以转基因动物研究类风湿关节炎的易感基因

Study on susceptible gene of rheumatoid arthritis by transgenic animal

目的通过人ＨＬＡＤＲ４转基因小鼠的胶原性关节炎动物模型研究易感基因对类风湿关节炎及药物治疗的影响。方法将人ＨＬＡＤＲα＋４β基因转入Ｃ５７ＢＬ／６×ＤＢＡ／１Ｆ１代小鼠复制出转基因动物，并予胶原和佐剂免疫建立胶原性关节炎模型。以分子生物学技术检测外源基因的整合和表达，以发病时间、病理评分及血清Ⅱ型胶原抗体水平研究ＨＬＡＤＲ４基因对关节炎易感性及甲氨蝶呤治疗的影响。在体外通过淋巴细胞增殖及阻断实验研究Ⅱ型胶原及抗ＨＬＡＤＲ、Ｌｙｔ２和Ｌ３Ｔ４单抗对淋巴细胞增殖的影响。结果①人ＨＬＡＤＲ４基因稳定整合于Ｃ５７ＢＬ／６×ＤＢＡ／１Ｆ１代小鼠，主要在脾、肾中表达。②ＨＬＡＤＲ４转基因小鼠对照组发病时间较非转基因小鼠组提早（Ｐ＜００５）。③ＨＬＡＤＲ４转基因小鼠的对照组总病理评分和软骨破坏评分较相应的甲氨蝶呤治疗组高（Ｐ＜００５），非转基因小鼠的对照组的总病理评分、软骨破坏评分和骨质侵蚀评分较相应的甲氨蝶呤治疗组高（Ｐ＜００５，Ｐ＜００１）。④ＨＬＡＤＲ４转基因小鼠对照组和甲氨蝶呤治疗组的Ⅱ型胶原抗体水平较相应的非转基因小鼠组高（Ｐ＜００５，Ｐ＜０００１），且均高于二组正常血清对照组（Ｐ＜?

Objective To determine the effect of susceptible gene on etiology,pathology and drug therapy of rheumatoid arthritis by collageninduced arthritis (CIA) of HLADR4 transgenic mice.Methods Human DRDR4 (Dw4) genes were introduced into C57BL/6DBA/1F1 mice to generate transgenic animal.then the model of CIA was constructed by systemic immunization with bovine collagen and adjuvants.By molecular techniques,the integration and expression of exogenous genes were detected.Day of onset,pathologic evaluation and serum level of anticollagen antibody of CIA made acquire associations between susceptible gene and arthritis and therapy effects of methotrexate (MTX).In vitro,the influence of collagen and anti HLADR,Lyt2 and L3T4 antibody to lymphocyte proliferation was studied. Results HLADR4 genes were detected steadily integrating in C57BL/6DBA/1F1 mice,which were expressed in spleen and kidney.The onset of disease in control group of HLADR4 transgenic mice of control group was earlier than that in nontransgenic mice (P<005).The total pathologic and cartilage damage evaluation in control group of HLADR4 transgenic mice was higher than in MTX group (P<005).The total pathologic,cartilage damage and bone erosion evaluation in control group of nontransgenic mice was higher than in MTX group (P<005,P<0001).The level of anticollagen antibody in control and MTX group of HLADR4 transgenic mice was higher than in these of nontransgenic mice (P<001,P<0001),which were all higher than their normal serum controls (P<001,P<0001).Collagen could stimulate proliferation of lymphocyte (P<005),which could be blocked by antiHLADR antibody and antiL3T4 antibody (P<001). Conclusion HLADR4 gene could enhance RA susceptiblility by earlier onset of arthritis and higher level of anticollagen antibody which would reduce response to drug therapy.MTX could ameliorate the influence of susceptible gene on onset of arthritis and alleviate cartilage damage and bone erosion of arthritis.