肾病综合征患者血浆中t-PA/PAI-1的变化及糖皮质激素治疗的影响

目的:探讨肾病综合征患者糖皮质激素治疗前后不同阶段组织型纤溶酶原激活物(t-PA)与纤溶酶原激活物抑制物-1(PA1-1)的变化.方法:分为健康对照组、肾病综合征组,采用酶联免疫吸附(ELISA)方法检测血浆中t-PA和PAI-1水平的变化.结果:t-PA的血浆水平在各组之间无统计学差异(P>0.05);PAI-1的水平在肾病综合征组较正常明显升高(P<0.05),激素治疗1周后进一步升高(P<0.05),治疗4周后比1周组有显著下降(P<0.05),但较正常仍高(P<0.05).结论:t-PA/PAI-1平衡的紊乱,可能参与了肾病综合征的损伤机制,经糖皮质激素治疗后高凝状态短期内无明显改善.     

采用激素联合内毒素建立兔中期股骨头坏死模型

目的探索激素联合内毒素建立兔股骨头坏死(ONFH)国际骨循环研究会(ARCO)中期(Ⅱb~Ⅲb期)模型的最佳时间,为进一步研究治疗中期ONFH提供动物模型。方法将周龄为28周的健康成年新西兰白兔36只雌雄各半,用单纯随机抽样法分为实验组(n=30),对照组(n=6)。实验组予耳缘静脉注射1次低剂量的脂多糖(LPS)后,以24 h为间隔给予肌肉注射3次高剂量的甲基强的松龙(MPS)。对照组与实验组相同时间点注射等量生理盐水。于注射前及注射后4~9周行MRI检查,观察股骨头影像学改变,并通过病理切片明确ONFH改变。根据MRI检查结果,算出各周ARCO中期ONFH的发生率,卡方检验分析统计学差异。结果实验组动物注射MPS与LPS后出现3只动物死亡,B组则无死亡。注射后实验组股骨头ARCO中期坏死率:第4周为22.2%,第5周为57.4%,第6周为75.9%,第7周68.5%,第8周42.6%,第9周20.3%。对照组股骨头MRI信号未见异常。实验组第6周的ARCO中期ONFH率与第4、8、9周比较,差异均有统计学意义(P0.01);出现ARCO中期ONFH的股骨头在光镜下表现为:空骨陷窝,骨小梁塌陷断裂,微血管的分布密度减少。结论用MPS联合LPS造模的ONFH发生率高,死亡率较低,是较好的非创伤性模型。本实验中ARCO中期发生率在第5~7周期间最高。     

PAI-1基因4G/5G多态性与股骨头坏死易感性关联的Meta分析

目的研究纤溶酶原激活物抑制物-1（plasminogen activator inhibitor-1,PAI-1）基因4G/5G多态性与股骨头缺血性坏死易感性的关联。方法 Meta分析采用Stata 12软件进行,相对风险率的计算（相对比,RR）和95%可信区间（95%CI）。股骨头缺血性坏死969例,健康对照者419例,Meta分析结果表明：PAI-1基因4G/5G多态性与缺血性股骨头坏死的增加有关（等位基因模型：RR=1.24,95%CI=1.16-1.33;显性遗传模型：RR=1.12,95%CI=1.05-1.18）。根据不同的种族分析发现,高加索人群与亚洲人群比较,PAI-1基因4G/5G多态性与股骨头缺血性坏死易感性的关联性增加显著（P＆amp;gt;0.05）。结论 PAI-1基因4G/5G多态性与股骨头缺血性坏死的易感性增加有关,为股骨头坏死的早期诊断及以后提供理论依据。     

激素性股骨头骨坏死发病机制的实验研究

目的通过建立动物实验模型,采用血清学、放免、病理组织学、透射电镜等方法,探讨PGE2和TXB2在激素性股骨头骨坏死中的意义.方法选用健康成年中国白兔40只,雌雄不限,体重(2.5±0.5)kg,随机等分两组.实验组,臀部肌注甲基强的松龙琥珀酸钠,每次8ml/kg,每周2次.对照组以等量生理盐水替代激素.各组于处理前和处理后2、4、6、8周,测定血脂、血浆前列腺素E2(PGE2)和血栓烷素B2(TXB2)含量.并于第4、8、10、周分批处死动物,取肝脏和股骨头作常规病理组织学和超微结构观察.结果(1)实验组血浆总胆固醇、甘油三酯、极低密度脂蛋白较对照组增高,差异有显著性.(2)从实验第4周开始实验组PGE2较对照组明显增高,差异非常显著(P＜0.01).(3)实验第4周,TXB2较对照组明显增高,经秩和检验差异有显著意义(P＜0.05).(4)实验组股骨头骨组织内早期见髓腔脂肪细胞增多且细胞肥大,造血细胞减少,周围可见大量炎性细胞.电镜下见核膜破坏,染色质碎裂,晚期见空泡骨陷窝形成.结论糖皮质激素所致的股骨头坏死是多种因素引起股骨头血液循环障碍所致.     

生物力学因素在激素性股骨头坏死中对软骨细胞的作用

目的建立大鼠激素性股骨头坏死正常负重和超负重模型,探究力学因素在激素性股骨头坏死中对于软骨细胞的作用。方法4月龄健康Wistar大鼠随机分成实验组与对照组。臀肌注射地塞米松磷酸钠(20mg/kg),每周1次,共计8w。实验组,置于1km/h跑步机中,强迫动物跑动,形成股骨头坏死超负重模型;对照组,生理状态下正常负重。分别于2、4、6、8w处死大鼠,取右侧股骨头标本,行免疫组织化学法Bcl-2和MMP-1染色,比较不同组之间的累计光密度(IOD)值。取左侧股骨头行组织病理学检查。结果实验组Bcl-2和MMP-1累计光密度(IOD)与对照组比较,第4周起有明显统计学差异(P0.05)。实验组Bcl-2和MMP-1表达随激素注射时间增加有统计学意义(P0.05),q检验显示实验组Bcl-2和MMP-1表达从4w开始两两之间有统计学意义,8w时表达最多(P0.05)。实验组8w时病理学呈股骨头坏死表现。结论力学因素通过影响Bcl-2和MMP-1的生成,调节着软骨细胞的凋亡,促进软骨组织的改建。     

股骨头坏死动物模型研究进展

股骨头坏死(ONFH)研究取决于高质量ONFH动物模型。应用创伤或非创伤方法制作的ONFH模型至今仍未达到与人类ONFH病理改变相似、能逐渐进展至股骨头塌陷的模型要求。目前报道的造模方法多采用多种方法联合,尤其是激素联合脂多糖或异体血清。两足大型动物鸸鹋ONFH模型成功出现关节面塌陷,其较大体型适合对不同保头手术效果进行比较,逐渐受到重视。     

应用低温手术建立三足犬股骨头坏死动物模型

目的 建立一种能模拟人股骨头坏死(osteoneerosis of the femoral head,ONFH)病理生理过程的实验动物模型. 方法 取健康成年雄性Beagle犬10只,体重(16.0 4±1.6)kg,建立三足犬模型后,于0.5 MPa加压液氮作用下冷冻16.5 min,复温10min至0℃,再冷冻16.5 rain建立ONFH动物模型.其中实验组9只,对照组假冷冻组,打隧道但并不冷冻).术中监测股骨头表面温度,术后6个月处死实验组9只犬进行脱钙骨组织学检查,对照组1只犬观察至24个月. 结果 术中监测第一次边界温度为(-27.9 4±4.3)℃,第二次边界温度为(-31.3 4±4.7)℃,第二次冷冻后股骨头表面温度较第一次平均下降3.4℃,差异有统计学意义(P<0.01).股骨头直径(17.7 4±1.1)mm,经线性回归分析,股骨头直径越小,股骨头表面温度下降越低,回归方程为y=-2.6-2.409x-(P<0.05).将股骨头直径与第一次边界温度进行Pearson相关性分析,成线性相关(r=-0.977,P<0.05).病理组织学检查示术后6个月实验组4只犬出现股骨头塌陷,塌陷率44.4%;对照组无异常改变. 结论 低温手术可建立进展至股骨头塌陷的三足犬ONFH模型.     

乳腺癌凝血纤溶指标与生物学行为关系的研究

目的 研究乳腺癌凝血纤溶异常与其转移、激素受体状态、分期等生物学行为的密切关系.方法 检测30例乳腺癌血浆6项凝血和纤溶指标,统计分析其变化与淋巴结转移、细胞增殖、受体状态等的关系.结果 良性对照组与乳腺癌组FiB由(3.05±0.44)g/L增高至(3.39±0.52)g/L(P＜0.05)、D-Di由(0.27±0.06)μg/ml增高至(0.36±0.16)μg/ml(P＜0.05)、PAI-1由(26.14±3.30)ng/ml增高至(34.59±3.68)ng/ml(P＜0.01),差异均有统计学意义.腋窝淋巴结转移阳性者的血浆FiB(3.70±0.47)g/L和PAI-1(37.36±2.71)ng/ml较无转移者增多(P＜0.05);细胞增殖抗原Ki67表达≥30%者PAI-1含量增加(36.40±3.57)ng/ml(P＜0.05).t-PA含量在不同激素受体状态下的差别有统计学意义(P＜0.05).不同TNM病理分期乳腺癌患者之间APTT(P＜0.01)、FiB(P＜0.01)、D-Di(P＜0.05)和PAI(P＜0.05)差异有统计学意义.结论 凝血纤溶指标与乳腺癌增殖、转移、分期等生物学行为有密切关系,可作为判断预后和术后复发转移的监测指标,并有利于血栓前状态的判断.     

赛百粉针对难治性肾病综合征凝血/纤溶系统的影响

目的:探讨赛百粉针对难治性肾病综合征(RNS)凝血/纤溶系统的影响.方法:30例RNS随机分为治疗组与对照组,在常规治疗基础上,治疗组加用纤溶酶制剂赛百粉针,对照组加用低分子肝素钙速碧林注射液,比较两组治疗前后24 h尿蛋白、组织型纤溶酶原激活物(t-PA)、纤溶酶原激活物抑制物1(PAI-1)活性及血浆纤维蛋白原(Fib),D-二聚体(D-D)、纤溶酶原(PLG)含量、部分凝血活酶时间(APTT)、凝血酶原时间(PT)的变化.结果:治疗1个疗程后,治疗组与对照组尿蛋白均显著下降(P≤0.001),两组间治疗前、后比较均无差异(P＞0.05);t-PA、PLG治疗前治疗组与对照组均较健康组明显下降(P＜0.005),PAI-1、D-D、Fib明显升高(P≤0.000),PT、APTT显著缩短(P≤0.000),两组间比较均无差异(P＞0.05).t-PA、PLG治疗后治疗组与对照组均较治疗前显著升高(P＜0.05),PAI-1、Fib、D-D显著下降(P≤0.005),除t-PA对照组升高更显著(P=0.018),PAI-1两组比较无统计学差异外(P＞0.05),D-D、Fib、PLG治疗组变化较对照组更显著(P≤0.005).治疗后两组PT延长均无统计学意义(P＞0.05),APTT对照组显著延长(P=0.021),治疗组无显著延长(P=0.155).结论:赛百粉针能显著改善RNS凝血/纤溶异常,减少蛋白尿,有助于RNS的缓解.     

Multiple bioimaging modalities in evaluation of an experimental osteonecrosis induced by a combination of lipopolysaccharide and methylprednisolone

AIMS: 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 lipopolysaccharide (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). 24 h later, three injections of 20 mg/kg of MPS were given intramuscularly at a time interval of 24 h. 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 hematological 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.     

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一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）。24h后，每隔24h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一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。     

甲强龙联合内毒素诱导犬股骨头坏死的实验研究

目的建立使用1次低剂量内毒素联合连续3次高剂量甲强龙诱导杂种犬股骨头坏死模型,并观察所造成股骨头坏死的特点。方法动物模型组采用12个月龄中国健康杂种犬12只,先予犬静脉注射内毒素1次（9.0μg/kg）,24 h后连续3 d每天行甲强龙肌内注射（18.0 mg/kg）。空白对照组4只,腹腔内注射等量生理盐水。末次注射2、4、6周后2组均行双髋关节数字化X线（DR）、磁共振成像（MRI）及病理组织学检查（Masson染色）。结果末次注射2周后动物模型组所有检查均未发现股骨头坏死的征象,4周后动物模型组Masson染色出现早期骨坏死表现,6周后动物模型组MRI及Masson染色出现较明显的早期股骨头坏死表现。结论采用1次低剂量内毒素联合连续3次高剂量甲强龙注射可造成杂种犬股骨头坏死模型。     

辛伐他汀联合BMSCs治疗激素性股骨头坏死的实验研究

目的 单纯BMSCs治疗激素性股骨头坏死的效果尚不理想,探讨辛伐他汀联合BMSCs治疗激素性股骨头坏死的可行性. 方法 取24只新西兰大白兔骨髓分离培养BMSCs,取第2代细胞制备为1×107/mL,细胞悬液,与明胶海绵复合.取70只新西兰大白兔经耳缘静脉注射10.μg/kg脂多糖,24 h后臀肌注射20 mg/kg甲基强的松龙琥珀酸钠,共3次,每次间隔24 h,制备股骨头坏死模型.选取制备成功的48只,随机分为4组,每组12只.A组不作任何处理B组单纯减压,并于减压通道植入明胶海绵;C组减压同时植入复合BMSCs的明胶海绵;D组减压同时植入复合BMSCs的明胶海绵,每日灌服辛伐他汀(10 mg/kg)至处死.观察动物一般情况,于术后4、8周各组取6只行MRI扫描后,处死取股骨头行组织学及免疫组织化学染色,扫描电镜观察. 结果 术后8周C、D组动物走路姿势逐步好转.术后4周各组MRI未见明显坏死区信号改变,8周时A组可见坏死低信号区明显扩大,B组无变化,C组范围缩小,D组明显缩小.组织病理学观察,术后4周A组见空骨陷窝,无新生毛细血管;B组空骨陷窝较多,有少量新生毛细血管;C、D组空骨陷窝减少,坏死区有大量增生活跃的成骨细胞及新生毛细血管.D组空骨陷窝阳性数及微血管密度分别为19.30±1.52和7.08±1.09,与其他组比较差异有统计学意义(P＜0.05).术后8周,A组可见部分骨小梁断裂;B组减压孔道有纤维性骨痂形成;C组空骨陷窝少见,髓腔形态不规则;D组大量新生骨形成,髓腔较规则,髓内脂肪细胞大小及分布均匀.D组空骨陷窝阳性数及微血管密度分别为11.31±1.28和12.37±1.32,与其他组比较差异有统计学意义(P＜0.05),与术后4周比较差异有统计学意义(P＜0.05).扫描电镜观察术后8周,A组骨小梁见多处断裂塌陷,骨小梁表面未见骨细胞,髓腔内有大量脂肪细胞堆积;B组部分骨小梁可见裂痕;C组骨小梁不致密;D组骨小梁结构规整致密,成骨细胞多,骨细胞及基质胶原纤维正常,髓腔规则. 结论 辛伐他汀能促进BMSCs成骨及成血管化,辛伐他汀联合BMSCs治疗激素性股骨头坏死具有较好的应用前景.     

激素性股骨头坏死与凝溶紊乱的实验研究

目的：观察激素性股骨头坏死家兔模型的血液凝溶功能变化,探讨激素性股骨头坏死的发病机理。方法：健康雄性成年家兔36只,随机分为对照组（注射生理盐水）,实验组（冲击注射地塞米松）。检测血脂、血浆组织纤溶酶原激活物（tPA)、纤溶酶原激活物抑制物（PAI）、血栓素A2（TXA2）、前列环素（PGI）的阶段性变化。观察实验35、70d双侧股骨头的病理变化。结果：实验组15d甘油三脂、总胆固醇、TXA2／PGI比值均显著高于对照组。实验30、60d,实验组tPA／PAI比值显著低于对照组。实验组35、70d双侧股骨头软骨下区骨陷窝空虚率均较对照组显著增高。结论：地塞米松冲击应用可引起家兔血液高凝、低纤溶。血液凝溶功能紊乱可能参与兔激素性股骨头坏死的发病。     

Blocking of tumor necrosis factor activity promotes natural repair of osteochondral defects in rabbit knee

Background and purpose Osteochondral defects have a limited capacity for repair. We therefore investigated the effects of tumor necrosis factor (TNF) signal blockade by etanercept (human recombinant soluble TNF receptor) on the repair of osteochondral defects in rabbit knees.Material and methods Osteochondral defects (5 mm in diameter) were created in the femoral patellar groove in rabbits. Soon after the procedure, a first subcutaneous injection of etanercept was performed. This single injection or, alternatively, 4 injections in total (twice a week for 2 weeks) were given. Each of these 2 groups was divided further into 3 subgroups: a low-dose group (0.05 μg/kg), an intermediate-dose group (0.4 μ g/kg), and a high-dose group (1.6 μ g /kg) with 19 rabbits in each. As a control, 19 rabbits were injected with water alone. The rabbits in each subgroup were killed 4 weeks (6 rabbits), 8 weeks (6 rabbits), or 24 weeks (7 rabbits) after surgery and repair was assessed histologically.Results Histological examination revealed that the natural process of repair of the osteochondral defects was promoted by 4 subcutaneous injections of intermediate-dose etanercept and by 1 or 4 injections of high-dose etanercept at the various time points examined postoperatively (4, 8, and 24 weeks). Western blot showed that rabbit TNFα had a high affinity for etanercept.Interpretation Blocking of TNF by etanercept enabled repair of osteochondral defects in rabbit knee. Anti-TNF therapy could be a strategy for the use of tissue engineering for bone and cartilage repair.     

Src siRNA prevents corticosteroid-associated osteoporosis in a rabbit model

In an established steroid-associated osteonecrosis (SAON) rabbit model we found recently that blockage Src by siRNA could improve reconstructive repair of osteonecrosis via enhancing osteogenesis and inhibiting bone resorption. The current study investigated if blocking Src was able to prevent steroid-associated osteoporosis (SAOP) in the same SAON animal model. Rabbits were treated with pulsed lipopolysaccharide (LPS) and corticosteroid methylprednisolone (MPS). At 2, 4, and 6 weeks after induction, Src siRNA, control siRNA and saline were intramedullary injected into proximal femur, respectively. Two fluorescent dyes xylenol orange and calcein green were injected before sacrificing the animals for in vivo labeling of the newly formed bone. At week 6 after induction, proximal femora of rabbits were dissected for micro-CT and histological analysis. Results showed significant bone loss in the metaphysis of femoral head in the control rabbits after SAON induction. Src siRNA treatment was able to prevent steroid-associate bone loss in trabecular bone and increase cortical bone thickness at femoral neck. Histomorphometry showed that Src siRNA increased the osteoblastic bone formation and decreased the eroded bone surfaces suggesting decreased osteoclastic bone resorption. This was the first study to report bone loss after SAON induction in rabbit model that could be prevented by knocking down Src by siRNA.     

Functional perfusion MRI predicts later occurrence of steroid-associated osteonecrosis: An experimental study in rabbits

Ischemia is the defined pathway leading to steroid-associated osteonecrosis (ON). Early detection of ischemic condition may help predict later ON occurrence. Bone marrow perfusion function evaluation by perfusion magnetic resonance imaging (MRI) may be a unique modality for this application. Twenty-five adult male New Zealand white rabbits were used in this study. Lipopolysaccharide (LPS) and methylprednisolone (MPS) were administrated for ON induction based on a published protocol. T1-weighted and fat suppression T2-weighted MR imaging (conventional MRI) were performed for ON lesion detection based on the abnormal signal in the proximal femora at week 0 as the baseline (before LPS injection), and week 1 and week 2 after MPS injection. At the same time, the blood perfusion function in the proximal femora was measured by perfusion MRI. Maximum enhancement (ME)—an index of MRI perfusion function was analyzed. After MRI scanning, the proximal femora were prepared histopathologically for ON lesion analysis. The rabbit with bilateral histopathological ON lesions was defined as an ON+ rabbit and included in the ON+ group evaluated at week 1 and week 2, respectively, and the rabbit without ON lesions in bilateral femora was classified into the ON− group. For the underlying mechanism of perfusion change, the extravascular marrow fat cells were measured and the intravascular endothelium inflammation injury indicator of tissue factor (TF) expression and thrombus formation were detected. In ON+ group, ME in perfusion MRI showed a significant decrease at week 1 and week 2 as compared with the baseline (p < 0.01). There was a more than 50% decrease in ME at week 1 in ON+ group; whereas there were no detectable ON lesions by conventional MRI at week 1, though 93% (14/15) rabbits could be detected at week 2 in ON+ group. In ON− group, ME showed a slight decrease at week 1 (less than 30%), and nearly recovered to normal at week 2 as compared with the baseline. Histological results showed a much larger average marrow fat area and more severe marrow blood sinusoids compression from surrounding crowded fat cells, and stronger positive TF expression in marrow endothelium and more thrombus formation in ON+ rabbits than ON− rabbits. This study demonstrated that functional perfusion MRI could predict development of steroid-associated ON. Our experimental data suggested that perfusion MRI might be a sensitive noninvasive modality for monitoring steroid-associated ON in patients. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 742–747, 2009     

Osteonecrosis induced by a single administration of low-dose lipopolysaccharide in rabbits

We succeeded in developing a novel rabbit model of nonsteroid and nontraumatic osteonecrosis (ON) by use of a single- and low-dose lipopolysaccharide (LPS) injection. This model is simple and highly reproducible for the frequent development of multifocal and widespread ON lesions. Male adult Japanese white rabbits intravenously injected with a single injection of 10 μg/kg body weight of LPS were histopathologically examined in the early phase (3 [n = 3], 5 [n = 3], and 24 h [n = 3]) and at 4 weeks (n = 22). Seventy-seven percent of the rabbits developed multifocal ON 4 weeks after LPS injection. ON was also observed in the femoral and humeral condyle. The average percentage of necrotic area/total area examined was 86.7 ± 29.1% and 78.8 ± 16.7% in the proximal one third of both the femoral and humeral bones, respectively. Organized thrombi in the intraosseous small-sized arteries and arterioles were frequently seen in and around the necrotic tissues. In the early phase, LPS treatment prominently induced thrombocytopenia, hyperlipidemia, and increased plasma levels of plasminogen activator inhibitor-1 (PAI-1). The plasma level of PAI-1 was significantly higher in the rabbits with ON than in those without ON (p < 0.01). The immunohistochemical expression of tissue factor was exaggerated in monocytes/macrophages and adipocytes in both the femoral and humeral bones of the LPS-treated rabbits. Histologically, marrow necrosis and fibrin thrombi could be observed at 24 h. In addition, pretreatment with an anticoagulant, warfarin potassium, significantly decreased the incidence of LPS-induced ON (33%, N = 9, p < 0.05) associated with elongation of prothrombin time. The results of our study show that a single administration of low-dose lipopolysaccharide induces multifocal and widespread ON characterized by the pathophysiological participation of hypercoagulability in ON development. Therefore, this model would be useful for elucidating the pathogenesis of nonsteroid ON in humans especially inflammatory hypercoagulability-induced as well as for developing preventive and therapeutic strategies.     

Treatment of experimental osteonecrosis of the hip in adult rabbits with a single local injection of recombinant human FGF-2 microspheres

Basic fibroblast growth factor (FGF-2) exerts anabolic actions on bone formation. Here we investigated the potential effects of recombinant human FGF-2 (rhFGF-2) on the repair process of osteonecrosis of the femoral head (ONFH) and the development of secondary osteoarthritis (OA) in adult rabbits. ONFH was induced by intramuscular injection with methylprednisolone, and vascular occlusion of the capital femoral epiphysis by electrocoagulation, in adult Japanese white rabbits. Animals were randomized into two groups: treatment and control. The treatment group was given a single local injection into the femoral head of 100 μg rhFGF-2 in 100 μl gelatin hydrogel microspheres 8 weeks after the ONFH procedure, and the control group was given phosphate-buffered saline in 100 μl gelatin hydrogel microspheres. Morphological, histopathological, and radiologic analyses, including micro-computed tomography scans and magnetic resonance imaging, showed collapse of the femoral head and progression of articular cartilage degeneration in the control group at 16 weeks after the single local injection of rhFGF-2. In contrast, rhFGF-2 treatment resulted in new bone formation in the femoral head and prevented the femoral head from collapsing. In addition, the changes in OA, assessed by the modified Mankin score, was significantly lower in the treatment group. Our results indicate that a single local injection of rhFGF-2 microspheres promoted the repair of the osteonecrotic femoral head and inhibited femoral head collapse and OA progression. rhFGF-2 may be a promising strategy for the treatment of ONFH.