O．01），TT4水平明显升高（P〈0．05），所有TT4升高的小鼠均出现了甲状腺组织增生件改变。此外，腹腔注射及肌肉注射高剂量耐受组CD4＋CD25＋Foxp3／CD4＋比例与对照组比较明显增高（P〈O．01）。腹腔注射及肌肉注射其余组群，与其对照组和造模组比较Graves病发生率差异无统计学意义。结论TSHR289基因可以通过腺病毒为载体，腹腔和肌肉注射两种途径诱导新生小鼠成年后对Graves病免疫耐受，抑制Graves病的发生。而高剂量的抗原刺激容易导致耐受产生。CD4＋cD25＋Foxp3＋T细胞存免疫耐受的诱导和维持中可能起重要作用。Objective To investigate the feasibility of inducing neonatal immunotolerance against Graves＇ disease by gene TSH receptor （TSHR） 289 and its possible mechanism. Methods Neonatal （ 0-24 h） female BALB/e mice were divided into intraperitoneal injection group, intramuscular injection group, model group, and normal control group. The intraperitoneal group and the intramuscular group were further divided into low-dosage, middle-dosage, high-dosage tolerance groups, and the corresponding control groups. The tolerance groups and the controls were intraperitoneally or intramuscularly pretreated with low-dosage （ 1× 10^6 particles ） , middle-dosage （ 1 × 10^8 particles）, high-dosage（ 1 ×10^10 particles） of Ad-TSHR 289 or Ad-laez respectively. 6 to 7 weeks later, the normal control group received intramuscular injection with Ad-lacz; the other groups were immunized with Ad-TSHR 289, three times at 3 weeks interval. 10 （lays afler the first immunization, serum TRAb was detected. 4 weeks after the last immunization, senlm TRAb, TT4, splenic CD4 ＋ CD25 ＋ Foxp3/CD4 ＋ were tested, and the thyroid tissues were examinated histologically. Results Ten days after the first immunization, no antibody response against TSHR was detected in the two high-dose tolerance groups, but the TRAb titer in respective controls was significantly higher（ P〈 0.05 ）. 4 weeks afler the last injection, in high-dose tolerance grnups, only 1/10 of mice immunized by intraperitoneal or intramuscular injection elicited anti-TSHR antibody, and no mice immunized intraperitoneally had elevated serum TT4. Two of ten mice challenged intramuscularly showed slightly increased TT4 levels, but the respective controls displayed a strong antibody response（ P〈0.01 ） and elevated TT4 level （P〈0.05）. The similar percentages of high TT4 and thyroid hyperplasia were found in all groups. Additionally, the frequencies of CD4 ＋CD25 ＋ Foxp3/CD4 ＋in two high-dose tolerance groups were significantly increased as compared t

关 键 词：格雷夫斯病 动物模型 小鼠 免疫耐受 

分 类 号：R581.1[医药卫生—内分泌]