假体无菌性松动过程中Co^2+对成骨前体细胞的生物学反应  

Biological response of Co^2+to preosteoblasts during aseptic loosening of the prosthesis

作　　者：蒋昇源 李丹 姜建浩 杨上游 杨淑野[1] Jiang Shengyuan;Li Dan;Jiang Jianhao;Shang-You Yang;Yang Shuye(Department of Traumatic Orthopedics,Binzhou Medical University Hospital,Binzhou 256600,Shandong Province,China;Chongqing Wushan County People’s Hospital,Chongqing 404700,China;Department of Orthopedic Surgery,the University of Kansas School of Medicine-Wichita,Wichita,KS,USA;Department of Biological Sciences,Wichita State University,Wichita,KS,USA)

机构地区：[1]滨州医学院附属医院创伤骨科,山东省滨州市256600 [2]巫山县人民医院,重庆市404700 [3]美国堪萨斯州堪萨斯大学威寄托骨科中心,堪萨斯州威寄托市,美国 [4]美国堪萨斯州威寄托州立大学生物系,堪萨斯州威寄托市,美国

出　　处：《中国组织工程研究》2021年第21期3292-3299,共8页Chinese Journal of Tissue Engineering Research

基　　金：山东省医药卫生科技发展计划项目(2016WS0023),项目负责人:杨淑野;滨州医学院科研计划(BY2016KYQD19),项目负责人:杨淑野。

摘　　要：背景:假体无菌性松动是人工关节置换后的主要并发症。目前金属离子已被证实参与人工假体无菌性松动的过程,如何控制和减缓假体松动成为当前研究的热点。目的:观察在假体松动过程中,不同浓度的二价钴离子(Co^2+)刺激成骨前体细胞的生物学反应。方法:(1)体外实验:小鼠成骨前体细胞(MC3T3-E1)分别与含不同浓度Co^2+的成骨细胞诱导液共同培养72 h,诱导为成骨细胞;应用MTT法检测细胞的增殖能力;通过测定乳酸脱氢酶活性来反映不同浓度Co^2+的细胞毒性;测定血清碱性磷酸酶蛋白浓度来检测成骨前体细胞向成骨细胞转化的能力;应用RT-PCR测定相关因子mRNA表达;(2)体内实验:将小鼠分为3组,稳定对照组将钛钉置入小鼠的胫骨近端、松动对照组置入钛钉和钴铬颗粒、钴离子组置入钛钉和钴铬颗粒并注入经过钴离子刺激的成骨前体细胞,术后立即使用MicroCT进行假体周围骨密度测定,5周后再次行假体周围骨密度测定,麻醉处死小鼠并离断患侧膝关节进行拔钉实验,并将拔钉后的组织进行苏木精-伊红染色。通过拔钉力量来测定假体松动程度;通过假体与骨界面之间膜的厚度来反映炎症反应的严重程度;通过抗酒石酸酸性磷酸酶染色观察假体周围组织中破骨细胞数量。结果与结论:(1)体外实验结果:当Co^2+浓度升高时,成骨前体细胞的增殖会受到抑制;Co^2+对成骨前体细胞表达血清碱性磷酸酶具有显著的抑制作用;Co^2+可促进单核细胞趋化蛋白1、肿瘤坏死因子α、白细胞介素6、核因子KB受体活化因子配基(RANKL)、活化T细胞核因子c1(NFATc1)mRNA表达,抑制骨保护素及成骨细胞特异性转录因子mRNA表达;低浓度Co^2+(62μmol/L)促进低密度脂蛋白受体相关蛋白(Lrp-5)和Runx2 mRNA表达,高浓度Co^2+(500μmol/L)抑制其表达;(2)体内实验结果:MicroCT扫描显示钴离子组小鼠骨密度值最低(P<0.05);钴离子组中BACKGROUND:Aseptic loosening of prosthesis is the main long-term complication after artificial joint replacement.Metalions have been proven to be one of the causes of aseptic loosening,How to control and mitigate aseptic loosening is an issue of concern.OBJECTIVE:To observe the biological response of preosteoblasts challenged with Co^2+during aseptic loosening of the prosthesis.METHODS:(1)In vitro:Preosteoblasts(MC3T3-E1)of mice were co-cultured with osteoblast induction solution of mice containing diffe rent concentrations of Co^2+for 72 hours,respectively,and nduced into osteoblast cells.The cell proliferation was tested by MTT assay and the cytotoxicity of diffe rent concentrations of Co^2+was measured with the activity of lactate dehydrogenase.The concentration of alkaline phosphatase protein was used to detect the transformation ability of preosteoblasts.RT-PCR was performed to detect the mRNA expression of related factors.(2)In vivo:titanium nails were implanted into the proximal tibia of mice.The mice were divided into three groups.Mice in the stable control group were implanted with titanium nails.Mice in the loosening control group were implanted with titanium nails and cobalt-chromium particles.Mice in the cobalt ion group were implanted with titanium nails and cobalt-chro mium particles and injected with cobalt-stimulated preosteoblasts.Bone mineral density around prosthesis was detected by MicroCT scanning immediately after surgery.Five weeks later,the bone density around the prosthesis was measured again.The mice were sacrificed and the affected knee joints were dissected for the pull-out test.The tissue after nail pull was stained with hematoxylin and eosin.The looseness of the prosthesis was determined by the force of the nail pull.The degree of inflammation was reflected by the thickness of the membrane between the prosthesis and the bone interface.The number of osteoclasts in the tissues around the prosthesis was observed by anti-tartrate acid phosphatase staining.RESULTS AND CONCLUSION:(1)In vit

关 键 词：成骨前体细胞 金属离子 磨损颗粒 无菌性松动 炎症因子 破骨细胞 肿瘤坏死因子 白细胞介素6 碱性磷酸酶 组织工程 

分 类 号：R453[医药卫生—治疗学] R363[医药卫生—临床医学]