新型双靶向姜黄素载药纳米粒治疗颗粒诱导骨溶解的实验研究

Aseptic loosening induced by periprosthetic osteolysis (PPO) is the leading complication of total joint arthroplasty (TJA), which makes patients have to accept revision surgery. No drug can effectively prevent or even slow down the pathological process at present. Recent studies showed curcumin can inhibit the whole activation of NF-κB signal pathway through inhibiting the IκB kinase (IKK); therefore it had the ability for the treatment of the osteolysis and bone loss induced by bone metastatic cancer and dentoalveolitis. This project hypothesizes that curcumin can inhibit the osteolysis induced by wear debris by inhibiting the IKK，and intend to prepare a novel dual targeting drug carrying nanoparticle. The nanoparticle was made by poly lactic-co-glycolic acid (PLGA) as the shell with curcumin on the surface; the PLGA shell was contained superparamagnetic iron oxide (SPIO) and coated with zoledronic acid. The nanoparticles had active targeting ability guided by zoledronic acid and passive targeting ability guided by SPIO under the action of magnetic field. The therapeutic effect to the osteolysis induced by wear debris and the targeting ability to the bone tissue of the novel drug carrying on nanoparticles were evaluated by in vitro and vivo experiments. This project will illustrate the mechanism of curcumin for the treatment of osteolysis induced by wear debris, and provide a novel method for targeting treatment for periprosthetic osteolysis after total joint arthroplasty.

假体周围骨溶解引起的假体无菌性松动是人工关节置换术（TJA）最主要的并发症，常致使患者必须接受翻修手术，目前缺乏有效的药物延缓假体周围骨溶解的病理进程。研究发现姜黄素能通过抑制核因子κB抑制蛋白激酶（IKK）的作用，抑制整个NF-κB信号通路的活化，从而对骨转移瘤，牙周炎等疾病引起的骨溶解和骨丢失起到一定的治疗效果。本课题假说姜黄素可通过抑制IKK的作用有效延缓颗粒诱导骨溶解的产生；并拟制备一种新型双靶向的载药纳米粒，以载姜黄素的PLGA作为纳米粒的外壳，外壳内添加超顺磁性氧化铁，外壳表面偶联唑来膦酸，从而使其在外加磁场的作用下，具备唑来膦酸主动靶向和磁场引导被动靶向能力。从体内、体外不同方面评价这种新型载药纳米粒对颗粒诱导骨溶解的治疗效果，以及对骨组织的靶向能力。本课题将阐明姜黄素治疗颗粒诱导骨溶解的作用机制，并为实现TJA术后假体周围骨溶解的靶向治疗提供新的方法和思路。

假体周围骨溶解引起的假体无菌性松动是人工关节置换术最主要的并发症，目前缺乏一种特定有效的药物延缓病理进程，且假体周围骨组织不甚丰富的血液循环也为药物治疗带来了阻碍。因此，两种或多种药物靶向骨组织的联合治疗或可收获更佳疗效。本课题组在体内、外实验中均证明姜黄素可以抑制IκBα蛋白的降解和磷酸化，从而进一步抑制了破骨相关基因c-Fos和NFATc1 mRNA及蛋白的表达，最终实现抑制破骨细胞的诱导分化作用以及颗粒诱导骨溶解的发生。在此基础上研制了一种新型双靶向姜黄素载药纳米粒，以载姜黄素的PLGA作为纳米粒的外壳，外壳内添加超顺磁性氧化铁，外壳表面偶联唑来膦酸，从而使其在外加磁场的作用下，具备唑来膦酸主动靶向和磁场引导被动靶向能力。这种纳米粒粒径足够小，在外加磁场靶向吸引及唑来膦酸的破骨细胞靶向效应下可以穿透血管内皮细胞，可以到达骨溶解部位聚集，从而对动物体内骨溶解部位产生靶向聚集效果。细胞毒性及凋亡实验则证实，制备的新型双靶向载药纳米粒无细胞毒性小，生物安全性好。本研究在颗粒诱导骨溶解动物模型上研究该双靶向载药纳米粒疗效，发现小鼠破骨相关基因P-IκBα、NFATc1和c-Fos的蛋白表达在颗粒诱导骨溶解组显著增高，而在双靶向载药纳米粒治疗后表达则明显降低，IκBα的表达水平则与之相反，表明双靶向载药纳米粒有效抑制了抑制IκBα蛋白的降解和磷酸化，以及抑制了整个NF-κB信号通路的活化，最终抑制了颗粒诱导骨溶解的发生。Micro CT结果可见双靶向纳米粒治疗组骨缺损几乎完全修复，TRAP染色示破骨细胞数量较骨溶解组明显减少，证明其具有较为显著的治疗效果。小鼠体内血药浓度代谢研究证实，新型纳米粒的靶向作用可明显延缓姜黄素在小鼠体内的代谢速度，具有明确的缓释作用。实验小鼠体内的重要脏器HE染色无异常、血肝肾功能重要指标均属正常范围，证明该双靶向载药纳米粒生物安全性好，在颗粒诱导骨溶解的靶向治疗上具有较大潜力和良好前景。