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1、MR定量监测经髓核抽吸法建立兔腰椎间盘退变模型的研究【基金项目】重庆市自然科学基金（cstc2011jjA10082）；国家临床重点专科建设项目【第一作者】谢光友，医学硕士，E-mail：【通讯作者】杨海涛，医学博士，E-mail：吕富荣，教授，硕士生导师， E-mail：谢光友1 杨海涛2* 吕富荣2* 刘昌杰1 杨明放1 邓奇平11.贵州省人民医院放射科，贵阳 5500022.重庆医科大学附属第一医院放射科，重庆 400016【摘要】 目的 构建兔腰椎间盘退变模型并提高其退变机制的认识。方法 18只新西兰大白兔，从脊柱侧后方手术入路暴露腰椎间盘，18G穿刺针抽吸12只L2/3、L3/4、L
2、4/5髓核，各约5mg，正常L1/2、L5/6和另6只L2-5间椎间盘分别为组内和组间对照。术后第4、8、12周行MR 矢状位T2WI和T2-mapping序列检查，取正中矢状面观察各椎间盘的信号并测量T2弛豫时间并与正常组比较。相应各时间点取2只L1-6间椎间盘行HE和Masson染色，观察纤维环形态、髓核细胞及基质的变化并与正常组比较。结果 18G穿刺针经脊柱旁路于横突根部可成功穿刺抽吸髓核。正常椎间盘T2WI序列为均匀高信号，实验组随时间延长信号逐渐降低，第8周明显降低，第12周基本完全呈低信号；第8周实验组与正常组T2弛豫时间差异有统计学意义（p0.05）。术后随时间进展，HE和Mas
3、son染色示椎间盘细胞和胶原逐渐减少，纤维环和髓核分界不清；第8周髓核基质退变纤维化，几乎被纤维组织取代；第12周椎间盘纤维软骨化，局部形成软骨。结论 髓核针刺抽吸法可成功建立兔腰椎间盘退变模型，MR可实时定量监测退变程度，为椎间盘退变机制的动物实验研究提供可靠的模型及影像学监测方法。【关键词】磁共振成像；定量；髓核抽吸；椎间盘；退变模型【中图法分类号】R332；R445.2 【文献标志码】AThe study of MR quantitatively monitoring the model of lumbar intervertebral disc degeneration by nucl
4、eus pulposus suction Xie Guangyou1，Yang Haitao2*, Lyu Furong2*, Liu Changjie1, Yang Mingfang1，Deng Qiping11.Department of Radiology,the GuizhouProvincialPeoplesHospital,Guiyang,550002,Guizhou,China2.Department of Radiology,the First Affiliated Hospital of Chongqing Medical University,Chongqing,40001
5、6,China【Abstract】 Objective To build a rabbit model of lumbar intervertebral disc degeneration and improve the recognition of its mechanism. Methods The nucleus pulposus of L2/3,L3/4,L4/5 discs of 12 rabbits were aspirated by using a 18G needle and the normal L1/2, L5/6 discs of each rabbit and the
6、discs between L2-5 vertebral body of 6 rabbits acted as the normal intra and inter control groups, respectively. The rabbits were performed with sagittal MR T2WI and T2mapping sequences scanning at the pre-and-post-operative 4th, 8th, 12th week, respectively. The signal intensity of each disc was ev
7、aluated and T2 relaxation time was measured carefully.The discs between L1-6 were dissected and then performed with HE and Masson trichrome staining to evaluate the form of discs,content change of nucleus pulposus cells and matrix between normal and experimental groups. Results The nucleus pulposus
8、could be punctured successfully with a 18G needle in the root of transverse process via spinal bypass.The normal discs manifested with uniform high signal in T2WI ,and the signal intensity of experimental group reduced gradually after operation.The signal intensity decreased sharply at the 8th week
9、and became completely low signal intensity at the 12th week post-operation.The significant difference of T2 relaxation time was found between normal and experimental at the 8th week groups(p 0.05).The results of HE and Masson staining showed that the cells and matrix decreased gradually,moreover,the
10、 boundary between annulus fibrosus and nucleus pulposus became obscure.The nucleus pulposus displayed the process of fibrosis and was almost replaced by fibrous tissue at 8th week.The discs was fibrocartilage and local cartilage can be observed at the 12th week. Conclusion The rabbit model of lumbar
11、 intervertebral disc degeneration can be established successfully by nucleus pulposus suction,and it can be dynamically monitored and quantitatively analyzed by MRI.The study provides a reliable experimental method to improve the recognition of the mechanism of intervertebral disc degeneration.【Key
12、words】Magnetic resonance imaging；Quantification；Intervertebral disc；Nucleus pulposus suction；Degenerative model腰椎间盘退行性疾病（intervertebral disc degeneration disease,IVDDD）是导致腰背部疼痛最常见的原因，为深入探讨椎间盘退变的病理及病理生理机制，建立一种操作简便、重复性和稳定性好的椎间盘退变模型，对指导治疗椎间盘退变均有重要价值。本研究先观察兔腰椎间盘与周围结构的解剖关系，然后经脊柱旁路针刺抽吸髓核组织，成功制作兔腰椎间盘退变模型，通
13、过MR动态定量监测并结合病理观察，综合评价该法建立椎间盘退变模型的特点。1 材料和方法1.1实验动物取3月龄健康成年新西兰大白兔18只，平均体重2.5kg，雌雄不拘，无脊柱畸形及椎间盘病变，肢体活动良好，分笼饲养，由重庆医科大学动物实验中心提供（许可证号：SCK(渝)2012-0001）。主要试剂仪器：HE和Masson染色试剂盒、15%EDTA脱钙液、无菌PBS( 武汉博士德公司)，戊巴比妥钠，青霉素、庆大霉素。T2000U倒置相差显微镜（日本Nikon公司），GE HD-X signa 3.0T MRI成像系统、N-V Array8通道相控阵线圈、adw 4.4工作站（美国GE公司），无菌
14、手术包，18G穿刺针。1.2 构建腰椎间盘退变模型18只兔行MRI检查，术前一天肌注青霉素20万U/kg,术前6h禁饮禁食。1%戊巴比妥钠（1ml/kg）经耳缘静脉注射成功麻醉后,取俯卧位固定于兔台，以L2-5间椎体为中心，剃毛备皮、术野消毒铺巾，以L3椎体左侧横突根部为中心，做一4-5cm纵切口，逐层切开皮肤、皮下组织、胸腰筋膜，神经剥离子沿横突向背侧剥离直至横突根部，充分暴露L3/4椎间盘并用18G穿刺针抽吸髓核组织，然后缓慢退针，同法处理L2/3、L4/5椎间盘，术中用庆大霉素生理盐水冲洗、止血。术后逐层缝合，回笼标准条件下饲养，连续一周消毒切口并肌注青霉素20万U/kg。1.3 MR动
15、态定量监测术前和术后第4、8、12周对实验组和正常组行腰椎MR检查。1%戊巴比妥钠（1ml/kg）经耳缘静脉注射成功麻醉后，取俯卧位并拉直脊柱后固定，以L3椎体为中心行MR矢状位检查。扫描序列及参数：FSE T2WI序列：TR2620 ms，TE102 ms，FOV20cm，层厚3 mm，层间距0.6 mm，NEX4，矩阵384256，T2maping序列：TR1600 ms，TE11.1、22.2、33.3、44.4、55.5、66.6、77.7、88ms，FOV 20cm，层厚3mm，层间距0.6 mm，NEX1，矩阵384256。MR资料由两名骨关节疾病影像诊断医师进行双盲评估，选腰椎正
16、中矢状面，观察T2WI序列上各组L1-6间椎间盘的信号及形态，参照Pfirrmann分级标准1。通过adw 4.4工作站Functool软件的时间自动测量系统，设置一椭圆形兴趣区包绕椎间盘，测量T2-mapping序列上相应椎间盘的T2弛豫时间，每个兴趣区重复测量3次，取均值。L2/3、L3/4、L4/5椎间盘为实验组，以组内L1/2、L5/6及正常兔L1-6间椎间盘为对照。1.4 病理染色观察分别在第4、8、12周用空气栓塞法处死实验组及正常组兔2只，迅速游离并取出L1-6间椎间盘，生理盐水反复冲洗，10%多聚甲醛固定24h，15% EDTA脱钙两周后梯度酒精脱水，二甲苯透明处理、石蜡包埋、5m切片，分别行HE和Masson染色，镜下观察椎间盘形态、髓核细胞及胶原的变化，L2/3、L3/4、L4
