现代生物影像方法评价脂多糖与甲基强的松龙联合诱导的实验性骨坏死

目的：建立脂多糖与甲基强的松龙联合诱导的激素性骨坏死家兔模型，同时应用静态和动态的影像学方法评价涉及骨坏死发病的血管内、外机制以及血管结构-功能的病理生理学异常。方法：取14只28周龄雄性新西兰白兔以10μg/kg静脉注射脂多糖(内毒素)。24 h后，每隔24 h以20mg/kg肌肉注射甲基强的松龙(糖皮质激素)。另取6只作为对照于相应时间点注射生理盐水。应用动态核磁共振于内毒素注射前后检查双侧股骨近端骨内灌注功能，并且进行血液学检查;切下双侧股骨脱钙进行基于Micro-CT的微血管造影检查。骨坏死病灶、血管内血栓和血管外骨髓脂肪细胞行组织学观察。 结果：实验组骨坏死家兔标本中均观察到血管内血栓。骨坏死家兔血管外骨髓脂肪细胞尺寸显著大于对照组(P＜0.05)。与基值比较，骨坏死家兔的血浆组织纤溶酶原激活剂/纤溶酶原激活剂抑制因子1及部分凝血激酶时间显著下降，而低密度脂蛋白/高密度脂蛋白显著升高(P＜0.05)。动态核磁共振检查显示骨坏死家兔局部灌注指数"最大信号增强"显著减少(P＜0.05)，同时基于Micro-CT的微血管造影检查显示血管堵塞。实验组93%家兔在接受内毒素与激素联合诱导下发生骨坏死，且未出现死亡。结论：血管内外病变以及血管结构-功能的病理生理学异常参与了激素性骨坏死的发生。该诱导方法在不导致家兔死亡的情况下可以产生高坏死发生率，适合作为未来评价干预激素性骨坏死方法的临床前效能的实验模型。：取14只28周龄雄性新西兰白兔以10μg/kg静脉注射脂多糖(内毒素)。24 h后，每隔24 h以20mg/kg肌肉注射甲基强的松龙(糖皮质激素)。另取6只作为对照于相应时间点注射生理盐水。应用动态核磁共振于内毒素注射前后检查双侧股骨近端骨内灌注功能，并且进行血液学检查;切下双侧股骨脱钙进行基于Micro-CT的微血管造影检查。骨坏死病灶、血管内血栓和血管外骨髓脂肪细胞行组织学观察。 结果 实验组骨坏死家兔标本中均观察到血管内血栓。骨坏死家兔血管外骨髓脂肪细胞尺寸显著大于对照组(P＜0。05)。与基值比较，骨坏死家兔的血浆组织纤溶酶原激活剂/纤溶酶原激活剂抑制因子1及部分凝血激酶时间显著下降，而低密度脂蛋白/高密度脂蛋白显著升高(P＜0。05)。动态核磁共振检查显示骨坏死家兔局部灌注指数"最大信号增强"显著减少(P＜0。05)，同时基于Micro-CT的微血管造影检查显示血管堵塞。实验组93%家兔在接受内毒素与激素联合诱导下发生骨坏死，且未出现死亡。 结论 血管内外病变以及血管结构-功能的病理生理学异常参与了激素性骨坏死的发生。该诱导方法在不导致家兔死亡的情况下可以产生高坏死发生率，适合作为未来评价干预激素性骨坏死方法的临床前效能的实验模型。

OBJECTIVE: The present study employed both static and dynamic imaging modalities to study both intra- and extravascular events attributing to steroid-associated osteonecrosis (ON) using an experimental protocol with a single low-dose lippolysaccharide (LPS) injection and subsequently three injections of high-dose methylprednisolone (MPS). METHODS: Fourteen 28-week-old male New Zealand white rabbits received one intravenous injection of LPS (10 microg/kg). After 24 hours, three injections of 20 mg/kg of MPS were given intramuscularly at a time interval of 24 hours. Additional 6 rabbits were used as controls. Dynamic MRI was performed on bilateral femora for local intraosseous perfusion before and after LPS injection. Blood samples were collected for haematological examinations before and after LPS injection. Bilateral femora were dissected and decalcified for microCT-based microangiography. ON lesion, intravascular thrombus and extravascular marrow fat cell size were examined histopathologically. RESULTS: Intravascular thrombus was observed in all ON rabbits. Extravascular marrow fat cell size was significantly increased in ON rabbits than that of the controls (P < 0.05). Compared to baseline, a significant decrease in ratio of tissue-type-plasminogen-activator/plasminogen-activator inhibitor 1, activated-partial- thromboplatin-time, and a significant increase in ratio of low-density-lipoprotein/high-density-lipoprotein were only found in ON rabbits (P < 0.05). Dynamic MRI showed a significant decrease in the perfusion index 'maximum enhancement' in the ON rabbits (P < 0.05) and microCT-based microangiography showed blocked stem vessels in ON samples. Overall, 93% of the rabbits (13/14) developed ON and no rabbits died throughout the experiment period. CONCLUSION: Both intra- and extravascular events were found attributing to the steroid-associated ON based on our experimental protocol with a single low-dose LPS injection and subsequent three injections of high-dose MPS. Both high ON incidence and no mortality in rabbits treated with this inductive protocol suggested its effectiveness for future studies on evaluation of therapeutic efficacy of interventions developed for prevention of steroid-associated ON.