【作者】 钱军；

【导师】 杨惠林；

【作者基本信息】 苏州大学 ， 外科学（专业学位）， 2014， 博士

【摘要】 第一部分振荡电场刺激大鼠脊髓损伤模型后脊髓少突胶质前体细胞的增殖和分化目的：观察振荡电场刺激后，损伤脊髓内少突胶质前体细胞增殖和分化为新生少突胶质细胞的数量和表型，揭示振荡电场刺激对少突胶质前体细胞的活化作用。方法：建立SD大鼠脊髓损伤模型，分为脊髓损伤组、振荡电场刺激组和正常组。运用BrdU标记及少突胶质细胞系细胞表面标记物标记少突胶质细胞系，采用冰冻切片、免疫荧光染色及电镜等观察方法，检测少突胶质前体细胞的分化及与受体共定位表达。结果：脊髓损伤组、振荡电场刺激组的少突胶质前体细胞的数量在4d至14d时呈现递减的趋势，而成熟少突胶质细胞的数量在4d至14d时呈现递增的趋势；振荡电场刺激组的少突胶质细胞系的细胞数量在4d、7d、10d和14d时均优于脊髓损伤组。振荡电场刺激组在4d、7d、10d和14d时的NG2-P1共定位表达均优于脊髓损伤组；但在振荡电场刺激组内，NG2-P1共定位的表达在14d时弱于4d、7d和10d的表达。结论：振荡电场刺激能改善损伤脊髓内少突胶质前体细胞的活化，促进少突胶质前体细胞分化为成熟少突胶质细胞，利于髓鞘的形成。第二部分振荡电场刺激促进少突胶质前体细胞活化过程中嘌呤能神经递质ATP和细胞因子LIF的含量变化目的：研究振荡电场刺激促进少突胶质前体细胞活化过程中嘌呤能神经递质和细胞因子的含量变化，揭示振荡电场刺激促进少突胶质前体细胞活化的可能机理。方法：建立SD大鼠脊髓损伤模型，分为脊髓损伤组、振荡电场刺激组和正常组。采用ELISA等方法，检测嘌呤神经递质（ATP）、细胞因子（LIF）的含量变化。结果：脊髓损伤组、振荡电场刺激组的损伤脊髓局部ATP、LIF的表达含量在4W、6W时均高于正常组，各组在各点的检测值与正常组比较，均存在统计学差异（P＜0.05）；并且，振荡电场刺激组与脊髓损伤组在损伤脊髓局部的ATP、LIF含量在4W、6W时的检测值比较也存在统计学差异（P＜0.05）；在同一组内的ATP、LIF的含量在脊髓损伤后4W、6W的检测值呈现逐渐增高的趋势。在6周以后，ATP、LIF的含量呈现下降趋势，但振荡电场刺激组在8W时的检测值仍高于正常对照组。结论：脊髓损伤后，在少突胶质前体细胞活化的过程中，振荡电场刺激能够促进嘌呤能神经递质ATP和细胞因子LIF在损伤脊髓局部的表达含量和活性增加，尤其是在振荡电场刺激的早期促进作用明显。表明嘌呤能神经递质ATP和细胞因子LIF在振荡电场刺激促进少突胶质前体细胞活化过程中起着重要作用。第三部分振荡电场刺激修复脊髓损伤后的神经功能恢复目的：评估振荡电场刺激修复脊髓损伤大鼠的神经功能恢复。方法：建立SD大鼠脊髓损伤模型组、振荡电场刺激组和正常对照组。记录并统计不同时点脊髓损伤大鼠的BBB评分、斜板试验评分和MEP检测，并进行统计学分析。结果：在振荡电场刺激的前2周，脊髓损伤组的BBB评分和斜板试验评分高于振荡电场刺激组，从第3周开始，脊髓损伤组的BBB评分和斜板试验评分明显低于振荡电场刺激组，两组之间的差异均存在统计学意义（P<0.05）。而诱发电位检测显示振荡电场刺激组MEP潜伏期在各时点均低于脊髓损伤组，两组之间存在统计学差异（P<0.05）。在建模后2周内，脊髓损伤组和振荡电场刺激组的腹水发生率分别在为14.7%和25.2%，两组之间比较存在统计学差异（P<0.05）。结论：振荡电场刺激能够有效修复大鼠脊髓损伤，促进脊髓损伤大鼠的神经功能恢复。在振荡电场刺激的早期，振荡电场刺激器造成了大鼠腹水的发生，可能恶化了大鼠的机体状态，影响了神经功能的评价。在振荡电场刺激的后期，神经功能的评价相对客观、真实。更多还原

【Abstract】 Part ⅠProliferation and differentiation ofoligodendrocyte precursor cells ofspinal cord injury model rats after oscillating field stimulation.Objective: This study is to evaluate the proliferation and differentiation ofoligodendrocyte precursor cells of spinal cord injury model rats after oscillating fieldstimulation. Methods: SD rats were divided into the spinal cord injury group, oscillatingfield stimulation group and the normal group. Oscillating field stimulation group wastreated by oscillating field stimulation. To detect the proliferation and differentiation ofoligodendrocyte precursor cells by means of frozen sections, immunofluorescence stainingand electron microscopy observation after labeled by Brdu and surface markers ofoligodendrocyte lineage cells. Results: The expression level of oligodendrocyte cell line ofoscillating field stimulation group was higher than that of spinal cord injury group in4d,7d,10d and14d, especially in10d and14d. Colocalization expression of NG2-P1ofoscillating field stimulation group was stronger than that in the spinal cord injury group in4d,7d,10d and14d. But in the oscillating electric field group, NG2-P1expression in14dwas weaker than that in4d,7d and10d. Conclusions: Oscillating field stimulation canimprove oligodendrocyte precursor cells proliferation in the injured spinal cord, whichwould contribute to the formation of myelin after spinal cord injury. Part ⅡTheATPand LIF expression level during the activation ofoligodendrocyte precursor cells after spinal cord injury treated by oscillating fieldstimulation.Objective: This study is to evaluate the expression level of theneurotransmitters purine (ATP) and cytokines (LIF)after promoting oligodendrocyteprecursor cells activation by oscillating field stimulation. Methods: SD rats were dividedinto the spinal cord injury group, oscillating field stimulation group and the normal group.Oscillating field stimulation group was treated by oscillating field stimulation. Detectingthe expression levels of ATP and LIF by method of ELISA. Results: The ATP and LIFexpression levels in the spinal cord injury group and oscillating electrical field stimulationgroup were higher than the normal group, there is a statistical difference between thenormal group and the two groups (P<0.05). Within the two groups, ATP and LIF showed agradual increasing trend from the first to the sixth week after spinal cord injury. But ATPand LIF showed a downward trend after the sixth week. Conclusions: Oscillating fieldstimulation can promote ATP and LIF expression levels in the injuried spinal cordespecially early atage after oscillating electric field stimulation. It showed that ATP andLIF played an important role for the activation of oligodendrocyte precursor cells afterspinal cord injury by the treatment of oscillating field stimulation. Part ⅢThe neurological function recovery of spinal cord injury rats treated byoscillating field stimulation.Objective: This study is to assessment the neurological function recoveryof spinal cord injury rats treated by oscillating field stimulation. Methods: SD rats weredivided into the spinal cord injury group, oscillating field stimulation group and the normalgroup. Basso-Beattie-Bresnahan(BBB)score, incline plane test and motor evokedpotential(MEP) were recorded and calculated. Results: The BBB score and incline plane score in the spinal cord injury group were more than that in the oscillating field stimulationgroup within two weeks after spinal cord injury. However, the BBB score and incline planescore in the spinal cord injury group were less than that in the oscillating field stimulationgroup from the third week, there is a statistical difference about the BBB score and inclineplane score between the two groups (P<0.05). The results of MEP in the the oscillatingfield stimulation group was better than that in the spinal cord injury group from first toeighth week. The incidence of ascites is14.7%and25.2%in the spinal cord injury groupand oscillating field stimulation group, there is a statistically significant difference (P <0.05)between the two groups(P<0.05). Conclusions: Oscillating field stimulation caneffectively repair spinal cord injury and promote neurological function recovery. Theinstrument of oscillatingl field stimulation may contribute to the occurrence of ascites inrats affecting the assessment of neurological function in the early stage.更多还原