钛颗粒诱导小鼠骨溶解模型的评价

Establishment and evaluation of titanium particle induced air-pouch osteolysis models in mice

背景:磨损颗粒引起的骨溶解是导致假体松动的主要原因,已成为骨科学的研究热点之一,但是准确模拟磨损颗粒诱导假体周围骨溶解发生过程的动物模型是首先需要解决的问题。RANKL是细胞因子中调控破骨细胞分化、成熟及激活其功能最关键的因子之一。目的:拟建立钛颗粒诱导小鼠air-pouch骨溶解模型,模拟人工关节无菌性松动发生的病理过程。设计、时间及地点:随机对照、动物体内实验,于2008-03/10在苏州大学附属第一医院骨科实验室完成。材料:选用雌性BALB/c小鼠20只,按随机数字表法分成实验组和对照组,每组10只。方法:背侧单点皮下注射3mL无菌空气,间隔1d补注1mL气体,连续5d。囊内植入同种系小鼠颅骨。术后实验组和对照组分别在囊内注射10g/L的钛颗粒0.5mL或0.5mL磷酸盐缓冲液,14d后处死小鼠将囊及植入颅骨一并取出。主要观察指标:应用苏木精-伊红染色观察囊壁组织学改变,以定量反转录-聚合酶链反应检测检测标本中RANK、RANKL和破骨细胞成熟标志因子之一蛋白酶K基因mRNA含量。结果:组织学观察发现,实验组标本囊壁增厚、细胞浸润增多。经钛颗粒刺激后,组织中RANK、RANKL及蛋白酶K基因表达显著高于对照组(P<0.05)。结论:钛颗粒诱导的小鼠air-pouch骨溶解模型,简单、灵敏、经济、重复性好,能准确模拟人工关节松动发生的病理过程。

BACKGROUND： Wear particle associated osteolysis is the leading cause of aseptic loosening, which is the Hotpoint in orthopaedics and tranmstology; however, how establish a titanium particle induced murine air-pouch model of bone resorption is the primary problem. RANKL is one of the key factors in regulating differentiation, maturation and activation of osteoclasts. OBJECTIVE： To develop a titanium particle induced murine air-pouch model of bone resorption to evaluate the pathogenesis of aseptic loosening. DESIGN, TIME AND SETTING： Randomized control study in animals was performed in the Orthopedic Laboratory of the First Affiliated Hospital of Soochow University between May and October in 2008. MATERIALS： Twenty female BALB/c mice were divided into experimental group and control group according to random number table, with 10 animals in each group. METHODS： Three milliliter sterilized air was injected subcutaneously to form an air pouch on the back of BALB/c mice. The air pouches were injected with 1 mL air every other day for 5 days. At the 6th days, a section of calvaria bone from a genetically identical donor mouse was implanted into the pouch. Totally 5 mg titanium particles suspended in 0.5 mL saline or 0.5 mL phosphate buffer saline was injected into pouches in the experimental group and control group, respectively. At the 14th day after bone implantation, the mice were sacrificed and tissues were harvested. MAIN OUTCOME MEASURES： Histological changes were observed with hematoxylin-eosin staining. Expression of RANK, RANKL and CPK gene transcripts were measured by real-time quantitative reverse transcription-polymerase chain reaction （RT-PCR）. RESULTS： The histological analysis showed that the pouch membrane was thicker with more cellular infiltration in the experimental group. Expression of osteolysis-related gene RANK/RANKL and CPK in air pouch tissues were at a high level after titanium particle stimulation than the control group （P 〈 0.05）. CONCLUSION： This air-pouch bone resorption model induced by titanium particle has the advantage of sensitive, rapid, economical and reproducible, which can mimic the pathogenesis of aseptic loosening.