一種新的兔脊髓型頸椎病動(dòng)物模型的建立
發(fā)布時(shí)間：2018-07-15 19:27
【摘要】：目的:本研究的目的是嘗試研制一種新的脊髓壓迫釘用以建立一種帶有典型脊髓型頸椎病(cervical spondylotic myelopathy,CSM)臨床特征的兔實(shí)驗(yàn)?zāi)Ｐ?這種動(dòng)物模型符合CSM神經(jīng)功能變化及病理變化,并可用于X線、CT、MRI等多種CSM臨床檢查方法及基礎(chǔ)研究。 材料與方法: 1材料 新西蘭家兔,30只,雄性。由河北醫(yī)科大學(xué)動(dòng)物實(shí)驗(yàn)中心提供,普通級(jí);核磁機(jī)由德國(guó)Siemens公司生產(chǎn)(Siemens Avanto MR);CT機(jī)由德國(guó)Siemens公司生產(chǎn)(SOMATOM Sensation 16 CT);X線機(jī)由法國(guó)Trophy公司生產(chǎn)( KODA2200 );肌電圖機(jī)由美國(guó)Nicolet公司(VikingQuest);聚甲基丙烯酸甲酯(PMMA)、牙膠尖和丁香油均購自上海醫(yī)療器械股份有限公司齒科材料廠。氧化鋅購自南昌白云藥業(yè)有限公司。 2方法 (1)應(yīng)用聚甲基丙烯酸甲酯(PMMA)、牙膠尖、氧化鋅和丁香油制備顯影樹脂脊髓壓迫釘。 (2)動(dòng)物飼養(yǎng)與分組:給予普通實(shí)驗(yàn)室飲食喂養(yǎng),飼養(yǎng)環(huán)境溫度為16-20℃。術(shù)前飼養(yǎng)1周。30只實(shí)驗(yàn)兔隨機(jī)分為實(shí)驗(yàn)組(20只)和對(duì)照組(10只)。實(shí)驗(yàn)組再分為3個(gè)月組(10只)和6個(gè)月組(10只)。 (3)動(dòng)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,備皮。取氣管右側(cè)旁開0.5cm,甲狀軟骨下1cm,行縱行切口長(zhǎng)約5cm,逐層分離至椎前,確定C3椎體,于C3椎體前開孔直徑約3×3mm,刮出松質(zhì)骨至椎管前壁骨皮質(zhì),將準(zhǔn)備好的顯影樹脂脊髓壓迫釘輕輕置入孔內(nèi),將肌肉復(fù)位逐層縫合。 (4)神經(jīng)功能觀察:分別于術(shù)前、術(shù)后第一天、術(shù)后3個(gè)月和術(shù)后6個(gè)月采用改良Tarlov’s運(yùn)動(dòng)功能評(píng)分法進(jìn)行運(yùn)動(dòng)功能評(píng)估,并記錄。 (5)皮層體感誘發(fā)電位(CSEP)檢查:與術(shù)前行雙下肢CSEP檢查,觀察圖形振幅并記錄潛伏期P15,術(shù)后立即行CSEP檢測(cè),術(shù)后3個(gè)月和6個(gè)月時(shí)測(cè)肌電圖并比較振幅以及潛伏期變化。 (6) X線檢查:術(shù)后1個(gè)月行X線檢查,以確定本模型可于X線下清晰顯影,并確定壓迫釘所在位置,壓迫釘與兔頸椎相關(guān)關(guān)系。 (7) CT檢查:術(shù)后1個(gè)月行CT檢查,以確定本模型可行CT檢查,在矢狀位確定壓迫釘位置,壓迫釘與頸椎關(guān)系;在軸位觀察壓迫釘突入椎管深度,并測(cè)量椎管橫徑、矢狀徑以及壓迫釘前端至椎管后壁距離(椎管最短經(jīng)),并計(jì)算椎管平均占用率(椎管占用率=(椎管前后徑-椎管最短徑)/椎管前后徑×100%)。 (8) MRI檢查:術(shù)后3個(gè)月和6個(gè)月分別進(jìn)行MRI平掃,觀察脊髓形態(tài),于T2加權(quán)像軸位觀察脊髓受壓情況,并測(cè)量脊髓前后徑、脊髓橫徑計(jì)算平均脊髓壓迫率(脊髓壓迫率=椎管矢狀徑/橫徑×100%)。于T2加權(quán)像矢狀面脊髓受壓信號(hào)增強(qiáng)最明顯部位或脊髓受壓最嚴(yán)重部位選取興趣區(qū)(面積為0.10 cm2)測(cè)量信號(hào)強(qiáng)度值,再于C6/7椎體脊髓節(jié)段取興趣區(qū)(面積為0.10cm2)測(cè)量信號(hào)強(qiáng)度值,計(jì)算并進(jìn)行比值;對(duì)照組則選取C3椎體節(jié)段脊髓和C6/7節(jié)段脊髓測(cè)量信號(hào)強(qiáng)度值,計(jì)算比值。 (9)光鏡下觀察:分別于3個(gè)月及6個(gè)月處死動(dòng)物,取脊髓受壓階段脊髓,HE染色及DAB染色觀察細(xì)胞變化及脊髓微血管變化。 3統(tǒng)計(jì)分析使用統(tǒng)計(jì)學(xué)軟件SPSS13.0處理數(shù)據(jù)進(jìn)行統(tǒng)計(jì)學(xué)分析。 (1)計(jì)算CSEP平均潛伏期、CT下平均椎管占用率、MRI下平均脊髓壓迫率。 (2) MRI信號(hào)比、Tarlov’s評(píng)分、CSEP潛伏期采用中位數(shù)(P50)和四分位數(shù)間距(P75? P25)進(jìn)行描述,Kruskal-Wallis H檢驗(yàn)分析組間差異,采用調(diào)整α水準(zhǔn)(α’=0.0167)后的兩樣本比較Wilcoxon秩和檢驗(yàn)進(jìn)行組間兩兩比較。 結(jié)果: 1神經(jīng)功能的變化實(shí)驗(yàn)組術(shù)后神經(jīng)癥狀最初出現(xiàn)在3個(gè)月內(nèi),6個(gè)月時(shí)運(yùn)動(dòng)功能差異較正常大。對(duì)照組,所有兔神經(jīng)功能均正常,無任何改變。 2 CSEP 實(shí)驗(yàn)組在行造模手術(shù)前雙下肢CSEP平均潛伏期為16.21±1.7ms,實(shí)驗(yàn)組肌電圖檢查與對(duì)照組比較,體感誘發(fā)電位在波幅與潛伏期上出現(xiàn)顯著變化,并隨時(shí)間加重。對(duì)照組較術(shù)前無變化。 3 X線檢查 X線檢查可見顯影樹脂脊髓壓迫釘位于C3椎體,突入椎管,顯影清晰。 4 CT檢查 CT顯像矢狀位及軸位像均可明顯觀察到顯影壓迫釘?shù)乃谖恢谩浩柔斖蝗胱倒芪恢眉按笮】汕逦^察。平均椎管占用率為40.2%。 5 MRI檢查 MRI檢查可在軸位像觀察到脊髓壓迫的存在,平均脊髓壓迫率為43.9%。在T2加權(quán)像矢狀位可觀察到受壓區(qū)域出現(xiàn)脊髓高信號(hào),肉眼可觀察到信號(hào)強(qiáng)度明顯高于未受壓區(qū)域及正常脊髓信號(hào),且隨時(shí)間逐漸增強(qiáng)。統(tǒng)計(jì)學(xué)結(jié)果顯示,脊髓信號(hào)比:對(duì)照組,3個(gè)月實(shí)驗(yàn)組和6個(gè)月實(shí)驗(yàn)組三者總體有統(tǒng)計(jì)學(xué)差異,信號(hào)隨時(shí)間逐漸增強(qiáng)。 6病理學(xué)檢 壓迫3個(gè)月后的兔脊髓灰質(zhì)神經(jīng)元數(shù)目減少,面積縮小,尼氏體淺染。白質(zhì)有空洞形成,脫髓鞘改變。壓迫6個(gè)月組,脊髓灰質(zhì)前角被壓扁,脊髓前角細(xì)胞消失或壞死,脊髓白質(zhì)成明顯的空泡樣變,軸索變性,脫髓鞘樣改變。DAB染色發(fā)現(xiàn)對(duì)照組血管形態(tài)正常,數(shù)目多。壓迫3個(gè)月組發(fā)現(xiàn)脊髓腹側(cè)受壓處血管減少,灰質(zhì)血管稍多。壓迫6個(gè)月組受壓處血管稀疏,灰質(zhì)有少量血管分布,血管較正常形態(tài)較短且細(xì)。 結(jié)論: 本實(shí)驗(yàn)應(yīng)用特制顯影樹脂脊髓壓迫釘建立的脊髓型頸椎病動(dòng)物模型具有典型脊髓型頸椎病臨床特征。這種模型可同時(shí)用于CSEP、X線、CT、MRI及病理學(xué)檢查等多種臨床CSM相關(guān)檢查,且模型制作簡(jiǎn)單,可重復(fù)性強(qiáng),是結(jié)合臨床綜合研究脊髓型頸椎病的可靠模型。
[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.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類號(hào)】：R687.3;R-332
本文編號(hào)：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.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類號(hào)】：R687.3;R-332
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