重复经颅磁刺激对局灶性脑缺血大鼠神经再生及功能恢复的相关研究
发布时间：2018-09-19 09:02
【摘要】：研究目的：利用重复经颅磁刺激对局灶性脑缺血再灌注成年大鼠进行干预,研究重复经颅磁刺激对脑缺血再灌注损伤大鼠运动功能及侧脑室室管膜下区NSCs增殖的影响,并探索可能的机制。材料与方法：将45只成年雄性SD大鼠进行随机分组,分成假手术组、模型组和rTMS组,每组15只。采用右侧大脑中动脉栓塞(middle artery occlusion,MCAO)模型,梗阻80分钟。rTMS组于造模成功后24h开始干预,每天一次,连续干预7天,期间假手术组和模型组不接受任何干预。每组大鼠在术后第1天和第7天给予神经功能评分(neurological severity scores,NSSs),部分大鼠(每组5只)在rTMS组最后一次治疗结束后即刻开始进行腹腔内注射5-溴脱氧尿嘧啶核苷(5-Bromo-2-deoxy Uridine, BrdU)以标记增殖的细胞,每隔4小时注射一次,连续注射3次。另一部分大鼠(每组10只)在术后第7天取患侧缺血脑组织。利用NSSs评分观察rTMS对MCAO大鼠运动功能的影响,利用免疫组化荧光双重标记技术检测患侧SVZ内源性NSCs增殖的情况,利用实时荧光定量反转录聚合酶链式反应(real-time fluorescent quantitative reverse transcription polymerase chain reaction,qRT-PCR)检测microRNA-25(miR-25)相对表达量,利用免疫组织蛋白印迹(Western blotting)检测miR-25增殖相关靶蛋白的变化。统计学数据分析采用单因素方差分析,组间两两比较采用Bonferroni检验。结果：1.模型组和rTMS组的大鼠术后1天时进行NSSs评分结果无显著差异(p0.05)；术后7天时两组大鼠NSSs评分于第1天相比较均得到明显改善(p0.005),且rTMS组大鼠的神经功能改善程度优于模型组大鼠(p=0.019)。2.利用BrdU+/Nestin+标记患侧SVZ增殖的NSCs数,结果发现术后7天,模型组较假手术组NSCs增殖数增多且存在显著的差异(p=0.044);rTMS组较模型组及假手术组NSCs增殖数目均有显著增多且存在统计学意义(p0.005)。3. qRT-PCR的结果显示术后7天时,模型组大鼠患侧脑缺血组织内miR-25的表达明显高于假手术组(p0.001); rTMS组患侧脑缺血组织miR-25表达较其他两组(p0.001)均显著增高。4. Western blotting的结果发现术后7天时,miR-25靶蛋白p57和PTEN在假手术组以及模型组的表达无显著性差异(p0.05)；而rTMS组大鼠患侧脑缺血组织内p57的表达较其他两组明显减少(p0.01),PTEN的表达则较其他两组显著增加(p0.01)。结论：10Hz rTMS干预患侧大脑皮层7天能促进局灶性脑缺血大鼠运动功能的恢复,并促进患侧SVZ内源性NSCs的增殖,且这种促NSCs增殖的作用可能与上调miR-25的表达进而降低其靶蛋白p57的表达有关。研究目的：通过侧脑室注射miR-25拮抗剂抑制miR-25的表达,探索参与调节NSCs增殖的miR-25/p57在rTMS促进局灶性脑缺血后患侧SVZ内源性NSCs增殖中的作用,进一步证实rTMS对内源性NSCs的增殖效应与miR-25/p57密切相关。材料与方法：将成年雄性SD大鼠进行随机分组,分成rTMS组、rTMS+拮抗剂组和rTMS+对照剂组,每组15只。造模方式为右侧大脑中动脉栓塞(middle artery occlusion,MCAO),梗阻80分钟。rTMS+拮抗剂组及rTMS+对照剂组分别予以不同剂量的miR-25拮抗剂和miR-25对照剂侧脑室注射后进行右侧MCAO造模。所有大鼠造模成功后24h开始给予rTMS干预,每天一次,连续干预7天。各组大鼠在术后7天取患侧的脑组织,通过qRT-PCR检测miR-106b-25家族的相对表达量,确定合适的拮抗剂剂量。在miR-25被特异性抑制后,部分大鼠(每组5只)在术后第7天取患侧的脑组织进行Western blotting检测相关靶蛋白的变化,另一部分大鼠(每组5只)在rTMS组最后一次治疗后即刻开始进行腹腔注射BrdU以标记增殖的细胞,每隔4小时一次,连续3次。利用免疫组化荧光双重标记技术检测miR-25抑制后患侧SVZ内源性NSCs的增殖情况。统计学数据分析采用单因素方差分析,组间两两比较采用Bonferroni检验。结果：1.miRNA拮抗剂可抑制相关miRNA表达,并表现出剂量依赖性。2. qRT-PCR结果显示2.5nmol Ant-25可以显著抑制miR-25的表达,且不影响miR-106b家族其他成员的表达。3.Western blotting结果显示miR-25被抑制后,拮抗剂组的p57表达较rTMS组(p=0.018)和对照剂组(p=0.012)显著提高,而p21的表达在各个组之间差异没有统计学意义(p0.05)。4.利用BrdU+/Nestin+标记患侧SVZ增殖的NSCs数,结果显示miR-25被抑制后,rTMS组和对照剂组NSCs增殖数无显著差异(p0.05)；拮抗剂组较rTMS组(p=0.006)及对照剂组(p=0.013)NSCs增殖数明显减少且有统计学意义。结论：MiR-25特异性有效性的抑制后,靶蛋白p57表达升高,而大鼠脑缺血侧SVZ内源性NSCs增殖降低,即10Hz rTMS可以通过干预miR-25来调节其靶蛋白p57的表达,进而促进NSCs的增殖。研究目的：利用rTMS对局灶性脑缺血大鼠进行干预,研究rTMS对脑缺血大鼠学习记忆功能的影响,并观察患侧海马区神经再生和凋亡的情况。材料与方法：将成年雄性SD大鼠进行随机分组,分成7天组和14天组,两组再进行随机分组,分成假手术组、模型组和rTMS三个亚组。采用右侧MCAO模型,梗阻80分钟。rTMS组于造模成功后24h开始干预,每天一次,直到相应治疗时间结束,期间假手术组和模型组不接受任何干预。各组大鼠在术后第1天、第7天和第14天利用磁共振(magnetic resonance imaging,MRI)进行弥散加权成像(Diffusion-weighted images,DWI)检查,并根据DWI获取表观弥散系数(apparent diffusion coefficient,ADC)值。7天组的大鼠(每组5只)在rTMS组最后一次治疗结束后即刻开始进行腹腔内注射BrdU以标记增殖的细胞,每隔4小时注射一次,连续注射3次。另一部分大鼠(每组10只)在术后第7天取患侧缺血脑组织。14天组的部分大鼠(每组5只)于术后第一天开始,每天腹腔注射一次BrdU来标记分化的细胞,连续注射一周。另一部分大鼠在术后第12天进行Morris水迷宫测试,并于第14天处死取患侧海马组织,分别进行Tunel染色(每组5只)和western blotting检测(每组5只)。利用缺血侧ADC值与健侧ADC比值研究rTMS对MCAO大鼠脑梗体积的影响,采用Morris水迷宫实验观察rTMS对MCAO大鼠的学习记忆的影响,利用免疫组化荧光双重标记技术检测患侧SGZ内源性NSCs增殖和分化的情况,利用Tunel染色观察患侧海马神经元凋亡的情况,利用western blotting证实患侧海马凋亡相关蛋白B细胞淋巴瘤／白血病基因2(B cell lymphoma/leukemia gene 2,Bcl-2)和Bcl基因相关蛋白(Bcl2-associated protein X,Bax)的表达变化。统计学数据分析采用单因素方差分析,组间两两比较采用Bonferroni检验。结果：1.模型组和rTMS组的大鼠术后1天时梗死灶体积差异无统计学意义(p=0.878),但两组大鼠都可见明显梗死灶且有统计学意义(p0.001)。第7天(p=0.246)和14天(p=0.132)时模型组和rTMS组之间梗死灶体积无统计学差异,但rTMS组大鼠脑梗死灶体积的改善较模型组的大鼠趋于更优。此外,虽然模型组和rTMS组大鼠在术后7天及14天时,梗死灶体积均较第1天明显减小(p0.001),但模型组(p=0.716)和rTMS组(p=0.264)第14天较第7天差异均没有统计学意义。2. Morris水迷宫结果显示,模型组大鼠与假手术组相比逃避潜伏期明显延长(p=0.042),60s内穿越目标区域的次数也明显减少(p=0.001); rTMS组较模型组大鼠逃避潜伏期显著缩短(p=0.006),60s内穿越目标区域的次数也显著增多(p=0.045)。3.利用BrdU+/Nestin+标记患侧SGZ区增殖的NSCs数,结果显示术后7天,模型组较假手术组增殖的NSCs数增多且有统计学意义(p=0.042);rTMS组较假手术组及模型组增殖的NSCs数均明显增多且有显著差异(p0.001)。4.利用BrdU+/NeuN+标记患侧SGZ区分化的NSCs数,结果显示术后14天,rTMS组较假手术组(p=0.006)及模型组(p=0.021)NSCs分化数目均显著增多且有统计学意义。而模型组较假手术组NSCs分化数未见统计学差异(p0.05)。5. Tunel染色结果显示术后14天,模型组大鼠患侧海马神经元凋亡数目较假手术组显著增多且有统计学意义(p0.001),而rTMS组大鼠患侧海马神经元凋亡数目较模型组明显减少且有统计学差异(p0.001)。6. Western blotting结果显示,与假手术组相比,模型组患侧海马Bcl-2表达下降(p0.01), Bax表达升高(p0.01)。而rTMS组的Bcl-2表达较模型组明显升高(p0.005), Bax的表达则较模型组显著降低(p0.005)。结论：局灶性脑缺血大鼠学习记忆能力下降,但lOHz rTMS能改善局灶性脑缺血大鼠学习记忆的恢复,并能有效促进局灶性脑缺血大鼠患侧SGZ内源性NSCs增殖和分化和通过调节凋亡相关蛋白Bcl-2和Bax抑制局灶性脑缺血大鼠患侧海马神经元的凋亡。
[Abstract]:AIM: To study the effects of repetitive transcranial magnetic stimulation on motor function and proliferation of NSCs in the subependymal zone of the lateral ventricle in rats with focal cerebral ischemia-reperfusion injury, and to explore the possible mechanism. Materials and Methods: Forty-five adult male SD rats were randomly divided into two groups. Fifteen rats in each group were divided into sham operation group, model group and rTMS group. The right middle cerebral artery occlusion (MCAO) model was used and the obstruction lasted for 80 minutes. Neurological severity scores (NSSs) were given on the 7th day. Some rats (5 rats in each group) were injected intraperitoneally with 5-bromo-2-deoxy Uridine (BrdU) to label proliferating cells, once every four hours, three times continuously after the last treatment in rTMS group. On the 7th day after operation, the ischemic brain tissue was taken from the affected side of the rats (10 rats in each group). The effects of rTMS on the motor function of MCAO rats were observed by NSS score. The proliferation of endogenous NSCs in the affected side of SVZ was detected by immunohistochemical fluorescence double labeling technique. Real-time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR) was used. Verse transcription polymerase chain reaction (qRT-PCR) was used to detect the relative expression of microRNA-25 (microRNA-25) and Western blotting was used to detect the changes of proliferation-related target proteins of microRNA-25. There was no significant difference in NSS scores between the rTMS group and the control group on the 1st day after operation (p0.05). The NSS scores of the two groups were significantly improved on the 7th day after operation (p0.005), and the neurological function of the rTMS group was better than that of the model group (p = 0.019). 2. The number of NSCs proliferated by BrdU + / Nestin + labeling SVZ on the affected side was found. On the 7th day after operation, the proliferation of NSCs in the model group was significantly higher than that in the sham operation group (p = 0.044), and the proliferation of NSCs in the rTMS group was significantly higher than that in the model group and sham operation group (p 0.005). 3. The results of qRT-PCR showed that the expression of microwave-25 in the affected cerebral ischemic tissue of the model group was significantly higher than that in the sham operation group on the 7th day after operation. The expression of microRNA-25 in the affected cerebral ischemic tissues of the rTMS group was significantly higher than that of the other two groups (p0.001). 4. Western blotting results showed that there was no significant difference in the expression of microRNA-25 target protein p57 and PTEN between the sham-operated group and the model group at 7 days after operation (p0.05); however, the expression of p57 in the affected cerebral ischemic tissues of the rTMS group was higher than that of the other groups (p0.001). Conclusion: 10Hz rTMS can promote the recovery of motor function and the proliferation of SVZ endogenous NSCs in the affected cerebral cortex in rats with focal cerebral ischemia for 7 days, and this effect may be related to the up-regulation of the expression of microRNA-25 and the down-regulation of its target. Objective: To investigate the role of microRNAs-25/p57 in promoting the proliferation of endogenous NSCs in SVZ after focal cerebral ischemia by rTMS through inhibiting the expression of microRNAs-25 by intracerebroventricular injection of microRNAs-25 antagonist, and further confirm that the proliferation effect of rTMS on endogenous NSCs is closely related to microRNAs-25/p57. Methods: Adult male SD rats were randomly divided into rTMS group, rTMS + antagonist group and rTMS + control group with 15 rats in each group. All rats were treated with rTMS once a day for 7 days. The brain tissues of each group were taken from the affected side 7 days after operation. The relative expression of the microRNA106 B-25 family was detected by qRT-PCR to determine the appropriate antagonist dose. After the microRNA25 was specifically inhibited, part of the brain tissues were partially inhibited. Western blotting was used to detect the changes of target proteins in the brain tissues of rats (5 rats in each group) on the 7th day after operation. BrdU was injected intraperitoneally to label proliferative cells immediately after the last treatment in the rTMS group, once every 4 hours for three consecutive times. Results: 1. MiRNA antagonists inhibited the expression of related microRNAs and showed a dose-dependent effect. 2. qRT-PCR results showed that 2.5 nmol Ant-25 significantly inhibited the expression of microRNAs-25. Western blotting results showed that the expression of p57 in the antagonist group was significantly higher than that in the rTMS group (p = 0.018) and the control group (p = 0.012), but the expression of p21 was not significantly different among the groups (p 0.05). 4. NSCs proliferated in the affected side of SVZ were labeled with BrdU + / Nestin +. The results showed that there was no significant difference in NSCs proliferation between rTMS group and control group (p0.05) after inhibition of microwave irradiation. The number of NSCs proliferation in antagonist group was significantly lower than that in rTMS group (p = 0.006) and control group (p = 0.013). Conclusion: The expression of target protein p57 was increased after inhibition of specificity and validity of MiR-25, but the endogenous SVZ in cerebral ischemic side of rats was significantly lower than that in rTMS group (p = 0.006) and control group (p = 0.01 Objective: To study the effects of rTMS on learning and memory function in rats with focal cerebral ischemia, and to observe the regeneration and apoptosis of hippocampal neurons. METHODS: Adult male SD rats were randomly divided into 7-day group and 14-day group. The rats were divided into sham-operation group, model group and rTMS subgroup. Diffusion-weighted imaging (DWI) was performed on the first day, the seventh day and the fourteenth day after operation, and the apparent diffusion coefficient (ADC) was obtained on the basis of DWI. After the last treatment, BrdU was injected intraperitoneally to label the proliferative cells, once every four hours, three times in succession. On the seventh day after operation, the ischemic brain tissue was taken from the affected side in another group of rats (10 rats in each group). Morris water maze test was performed on the 12th day after operation, and the hippocampus was sacrificed on the 14th day after operation. Tunel staining (5 in each group) and Western blotting (5 in each group) were used to detect the volume of cerebral infarction in MCAO rats. Morris water maze test was used to observe the effect of rTMS on learning and memory in MCAO rats. The proliferation and differentiation of endogenous NSCs in SGZ were detected by immunohistochemical double labeling technique. The apoptosis of neurons in the affected hippocampus was observed by Tunel staining. The apoptosis-related protein B cells in the affected hippocampus were confirmed by Western blotting. The expression of B cell lymphoma/leukemia gene 2 (Bcl-2) and Bcl 2-associated protein X (Bax) was analyzed by one-way ANOVA. Bonferroni test was used to compare the two groups. Results: 1. The infarct volume of the model group and rTMS group was different at 1 day after operation. There was no significant difference in infarct volume between the model group and the rTMS group on the 7th day (p = 0.246) and the 14th day (p = 0.132), but the improvement of infarct volume in the rTMS group was better than that in the model group. The infarct volume of rats in MS group decreased significantly on the 7th and 14th day after operation (p0.001), but there was no significant difference between model group (p = 0.716) and rTMS group (p = 0.264) on the 14th day and the 7th day. The escape latency of rTMS group was significantly shorter than that of model group (p = 0.006), and the number of NSCs proliferating within 60 seconds was also significantly increased (p = 0.045). 3. The number of NSCs proliferating in SGZ of affected side was marked by BrdU + / Nestin +, and the results showed that the number of NSCs proliferating in model group was significantly higher than that in sham operation group 7 days after operation. The number of NSCs differentiated by BrdU+/NeuN+ labeling of SGZ on the affected side was significantly higher in rTMS group than that in sham operation group and model group (p = 0.042), and the number of NSCs differentiated by BrdU+/NeuN+ labeling was significantly higher in rTMS group than that in sham operation group (p = 0.006) and model group (p = 0.021). There was no significant difference in the number of NSCs differentiated between the model group and the sham group (p0.05). 5. Tunel staining showed that the number of neuronal apoptosis in the hippocampus of the model group was significantly higher than that of the sham group at 14 days after operation (p0.001). The results of Western blotting showed that the expression of Bcl-2 decreased (p0.01) and the expression of Bax increased (p0.01) in the affected hippocampus of the model group compared with the sham-operated group. The expression of Bcl-2 in the rTMS group was significantly higher than that in the model group (p0.005), but the expression of Bax was significantly lower than that in the model group (p0.005). However, lOHz rTMS can improve the recovery of learning and memory in rats with focal cerebral ischemia, and can effectively promote the proliferation and differentiation of endogenous NSCs in SGZ and inhibit the apoptosis of hippocampal neurons in rats with focal cerebral ischemia by regulating apoptosis-related proteins Bcl-2 and Bax.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R743.3
[Abstract]:AIM: To study the effects of repetitive transcranial magnetic stimulation on motor function and proliferation of NSCs in the subependymal zone of the lateral ventricle in rats with focal cerebral ischemia-reperfusion injury, and to explore the possible mechanism. Materials and Methods: Forty-five adult male SD rats were randomly divided into two groups. Fifteen rats in each group were divided into sham operation group, model group and rTMS group. The right middle cerebral artery occlusion (MCAO) model was used and the obstruction lasted for 80 minutes. Neurological severity scores (NSSs) were given on the 7th day. Some rats (5 rats in each group) were injected intraperitoneally with 5-bromo-2-deoxy Uridine (BrdU) to label proliferating cells, once every four hours, three times continuously after the last treatment in rTMS group. On the 7th day after operation, the ischemic brain tissue was taken from the affected side of the rats (10 rats in each group). The effects of rTMS on the motor function of MCAO rats were observed by NSS score. The proliferation of endogenous NSCs in the affected side of SVZ was detected by immunohistochemical fluorescence double labeling technique. Real-time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR) was used. Verse transcription polymerase chain reaction (qRT-PCR) was used to detect the relative expression of microRNA-25 (microRNA-25) and Western blotting was used to detect the changes of proliferation-related target proteins of microRNA-25. There was no significant difference in NSS scores between the rTMS group and the control group on the 1st day after operation (p0.05). The NSS scores of the two groups were significantly improved on the 7th day after operation (p0.005), and the neurological function of the rTMS group was better than that of the model group (p = 0.019). 2. The number of NSCs proliferated by BrdU + / Nestin + labeling SVZ on the affected side was found. On the 7th day after operation, the proliferation of NSCs in the model group was significantly higher than that in the sham operation group (p = 0.044), and the proliferation of NSCs in the rTMS group was significantly higher than that in the model group and sham operation group (p 0.005). 3. The results of qRT-PCR showed that the expression of microwave-25 in the affected cerebral ischemic tissue of the model group was significantly higher than that in the sham operation group on the 7th day after operation. The expression of microRNA-25 in the affected cerebral ischemic tissues of the rTMS group was significantly higher than that of the other two groups (p0.001). 4. Western blotting results showed that there was no significant difference in the expression of microRNA-25 target protein p57 and PTEN between the sham-operated group and the model group at 7 days after operation (p0.05); however, the expression of p57 in the affected cerebral ischemic tissues of the rTMS group was higher than that of the other groups (p0.001). Conclusion: 10Hz rTMS can promote the recovery of motor function and the proliferation of SVZ endogenous NSCs in the affected cerebral cortex in rats with focal cerebral ischemia for 7 days, and this effect may be related to the up-regulation of the expression of microRNA-25 and the down-regulation of its target. Objective: To investigate the role of microRNAs-25/p57 in promoting the proliferation of endogenous NSCs in SVZ after focal cerebral ischemia by rTMS through inhibiting the expression of microRNAs-25 by intracerebroventricular injection of microRNAs-25 antagonist, and further confirm that the proliferation effect of rTMS on endogenous NSCs is closely related to microRNAs-25/p57. Methods: Adult male SD rats were randomly divided into rTMS group, rTMS + antagonist group and rTMS + control group with 15 rats in each group. All rats were treated with rTMS once a day for 7 days. The brain tissues of each group were taken from the affected side 7 days after operation. The relative expression of the microRNA106 B-25 family was detected by qRT-PCR to determine the appropriate antagonist dose. After the microRNA25 was specifically inhibited, part of the brain tissues were partially inhibited. Western blotting was used to detect the changes of target proteins in the brain tissues of rats (5 rats in each group) on the 7th day after operation. BrdU was injected intraperitoneally to label proliferative cells immediately after the last treatment in the rTMS group, once every 4 hours for three consecutive times. Results: 1. MiRNA antagonists inhibited the expression of related microRNAs and showed a dose-dependent effect. 2. qRT-PCR results showed that 2.5 nmol Ant-25 significantly inhibited the expression of microRNAs-25. Western blotting results showed that the expression of p57 in the antagonist group was significantly higher than that in the rTMS group (p = 0.018) and the control group (p = 0.012), but the expression of p21 was not significantly different among the groups (p 0.05). 4. NSCs proliferated in the affected side of SVZ were labeled with BrdU + / Nestin +. The results showed that there was no significant difference in NSCs proliferation between rTMS group and control group (p0.05) after inhibition of microwave irradiation. The number of NSCs proliferation in antagonist group was significantly lower than that in rTMS group (p = 0.006) and control group (p = 0.013). Conclusion: The expression of target protein p57 was increased after inhibition of specificity and validity of MiR-25, but the endogenous SVZ in cerebral ischemic side of rats was significantly lower than that in rTMS group (p = 0.006) and control group (p = 0.01 Objective: To study the effects of rTMS on learning and memory function in rats with focal cerebral ischemia, and to observe the regeneration and apoptosis of hippocampal neurons. METHODS: Adult male SD rats were randomly divided into 7-day group and 14-day group. The rats were divided into sham-operation group, model group and rTMS subgroup. Diffusion-weighted imaging (DWI) was performed on the first day, the seventh day and the fourteenth day after operation, and the apparent diffusion coefficient (ADC) was obtained on the basis of DWI. After the last treatment, BrdU was injected intraperitoneally to label the proliferative cells, once every four hours, three times in succession. On the seventh day after operation, the ischemic brain tissue was taken from the affected side in another group of rats (10 rats in each group). Morris water maze test was performed on the 12th day after operation, and the hippocampus was sacrificed on the 14th day after operation. Tunel staining (5 in each group) and Western blotting (5 in each group) were used to detect the volume of cerebral infarction in MCAO rats. Morris water maze test was used to observe the effect of rTMS on learning and memory in MCAO rats. The proliferation and differentiation of endogenous NSCs in SGZ were detected by immunohistochemical double labeling technique. The apoptosis of neurons in the affected hippocampus was observed by Tunel staining. The apoptosis-related protein B cells in the affected hippocampus were confirmed by Western blotting. The expression of B cell lymphoma/leukemia gene 2 (Bcl-2) and Bcl 2-associated protein X (Bax) was analyzed by one-way ANOVA. Bonferroni test was used to compare the two groups. Results: 1. The infarct volume of the model group and rTMS group was different at 1 day after operation. There was no significant difference in infarct volume between the model group and the rTMS group on the 7th day (p = 0.246) and the 14th day (p = 0.132), but the improvement of infarct volume in the rTMS group was better than that in the model group. The infarct volume of rats in MS group decreased significantly on the 7th and 14th day after operation (p0.001), but there was no significant difference between model group (p = 0.716) and rTMS group (p = 0.264) on the 14th day and the 7th day. The escape latency of rTMS group was significantly shorter than that of model group (p = 0.006), and the number of NSCs proliferating within 60 seconds was also significantly increased (p = 0.045). 3. The number of NSCs proliferating in SGZ of affected side was marked by BrdU + / Nestin +, and the results showed that the number of NSCs proliferating in model group was significantly higher than that in sham operation group 7 days after operation. The number of NSCs differentiated by BrdU+/NeuN+ labeling of SGZ on the affected side was significantly higher in rTMS group than that in sham operation group and model group (p = 0.042), and the number of NSCs differentiated by BrdU+/NeuN+ labeling was significantly higher in rTMS group than that in sham operation group (p = 0.006) and model group (p = 0.021). There was no significant difference in the number of NSCs differentiated between the model group and the sham group (p0.05). 5. Tunel staining showed that the number of neuronal apoptosis in the hippocampus of the model group was significantly higher than that of the sham group at 14 days after operation (p0.001). The results of Western blotting showed that the expression of Bcl-2 decreased (p0.01) and the expression of Bax increased (p0.01) in the affected hippocampus of the model group compared with the sham-operated group. The expression of Bcl-2 in the rTMS group was significantly higher than that in the model group (p0.005), but the expression of Bax was significantly lower than that in the model group (p0.005). However, lOHz rTMS can improve the recovery of learning and memory in rats with focal cerebral ischemia, and can effectively promote the proliferation and differentiation of endogenous NSCs in SGZ and inhibit the apoptosis of hippocampal neurons in rats with focal cerebral ischemia by regulating apoptosis-related proteins Bcl-2 and Bax.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R743.3
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