一种新的兔脊髓型颈椎病动物模型的建立
发布时间：2018-07-15 19:27
【摘要】：目的:本研究的目的是尝试研制一种新的脊髓压迫钉用以建立一种带有典型脊髓型颈椎病(cervical spondylotic myelopathy,CSM)临床特征的兔实验模型,这种动物模型符合CSM神经功能变化及病理变化,并可用于X线、CT、MRI等多种CSM临床检查方法及基础研究。 材料与方法: 1材料 新西兰家兔,30只,雄性。由河北医科大学动物实验中心提供,普通级;核磁机由德国Siemens公司生产(Siemens Avanto MR);CT机由德国Siemens公司生产(SOMATOM Sensation 16 CT);X线机由法国Trophy公司生产( KODA2200 );肌电图机由美国Nicolet公司(VikingQuest);聚甲基丙烯酸甲酯(PMMA)、牙胶尖和丁香油均购自上海医疗器械股份有限公司齿科材料厂。氧化锌购自南昌白云药业有限公司。 2方法 (1)应用聚甲基丙烯酸甲酯(PMMA)、牙胶尖、氧化锌和丁香油制备显影树脂脊髓压迫钉。 (2)动物饲养与分组:给予普通实验室饮食喂养,饲养环境温度为16-20℃。术前饲养1周。30只实验兔随机分为实验组(20只)和对照组(10只)。实验组再分为3个月组(10只)和6个月组(10只)。 (3)动物模型制作:首先腹腔注射地西泮0.5ml,之后腹腔注射速眠新Ⅱ(0.1ml/kg)全身麻醉,备皮。取气管右侧旁开0.5cm,甲状软骨下1cm,行纵行切口长约5cm,逐层分离至椎前,确定C3椎体,于C3椎体前开孔直径约3×3mm,刮出松质骨至椎管前壁骨皮质,将准备好的显影树脂脊髓压迫钉轻轻置入孔内,将肌肉复位逐层缝合。 (4)神经功能观察:分别于术前、术后第一天、术后3个月和术后6个月采用改良Tarlov’s运动功能评分法进行运动功能评估,并记录。 (5)皮层体感诱发电位(CSEP)检查:与术前行双下肢CSEP检查,观察图形振幅并记录潜伏期P15,术后立即行CSEP检测,术后3个月和6个月时测肌电图并比较振幅以及潜伏期变化。 (6) X线检查:术后1个月行X线检查,以确定本模型可于X线下清晰显影,并确定压迫钉所在位置,压迫钉与兔颈椎相关关系。 (7) CT检查:术后1个月行CT检查,以确定本模型可行CT检查,在矢状位确定压迫钉位置,压迫钉与颈椎关系;在轴位观察压迫钉突入椎管深度,并测量椎管横径、矢状径以及压迫钉前端至椎管后壁距离(椎管最短经),并计算椎管平均占用率(椎管占用率=(椎管前后径-椎管最短径)/椎管前后径×100%)。 (8) MRI检查:术后3个月和6个月分别进行MRI平扫,观察脊髓形态,于T2加权像轴位观察脊髓受压情况,并测量脊髓前后径、脊髓横径计算平均脊髓压迫率(脊髓压迫率=椎管矢状径/横径×100%)。于T2加权像矢状面脊髓受压信号增强最明显部位或脊髓受压最严重部位选取兴趣区(面积为0.10 cm2)测量信号强度值,再于C6/7椎体脊髓节段取兴趣区(面积为0.10cm2)测量信号强度值,计算并进行比值;对照组则选取C3椎体节段脊髓和C6/7节段脊髓测量信号强度值,计算比值。 (9)光镜下观察:分别于3个月及6个月处死动物,取脊髓受压阶段脊髓,HE染色及DAB染色观察细胞变化及脊髓微血管变化。 3统计分析使用统计学软件SPSS13.0处理数据进行统计学分析。 (1)计算CSEP平均潜伏期、CT下平均椎管占用率、MRI下平均脊髓压迫率。 (2) MRI信号比、Tarlov’s评分、CSEP潜伏期采用中位数(P50)和四分位数间距(P75? P25)进行描述,Kruskal-Wallis H检验分析组间差异,采用调整α水准(α’=0.0167)后的两样本比较Wilcoxon秩和检验进行组间两两比较。 结果: 1神经功能的变化实验组术后神经症状最初出现在3个月内,6个月时运动功能差异较正常大。对照组,所有兔神经功能均正常,无任何改变。 2 CSEP 实验组在行造模手术前双下肢CSEP平均潜伏期为16.21±1.7ms,实验组肌电图检查与对照组比较,体感诱发电位在波幅与潜伏期上出现显著变化,并随时间加重。对照组较术前无变化。 3 X线检查 X线检查可见显影树脂脊髓压迫钉位于C3椎体,突入椎管,显影清晰。 4 CT检查 CT显像矢状位及轴位像均可明显观察到显影压迫钉的所在位置。压迫钉突入椎管位置及大小可清晰观察。平均椎管占用率为40.2%。 5 MRI检查 MRI检查可在轴位像观察到脊髓压迫的存在,平均脊髓压迫率为43.9%。在T2加权像矢状位可观察到受压区域出现脊髓高信号,肉眼可观察到信号强度明显高于未受压区域及正常脊髓信号,且随时间逐渐增强。统计学结果显示,脊髓信号比:对照组,3个月实验组和6个月实验组三者总体有统计学差异,信号随时间逐渐增强。 6病理学检 压迫3个月后的兔脊髓灰质神经元数目减少,面积缩小,尼氏体浅染。白质有空洞形成,脱髓鞘改变。压迫6个月组,脊髓灰质前角被压扁,脊髓前角细胞消失或坏死,脊髓白质成明显的空泡样变,轴索变性,脱髓鞘样改变。DAB染色发现对照组血管形态正常,数目多。压迫3个月组发现脊髓腹侧受压处血管减少,灰质血管稍多。压迫6个月组受压处血管稀疏,灰质有少量血管分布,血管较正常形态较短且细。 结论: 本实验应用特制显影树脂脊髓压迫钉建立的脊髓型颈椎病动物模型具有典型脊髓型颈椎病临床特征。这种模型可同时用于CSEP、X线、CT、MRI及病理学检查等多种临床CSM相关检查,且模型制作简单,可重复性强,是结合临床综合研究脊髓型颈椎病的可靠模型。
[Abstract]:Objective: the purpose of this study was to try to develop a new spinal cord compression nail to establish a rabbit experimental model with typical cervical spondylotic myelopathy (CSM) clinical features. This animal model accords with the changes of CSM nerve function and pathological changes, and can be used in various CSM clinical examination methods such as X, CT, MRI and so on. And basic research.
Materials and methods:
1 material
New Zealand rabbit, 30, male. Provided by the animal experiment center of Hebei Medical University, general level; nuclear magnetic machine is produced by Siemens Avanto MR (German Siemens); the CT machine is produced by Siemens company of Germany (SOMATOM Sensation 16 CT); the X ray machine is produced by the French Trophy company (KODA2200); the electromyogram machine is made by Nicolet company (VikingQuest) of the United States. Methyl methacrylate (PMMA), gum cusp and lilac oil are purchased from Shanghai medical equipment Limited by Share Ltd dental materials factory. Zinc Oxide is purchased from Nanchang Baiyun Pharmaceutical Co., Ltd.
2 method
(1) polymethylmethacrylate (PMMA), gutta percha, Zinc Oxide and clove oil were used to prepare developing resin compression screws for spinal cord.
(2) animal feeding and grouping: feeding the ordinary laboratory and feeding the environment at 16-20. The experimental rabbits were randomly divided into the experimental group (20) and the control group (10) for 1 weeks before the operation for 1 weeks. The experimental group was divided into 3 months (10) and 6 months (10).
(3) animal model making: first intraperitoneal injection of diazepam 0.5ml, then intraperitoneal injection of fast dormant new II (0.1ml/kg) general anesthesia, skin preparation, taking the right side of the trachea on the right side of 0.5cm, subchondral 1cm, longitudinal incision length about 5cm, separating the layer by layer to the anterior vertebra, determining the C3 vertebral body, the diameter of the anterior opening of the C3 vertebral body about 3 x 3mm, scraping the cancellous bone to the anterior spinal canal wall of the vertebral canal. The prepared resin compression spinal cord is placed into the hole gently, and the muscle is reset and stitched by layer.
(4) observation of nerve function: before the operation, the first day after operation, 3 months after operation and 6 months after the operation, the modified Tarlov 's motor function scoring method was used to evaluate the motor function, and the records were recorded.
(5) the cortical somatosensory evoked potential (CSEP) examination: CSEP examination of the two lower limbs was performed before the operation, the amplitude of the graph was observed and the latency P15 was recorded. The CSEP was measured immediately after the operation. The EMG was measured at 3 and 6 months after the operation, and the amplitude and the latent period were compared.
(6) x - ray examination: the X - ray examination was performed 1 months after the operation to determine that the model could be clearly developed under X - ray, and the position of the compression nail was determined, and the correlation between the compression nail and the cervical spine of the rabbit was determined.
(7) CT examination: 1 months after the operation, CT examination was performed to determine the feasible CT examination of the model, to determine the position of the compression nail in the sagittal position, to press the relationship between the nail and the cervical spine; to observe the depth of the vertebral canal in the axial position, and to measure the transverse diameter of the spinal canal, the sagittal diameter and the distance between the front of the compression nail and the posterior wall of the spinal canal (the shortest canal of the vertebral canal), and calculate the average occupancy rate of vertebral canal (vertebrae). Tube occupancy rate = (anterior and posterior spinal canal diameter shortest) / vertebral canal anteroposterior diameter * 100%.
(8) MRI examination: the spinal cord compression was observed at 3 months and 6 months after the operation, and the spinal cord compression was observed on the T2 weighted image axis. The spinal cord compression rate (spinal cord compression ratio = sagittal diameter / transverse diameter * 100%) was measured with the transverse diameter of the spinal cord. The most obvious position in the T2 weighted sagittal spinal cord compression signal was enhanced. The intensity of signal intensity was measured in the most serious part of the spinal cord compression (area 0.10 cm2), and the intensity of signal intensity was measured in the area of C6/7 vertebral spinal cord segment (area 0.10cm2), and the ratio was calculated and carried out. In the control group, the signal intensity of the spinal cord and C6/7 segment of the C3 vertebral body was measured and the ratio was calculated.
(9) light microscope observation: animals were sacrificed at 3 months and 6 months respectively. Spinal cord compression stage was performed. HE staining and DAB staining were used to observe cell changes and changes in spinal cord microvasculature.
3 statistical analysis used statistical software SPSS13.0 to process data for statistical analysis.
(1) calculate the average latency of CSEP, the average spinal canal occupancy rate under CT, and the average spinal cord compression rate under MRI.
(2) the MRI signal ratio, the Tarlov 's score, the median of the CSEP latency (P50) and the four quantile spacing (P75? P25) were described. The difference between the groups was analyzed by the Kruskal-Wallis H test, and the two samples after the adjustment of alpha level (alpha' =0.0167) were compared with the Wilcoxon rank sum test to compare 22 of the groups.
Result:
1 nerve function changes in the experimental group first appeared in 3 months, and the motor function difference was larger than normal at 6 months. In the control group, all the rabbits' nerve function was normal, without any change.
2 CSEP
The average latent period of CSEP in the lower limbs of the experimental group was 16.21 + 1.7ms before the operation of the experimental group. The electromyogram of the experimental group was compared with the control group. The somatosensory evoked potential was significantly changed in the amplitude and latency, and increased with the time. The control group had no change compared with the pre operation.
3 x - ray examination
X-ray examination showed that the developing resin compression screw was located in the C3 vertebral body, and it developed into the vertebral canal clearly.
4 CT examination
The position of the compression nail can be observed obviously in the sagittal and axial images of CT. The position and size of the compression nail into the spinal canal can be clearly observed. The average occupancy rate of the spinal canal is 40.2%.
5 MRI examination
MRI examination could observe the presence of spinal cord compression in the axial image. The average compression rate of the spinal cord was 43.9%. at the T2 weighted image sagittal position. The signal intensity of the spinal cord was observed in the compression area. The signal intensity of the naked eye was significantly higher than that in the uncompressed area and the normal spinal signal, and the signal was gradually enhanced with time. According to the group, there was a statistically significant difference between the 3 months and the 6 months in the three groups.
6 pathological examination
3 months after compression, the number of neurons in the spinal cord of the spinal cord of the rabbit decreased, the area was reduced, the Nissl body was shallow. The white matter was formed and demyelinating. The anterior horn of the spinal gray matter was compressed, the anterior horn cells of the spinal cord were flattened, the spinal cord anterior horn cells disappeared or necrotic, the leukocyte degeneration, axonosis, and demyelinating changes of the.DAB staining were found in the control group. The shape of the tube was normal and the number was more. The blood vessels of the ventral ventral compression of the spinal cord were reduced and the gray matter vessels were a little more. The blood vessels were sparse in the compression group for 6 months, and a small amount of blood vessels were distributed in the gray matter. The blood vessels were shorter and thinner than the normal form in the 3 month group.
Conclusion:
The animal model of cervical spondylotic myelopathy, established by a special developing resin spinal cord compression nail, has a typical clinical feature of cervical spondylotic myelopathy. This model can be used for various clinical CSM related tests such as CSEP, X ray, CT, MRI and pathological examination. The model is simple and reproducible. It is a combined clinical study of the spinal cord type. A reliable model of vertebral disease.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R687.3;R-332
本文编号：2125140
[Abstract]:Objective: the purpose of this study was to try to develop a new spinal cord compression nail to establish a rabbit experimental model with typical cervical spondylotic myelopathy (CSM) clinical features. This animal model accords with the changes of CSM nerve function and pathological changes, and can be used in various CSM clinical examination methods such as X, CT, MRI and so on. And basic research.
Materials and methods:
1 material
New Zealand rabbit, 30, male. Provided by the animal experiment center of Hebei Medical University, general level; nuclear magnetic machine is produced by Siemens Avanto MR (German Siemens); the CT machine is produced by Siemens company of Germany (SOMATOM Sensation 16 CT); the X ray machine is produced by the French Trophy company (KODA2200); the electromyogram machine is made by Nicolet company (VikingQuest) of the United States. Methyl methacrylate (PMMA), gum cusp and lilac oil are purchased from Shanghai medical equipment Limited by Share Ltd dental materials factory. Zinc Oxide is purchased from Nanchang Baiyun Pharmaceutical Co., Ltd.
2 method
(1) polymethylmethacrylate (PMMA), gutta percha, Zinc Oxide and clove oil were used to prepare developing resin compression screws for spinal cord.
(2) animal feeding and grouping: feeding the ordinary laboratory and feeding the environment at 16-20. The experimental rabbits were randomly divided into the experimental group (20) and the control group (10) for 1 weeks before the operation for 1 weeks. The experimental group was divided into 3 months (10) and 6 months (10).
(3) animal model making: first intraperitoneal injection of diazepam 0.5ml, then intraperitoneal injection of fast dormant new II (0.1ml/kg) general anesthesia, skin preparation, taking the right side of the trachea on the right side of 0.5cm, subchondral 1cm, longitudinal incision length about 5cm, separating the layer by layer to the anterior vertebra, determining the C3 vertebral body, the diameter of the anterior opening of the C3 vertebral body about 3 x 3mm, scraping the cancellous bone to the anterior spinal canal wall of the vertebral canal. The prepared resin compression spinal cord is placed into the hole gently, and the muscle is reset and stitched by layer.
(4) observation of nerve function: before the operation, the first day after operation, 3 months after operation and 6 months after the operation, the modified Tarlov 's motor function scoring method was used to evaluate the motor function, and the records were recorded.
(5) the cortical somatosensory evoked potential (CSEP) examination: CSEP examination of the two lower limbs was performed before the operation, the amplitude of the graph was observed and the latency P15 was recorded. The CSEP was measured immediately after the operation. The EMG was measured at 3 and 6 months after the operation, and the amplitude and the latent period were compared.
(6) x - ray examination: the X - ray examination was performed 1 months after the operation to determine that the model could be clearly developed under X - ray, and the position of the compression nail was determined, and the correlation between the compression nail and the cervical spine of the rabbit was determined.
(7) CT examination: 1 months after the operation, CT examination was performed to determine the feasible CT examination of the model, to determine the position of the compression nail in the sagittal position, to press the relationship between the nail and the cervical spine; to observe the depth of the vertebral canal in the axial position, and to measure the transverse diameter of the spinal canal, the sagittal diameter and the distance between the front of the compression nail and the posterior wall of the spinal canal (the shortest canal of the vertebral canal), and calculate the average occupancy rate of vertebral canal (vertebrae). Tube occupancy rate = (anterior and posterior spinal canal diameter shortest) / vertebral canal anteroposterior diameter * 100%.
(8) MRI examination: the spinal cord compression was observed at 3 months and 6 months after the operation, and the spinal cord compression was observed on the T2 weighted image axis. The spinal cord compression rate (spinal cord compression ratio = sagittal diameter / transverse diameter * 100%) was measured with the transverse diameter of the spinal cord. The most obvious position in the T2 weighted sagittal spinal cord compression signal was enhanced. The intensity of signal intensity was measured in the most serious part of the spinal cord compression (area 0.10 cm2), and the intensity of signal intensity was measured in the area of C6/7 vertebral spinal cord segment (area 0.10cm2), and the ratio was calculated and carried out. In the control group, the signal intensity of the spinal cord and C6/7 segment of the C3 vertebral body was measured and the ratio was calculated.
(9) light microscope observation: animals were sacrificed at 3 months and 6 months respectively. Spinal cord compression stage was performed. HE staining and DAB staining were used to observe cell changes and changes in spinal cord microvasculature.
3 statistical analysis used statistical software SPSS13.0 to process data for statistical analysis.
(1) calculate the average latency of CSEP, the average spinal canal occupancy rate under CT, and the average spinal cord compression rate under MRI.
(2) the MRI signal ratio, the Tarlov 's score, the median of the CSEP latency (P50) and the four quantile spacing (P75? P25) were described. The difference between the groups was analyzed by the Kruskal-Wallis H test, and the two samples after the adjustment of alpha level (alpha' =0.0167) were compared with the Wilcoxon rank sum test to compare 22 of the groups.
Result:
1 nerve function changes in the experimental group first appeared in 3 months, and the motor function difference was larger than normal at 6 months. In the control group, all the rabbits' nerve function was normal, without any change.
2 CSEP
The average latent period of CSEP in the lower limbs of the experimental group was 16.21 + 1.7ms before the operation of the experimental group. The electromyogram of the experimental group was compared with the control group. The somatosensory evoked potential was significantly changed in the amplitude and latency, and increased with the time. The control group had no change compared with the pre operation.
3 x - ray examination
X-ray examination showed that the developing resin compression screw was located in the C3 vertebral body, and it developed into the vertebral canal clearly.
4 CT examination
The position of the compression nail can be observed obviously in the sagittal and axial images of CT. The position and size of the compression nail into the spinal canal can be clearly observed. The average occupancy rate of the spinal canal is 40.2%.
5 MRI examination
MRI examination could observe the presence of spinal cord compression in the axial image. The average compression rate of the spinal cord was 43.9%. at the T2 weighted image sagittal position. The signal intensity of the spinal cord was observed in the compression area. The signal intensity of the naked eye was significantly higher than that in the uncompressed area and the normal spinal signal, and the signal was gradually enhanced with time. According to the group, there was a statistically significant difference between the 3 months and the 6 months in the three groups.
6 pathological examination
3 months after compression, the number of neurons in the spinal cord of the spinal cord of the rabbit decreased, the area was reduced, the Nissl body was shallow. The white matter was formed and demyelinating. The anterior horn of the spinal gray matter was compressed, the anterior horn cells of the spinal cord were flattened, the spinal cord anterior horn cells disappeared or necrotic, the leukocyte degeneration, axonosis, and demyelinating changes of the.DAB staining were found in the control group. The shape of the tube was normal and the number was more. The blood vessels of the ventral ventral compression of the spinal cord were reduced and the gray matter vessels were a little more. The blood vessels were sparse in the compression group for 6 months, and a small amount of blood vessels were distributed in the gray matter. The blood vessels were shorter and thinner than the normal form in the 3 month group.
Conclusion:
The animal model of cervical spondylotic myelopathy, established by a special developing resin spinal cord compression nail, has a typical clinical feature of cervical spondylotic myelopathy. This model can be used for various clinical CSM related tests such as CSEP, X ray, CT, MRI and pathological examination. The model is simple and reproducible. It is a combined clinical study of the spinal cord type. A reliable model of vertebral disease.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R687.3;R-332
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