课题背景：课题受国家自然科学基金项目（30560155）和教育部科学技术研究重点项目（206161）资助。课题组在前期实验中证实依那西普在体外可以抑制破骨细胞的增殖与活性，并且可以抑制磨屑诱导巨噬细胞分泌肿瘤坏死因子α。本实验进一步观察依那西普在体内对钛颗粒诱导小鼠骨溶解及破骨细胞增殖的影响，阐释依那西普抑制骨溶解的机制。

同行评价：选择肿瘤坏死因子α的拮抗剂依那西普为切入点，采用动物体内实验着重探讨关节置换后假体松动的早期防治，有一定的创新点，为临床解决关节置换后假体松动问题提供了一定的实验依据。

偏倚或不足：实验未在分子生物学等方面对依那西普防治骨溶解及破骨细胞增殖的原理进行进一步观察，在可能应用于临床之前，还需用大动物作进一步验证及长时间的观察。

摘要
目的：已证实肿瘤坏死因子α在人工关节无菌性松动和类风湿关节炎的发病机制中均发挥重要的作用。依那西普作为肿瘤坏死因子α拮抗剂在治疗类风湿关节炎中疗效显著。拟通过实验观察依那西普对钛颗粒诱导小鼠骨溶解及破骨细胞增殖的影响，为依那西普防治人工关节无菌性松动提供实验依据。
方法：实验于2007-02/2007-08在宁夏医学院生殖及遗传学重点实验室和宁夏医学院附属医院病理学实验室完成。①60只BALB/C小鼠随机分为6组，每组10只。分别为空白对照组，单纯钛处理组，20，50，100，200 μg/kg依那西普+钛处理组。实验过程中对动物处置符合动物伦理学要求。依那西普由美国Wyeth公司提供（生产批号：P321686）；钛金属颗粒由美国Zimmer公司提供。②建立钛颗粒骨吸收模型，其中空白对照组行原位缝合，不置入钛颗粒。依那西普各组分别在第0，2，4，6，8天腹腔注射不同剂量的依那西普(20，50，100，200 μg/kg)。10 d后取出颅骨，行常规苏木精-伊红染色，了解骨吸收情况。抗酒石酸酸性磷酸酶染色计算破骨细胞数量。
结果：苏木精-伊红染色结果为单纯钛处理组骨吸收明显增加，骨小梁结构紊乱及骨质疏松,破骨细胞的形态变大，数目变多；依那西普干预组上述表现减轻，尤以200 μg/kg剂量为明显。与空白对照组相比，单纯钛处理组可明显增加骨溶解（P < 0.001）及破骨细胞生成（P < 0.001）；与单纯钛处理组相比，依那西普各组可减少骨溶解（P < 0.05）及破骨细胞生成（P < 0.05），尤以200 μg/ kg剂量最为明显（P < 0.05），抗酒石酸酸性磷酸酶染色与苏木精-伊红染色结果相符。
结论：依那西普可抑制钛颗粒诱导的骨溶解及破骨细胞的增殖，有望成为预防人工关节无菌性松动的药物。
关键词：磨损颗粒；依那西普；破骨细胞；骨溶解; 人工关节

张亮，陈志荣，吴兴临，金群华.依那西普对钛颗粒诱导小鼠骨溶解及破骨细胞增殖的影响[J].中国组织工程研究与临床康复，2008，12(9):1601-1604 [www.zglckf.com/zglckf/ejournal/upfiles/08-9/9k-1601(ps).pdf]

Effects of etanercept on Ti particle-induced osteolysis and osteoclastogenesis in mice

Abstract

AIM: It has been demonstrated that tumor necrosis factor-α (TNF-α) plays an important role in the pathogenesy of artificial joint aseptic loosening and rheumatoid arthritis. Etanercept, antagon of TNF-α, shows notable effect in treating rheumatoid arthritis. In this study, we investigated the influence of etanercept on Ti particles-induced osteolysis and osteoclastogenesis in mice and evaluate the feasibility of etanercept for treatment of artificial joint aseptic loosening.
METHODS: The experiment was performed at the Key Laboratory of Genesiology and Genetics, and Pathological Laboratory of Hospital, Ningxia Medical College from February to August 2007. ①Sixty BALB/C mice were randomly divided into 6 groups (n =10): blank control group, Ti (Zimmer, USA) treatment group, and 20, 50, 100, 200μg/kg etanercept (Wyeth, USA, No. P321686) plus Ti treatment group. ②Bone resorption models of Ti particles were established in all groups exception blank control group (in situ suture with no Ti particles). Different doses of etanercept (20, 50, 100, 200μg/kg) were intraperitoneally injected into etanercept groups on days 0, 2, 4, 6, and 8, respectively. Ten days later, mice cranial bones were harvested for HE staining to observe bone resorption. Number of osteoclast was counted using tartrate-resistant acid phosphatase (TRAP).
RESULTS: HE staining showed that bone resorption in Ti treatment group was increased obviously and the structure of bone trabecula was disorderly and osteoporotic; the volume of osteoclast was enlarged and number was increased. The findings were reduced in etanercept groups, especially in 200μg/kg group. Compared with blank control group, Ti particles significantly increased osteolysis (P < 0.001) and osteoclastogenesis (P < 0.001); compared with Ti particles group, etanercept effectively inhibited Ti particles induced osteolysis (P < 0.05) and osteoclastogenesis (P < 0.05), especially in 200μg/kg group (P < 0.05). The result of TRAP was accorded with HE staining.
CONCLUSION: Etanercept can inhibit Ti particles-induced osteolysis and osteoclastogenesis in vivo and is promising to be a therapeutic candidate for the prevention of artificial joint aseptic loosening.

Zhang L, Chen ZR, Wu XL, Jin QH.Effects of etanercept on Ti particle-induced osteolysis and osteoclastogenesis in mice. Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu 2008;12(9):1601-1604(China)
[www.zglckf.com/zglckf/ejournal/upfiles/08-9/9k-1601(ps).pdf]