中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2014年
6期
512-517
,共6页
实验性自身免疫性葡萄膜视网膜炎%内毒素,脂多糖%动物模型%C57BL/6(H-2b)小鼠
實驗性自身免疫性葡萄膜視網膜炎%內毒素,脂多糖%動物模型%C57BL/6(H-2b)小鼠
실험성자신면역성포도막시망막염%내독소,지다당%동물모형%C57BL/6(H-2b)소서
Experimental autoimmunity uveoretinitis%Endotoxin,lipopolysaccharide%Model%C57BL/6 (H-2b) mouse
背景 葡萄膜炎的发病机制和治疗仍是目前的研究热点,但多年来该领域的基础研究仍是沿用传统的造模方法制备相关的动物模型,与人类的葡萄膜炎自然病程有较大偏差. 目的 本研究用大肠杆菌内毒素,即脂多糖(LPS)替代百日咳毒素(PTX)作为主要诱发因素,建立更符合人类自然发病环境的新型实验性自身免疫性葡萄膜视网膜炎(EAU)的动物模型,并与传统的造模方法进行比较,为研究该病的发病机制和有效的治疗方案提供实验依据.方法 6～8周龄的无特定病原体级雌性C57BL/6(H-2b)小鼠20只,按随机数字表法分为正常对照组、单纯内毒素注射组(EIU组)、多肽+完全弗氏佐剂(CFA)注射组(EAU组)、多肽+CFA+LPS组(LPS-EAU组).LPS-EAU组先用人类光感受器间维生素A类结合蛋白(IRBP 1-20)+CFA免疫小鼠,免疫后第7天小鼠足底注射LPS,诱发小鼠EAU模型.采用组织病理学损害、眼球组织病理学评分、迟发型过敏反应、特异性淋巴细胞增生反应等评价指标对动物模型进行鉴定,并与LPS诱导的EIU及IRBP 1-20+ CFA免疫诱导的EAU进行比较.结果 正常对照组小鼠虹膜睫状体及视网膜组织结构未见异常;EIU组小鼠虹膜睫状体可见轻微血管扩张、蛋白及纤维素渗出,但玻璃体和视网膜组织内未见血管异常及炎症反应;EAU组小鼠虹膜睫状体未见血管扩张及炎性渗出,但可见视网膜轻微血管周围炎及神经纤维层肿胀;LPS-EAU组小鼠视网膜结构紊乱,可见较多的炎性细胞浸润、光感受器细胞损伤及视网膜全层破坏.正常对照组小鼠和EIU组小鼠病理评分均为0分,EAU组病理评分为0.5分,而LPS-EAU组小鼠病理评分为3.0分,显著高于EAU组,差异有统计学意义(U=16.246,P=0.001).LPS-EAU组小鼠耳廓增厚值为(35.60±0.55) tm,显著高于EIU组小鼠的(12.60±0.55)μm,差异均有统计学意义(q=23.003,P＜0.01);但与EAU组小鼠的(34.80±0.84)μm比较,差异无统计学意义(q=0.820,P＞0.05).LPS-EAU组小鼠的脾细胞体外培养的克隆数显著增加,其3 HTdR掺入值(CPM)为(8 540.00±54.77)/min,而EAU组的cpm为(8 484.00±47.75)/min,差异无统计学意义(q=56.634,P=0.069),但与EIU组的cpm(2 050.00±50.00)/min比较,LPS-EAU组明显升高,差异有统计学意义(q=195.683,P=0.000). 结论 LPS可以成功诱发小鼠的EAU,该动物模型在模拟病因方面更符合人类自然发病环境,为研究人类EAU的病因和发病机制提供了一个可能更好的动物模型.
揹景 葡萄膜炎的髮病機製和治療仍是目前的研究熱點,但多年來該領域的基礎研究仍是沿用傳統的造模方法製備相關的動物模型,與人類的葡萄膜炎自然病程有較大偏差. 目的 本研究用大腸桿菌內毒素,即脂多糖(LPS)替代百日咳毒素(PTX)作為主要誘髮因素,建立更符閤人類自然髮病環境的新型實驗性自身免疫性葡萄膜視網膜炎(EAU)的動物模型,併與傳統的造模方法進行比較,為研究該病的髮病機製和有效的治療方案提供實驗依據.方法 6～8週齡的無特定病原體級雌性C57BL/6(H-2b)小鼠20隻,按隨機數字錶法分為正常對照組、單純內毒素註射組(EIU組)、多肽+完全弗氏佐劑(CFA)註射組(EAU組)、多肽+CFA+LPS組(LPS-EAU組).LPS-EAU組先用人類光感受器間維生素A類結閤蛋白(IRBP 1-20)+CFA免疫小鼠,免疫後第7天小鼠足底註射LPS,誘髮小鼠EAU模型.採用組織病理學損害、眼毬組織病理學評分、遲髮型過敏反應、特異性淋巴細胞增生反應等評價指標對動物模型進行鑒定,併與LPS誘導的EIU及IRBP 1-20+ CFA免疫誘導的EAU進行比較.結果 正常對照組小鼠虹膜睫狀體及視網膜組織結構未見異常;EIU組小鼠虹膜睫狀體可見輕微血管擴張、蛋白及纖維素滲齣,但玻璃體和視網膜組織內未見血管異常及炎癥反應;EAU組小鼠虹膜睫狀體未見血管擴張及炎性滲齣,但可見視網膜輕微血管週圍炎及神經纖維層腫脹;LPS-EAU組小鼠視網膜結構紊亂,可見較多的炎性細胞浸潤、光感受器細胞損傷及視網膜全層破壞.正常對照組小鼠和EIU組小鼠病理評分均為0分,EAU組病理評分為0.5分,而LPS-EAU組小鼠病理評分為3.0分,顯著高于EAU組,差異有統計學意義(U=16.246,P=0.001).LPS-EAU組小鼠耳廓增厚值為(35.60±0.55) tm,顯著高于EIU組小鼠的(12.60±0.55)μm,差異均有統計學意義(q=23.003,P＜0.01);但與EAU組小鼠的(34.80±0.84)μm比較,差異無統計學意義(q=0.820,P＞0.05).LPS-EAU組小鼠的脾細胞體外培養的剋隆數顯著增加,其3 HTdR摻入值(CPM)為(8 540.00±54.77)/min,而EAU組的cpm為(8 484.00±47.75)/min,差異無統計學意義(q=56.634,P=0.069),但與EIU組的cpm(2 050.00±50.00)/min比較,LPS-EAU組明顯升高,差異有統計學意義(q=195.683,P=0.000). 結論 LPS可以成功誘髮小鼠的EAU,該動物模型在模擬病因方麵更符閤人類自然髮病環境,為研究人類EAU的病因和髮病機製提供瞭一箇可能更好的動物模型.
배경 포도막염적발병궤제화치료잉시목전적연구열점,단다년래해영역적기출연구잉시연용전통적조모방법제비상관적동물모형,여인류적포도막염자연병정유교대편차. 목적 본연구용대장간균내독소,즉지다당(LPS)체대백일해독소(PTX)작위주요유발인소,건립경부합인류자연발병배경적신형실험성자신면역성포도막시망막염(EAU)적동물모형,병여전통적조모방법진행비교,위연구해병적발병궤제화유효적치료방안제공실험의거.방법 6～8주령적무특정병원체급자성C57BL/6(H-2b)소서20지,안수궤수자표법분위정상대조조、단순내독소주사조(EIU조)、다태+완전불씨좌제(CFA)주사조(EAU조)、다태+CFA+LPS조(LPS-EAU조).LPS-EAU조선용인류광감수기간유생소A류결합단백(IRBP 1-20)+CFA면역소서,면역후제7천소서족저주사LPS,유발소서EAU모형.채용조직병이학손해、안구조직병이학평분、지발형과민반응、특이성림파세포증생반응등평개지표대동물모형진행감정,병여LPS유도적EIU급IRBP 1-20+ CFA면역유도적EAU진행비교.결과 정상대조조소서홍막첩상체급시망막조직결구미견이상;EIU조소서홍막첩상체가견경미혈관확장、단백급섬유소삼출,단파리체화시망막조직내미견혈관이상급염증반응;EAU조소서홍막첩상체미견혈관확장급염성삼출,단가견시망막경미혈관주위염급신경섬유층종창;LPS-EAU조소서시망막결구문란,가견교다적염성세포침윤、광감수기세포손상급시망막전층파배.정상대조조소서화EIU조소서병리평분균위0분,EAU조병리평분위0.5분,이LPS-EAU조소서병리평분위3.0분,현저고우EAU조,차이유통계학의의(U=16.246,P=0.001).LPS-EAU조소서이곽증후치위(35.60±0.55) tm,현저고우EIU조소서적(12.60±0.55)μm,차이균유통계학의의(q=23.003,P＜0.01);단여EAU조소서적(34.80±0.84)μm비교,차이무통계학의의(q=0.820,P＞0.05).LPS-EAU조소서적비세포체외배양적극륭수현저증가,기3 HTdR참입치(CPM)위(8 540.00±54.77)/min,이EAU조적cpm위(8 484.00±47.75)/min,차이무통계학의의(q=56.634,P=0.069),단여EIU조적cpm(2 050.00±50.00)/min비교,LPS-EAU조명현승고,차이유통계학의의(q=195.683,P=0.000). 결론 LPS가이성공유발소서적EAU,해동물모형재모의병인방면경부합인류자연발병배경,위연구인류EAU적병인화발병궤제제공료일개가능경호적동물모형.
Background The pathogenesis and management of human autoimmunity uveoretinitis is a focus in ophthalmology.For decades,a traditional experimental autoimmunity uveoretinitis (EAU) induced by pertussis toxin (PTX) was used for the basic investigation,which was thought to have a large deviation from the natural course of human autoimmunity uveoretinitis.Objective This study was to establish a new mice model of autoimmunity uveoretinitis which mimics the human autoimmunity uveoretinitis pathogenesis and offer a basis for the investigation and treatment of uveoretinitis.Methods Twenty 6-8 weeks old specific pathogen-free female C57BL/6 (H-2b) mice were randomized into normal control group,only endotoxin (lipopolysaccharide,LPS) induced uveitis group (endotoxin induced uveitis [EIU] group),interphotoreceptor retinoid-binding protein (IRBP1-20) +complete Freund adjuvant (CFA) induced uveoretinitis group (EAU group) and IRBP+CFA+LPS induced uveoretinitis group (LPS-EAU group).The mice of the EAU were only immunized with IRBP emulsified in CFA,and LPS-EAU group firstly were immunized with IRBP emulsified in CFA and then LPS was injected in the footpad of the mice on 7 days following immunization.The ocular pathological examination,histopathological scoring,delayed-type hypersensitivity and specific lymphocyte proliferating response were evaluated and compared with the EIU models,traditional EAU models without PTX and LPS-EAU models.The use and care of experimental animals complied with Regulation for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results No inflammatory response was found in the iris,cilliary body and retina of mice in the normal control group.However,mild blood vessels dilation and fibrin exudation were seen in the iris and cilliary body of mice in the EIU group.In the EAU group,mild vasculitis and swelling of nerve fiber layer were exhibited in the retinas; while in the LPS-EAU group,severe disorder of retinal structure,infiltration of inflammatory cells and damage of photoreceptor were found under the optical microscope.The pathological score was 0 in the models of the normal control group and EIU group,0.5 score in the EAU group and 3.0 scores in the LPS-EAU group,with a significant difference in the pathological scores between the EAU group and the LPS-EAU group (U=16.246,P =0.001).The earthickness of the mice was (35.60±0.55) μm in the LPS-EAU group,and this value was significantly higher than (12.60±0.55) μm of the EIU group (q =23.003,P＜0.01),but closed to (34.80±0.84) μm of the EAU group (t =0.820,P＞0.05).The obvious cloning were seen and theradiation count per minute was (8 540.00 ±54.77)/min in the model mice of the LPS-EAU group,and that in the EAU group was (8 484.00±47.75)/min,without significant difference between them (q =56.634,P =0.069).Compared with the β particle number (2 050.00±50.00)/min in the EIU group,that of the LPS-EAU group was significantly elevated (q =195.683,P =0.000).Conclusions LPS injection can induce EAU in mice,and this model can better imitate the pathogenesis of human autoimmunity uveoretinitis.
