功能MRI动态监测大鼠急性缺血性脑卒中缺血再灌注损伤及其与预后相关的实验研究

发布时间：2018-06-04 00:25

本文选题：缺血再灌注损伤 + 大鼠大脑中动脉闭塞模型　；参考：《复旦大学》2014年博士论文

【摘要】：第一部分大鼠急性缺血性脑卒中BBB通透性动态变化研究目的：本实验采用大鼠大脑中动脉闭塞(Middle cerebral artery occlusion, MCAO)模型,运用动态对比增强(Dynamic contrast enhanced, DCE)磁共振成像(Magnetic resonance imaging, MRI)技术,研究急性缺血性脑卒中血脑屏障(Blood brain barrier, BBB)通透性动态变化规律。材料与方法：采用线栓栓塞法制备SD大鼠MCAO模型60只,根据缺血时间(3 h、6 h、永久栓塞)及再通时间(2 h、6 h、12 h、24 h),随机分为12组,每组5只。各组相应时间点行MRI检查,所得图像经西门子工作站处理后测量BBB通透性参数：Ktrans?Ve、Kep及rKtrans、rVe、 rKep,观察各组BBB通透性变化,并与病理学“金标准”伊文思蓝(Evans blue, EB)染色漏出量相对比。结果：缺血3小时、6小时组再灌注后BBB通透性均呈现双相性改变,即再通后2小时及6小时rKtrans值与rVe值升高,rKep值降低,BBB通透性增高,再通12小时BBB通透性降低,再通24小时通透性再次升高,均以再通6小时BBB通透性最大,缺血6小时组各再通时间点均较缺血3小时组BBB通透性大。永久栓塞组BBB通透性呈现单相性改变,即随着缺血时间延长BBB通透性持续轻度增高。MRI观察结果与EB染色漏出量相一致。各组组内及组间比较rKtrans值、rVe值与rKep值均具统计学差异(P0.05),且Ktrans值与Ve值呈正相关(相关系数为0.93,P0.01)；Ktrans值与Kep值呈负相关(相关系数为-0.84,P0.01)；Ve值与Kep值呈负相关(相关系数为-0.8,P0.01)。结论：缺血再灌注组BBB通透性呈双相性变化,永久栓塞组BBB通透性呈单相性变化。DCE-MRI技术可准确反映脑缺血后BBB通透性改变,BBB通透性受缺血时间及再通时间两个因素影响,随着缺血时间的延长,BBB通透性增加所需的时间就越短,BBB破坏的时间较早,且破坏程度明显增加。第二部分DCE-MRI技术预测急性缺血性脑卒中HT的实验研究目的：本实验通过观察缺血再灌注损伤引起的出血性转换(Hemorrhagic transformation, HT)发生率,从而进一步探讨DCE-MRI技术早期预测HT可行性。材料与方法：健康雄性SD大鼠50只,随机分为5组,每组10只,分别为3 h、4.5 h、6h、7.5h、9h缺血再灌注组。所有实验动物均建立MCAO缺血再灌注模型,并于再灌注前即刻及灌注后6小时分别行MRI检查,扫描序列包括：DWI(Diffusion weighted imaging, DWI)、DCE-MRI及磁敏感加权成像(Susceptibility weighted imaging, SWI)。根据SWI图像及最终病理结果,比较性分析再灌注前HT组及无HT组组间rKtrans、rVe及rKep值差异性,并行二分类Logistic回归及ROC曲线分析,判断再灌注前预测HT发生的危险因素及其敏感性、特异性。结果：3 h、4.5h缺血再灌注组均未见HT发生,6h、7.5h、9h缺血再灌注组中HT发生分别为1只、3只及4只,缺血6h后HT发生率为26.7%。回顾性分析HT组及无HT组再灌注前rKtrans、rKep、rVe值,再灌注前纹状体区及皮层区rKtrans及rVe值HT组均高于无HT组(P0.05),而再灌注前纹状体区及皮层区rKep值两组间无统计学差异(P0.05)。二分类Logistic回归分析显示：仅纹状体区rKtrans值是预测HT的危险因素,其OR值为263.614,结合ROC曲线,取Youden指数最大值为预测出血的最佳临界点,纹状体rKtrans值为1.565时,预测HT的敏感性91.4%,特异性为73.9%。结论：纹状体区rKtrans值可成为预测急性缺血性脑卒中继发HT的影像学指标。除了时间点外(大于6小时),结合纹状体区rKtrans值,将有助于早期预测HT。第三部分DCE-MRI技术预测缺血再灌注损伤的实验研究目的：本实验观察不同缺血时间点大鼠MCAO模型缺血再灌注前、后影像学及行为学动态变化,分析DCE.MRI参数与急性缺血性脑卒中预后的关系,探讨有效预测及监测缺血再灌注损伤的影像学指标。材料与方法：健康雄性SD大鼠50只,随机分为5组,每组10只,分别为3h、4.5h、6h、7.5h、9h缺血再灌注组。所有实验动物均建立MCAO缺血再灌注模型,每组动物于再灌注前即刻及再灌注后6小时分别行MRI检查,根据SWI图像及病理学结果将发生HT的实验动物剔除,其余实验动物为入组实验动物,入组实验动物的DCE原始数据经工作站后处理,测量得到缺血脑组织皮层区及纹状体区rKtrans、rVe及rKep值,并进一步求得灌注前、后的各参数差值(△rKtrans、△rVe及△rKep)；入组实验动物于再灌注前即刻及再灌注后24小时行神经功能行为学评分,求得再灌注前、后神经功能行为学评分差值,神经功能行为学评分差值为正值认为预后良好,神经功能行为学评分差值为负值或0认为预后不良；将代表BBB通透性的各参数差值与代表最终预后的神经功能行为学评分差值进行统计分析,寻找可以预测脑卒中缺血再灌注损伤的DCE指标,确定最佳预测指标及其最佳临界点、敏感性及特异性。结果：不同缺血时间点治疗前后纹状体区rKtrans值有显著差异(F=64.94,P=0.000),多重比较,组间ArKtrans值差异均有统计学意义,ArKtrans值随缺血时间延长逐渐增高。不同缺血时间点治疗前后皮层区rKtrans值有显著差异(F=6.973,P=0.000),多重比较,缺血113h、4.5h及6h组间ArKtrans值差异无统计学意义(P0.05),但均高于缺血7.5h及9h组的ArKtrans(P0.05),缺血7.5h与9h组间ArKtrans值差异无统计学意义(P0.05)。不同缺血时间点治疗后纹状体区rVe间有显著差异(F=67.935,P=0.000),多重比较,缺血4.5 h及6h组间△rVe值差异无统计学意义(P0.05),余组间ArVe值比较差异均有统计学意义(P0.05)。不同缺血时间点治疗后皮层区rVe值间有显著差异(F=70.352,P=0.000),多重比较,缺血4.5 h及6 h间ArVe值差异无统计学意义(P0.05),余组间ArVe值比较差异均有统计学意义(P0.05)。不同缺血时间点治疗后纹状体区rKep值间有显著差异(F=46.808,P=0.000),多重比较,缺血3 h及4.5 h间ΔrKep值差异无统计学意义(P0.05),余组间ΔrKep值比较差异均有统计学意义(P0.05)。不同缺血时间点治疗后皮层区rKep值间有显著差异(F=143.317,P=0.000),多重比较,缺血3h及4.5h间ArKep值差异无统计学意义(P0.05),余组间ArKep值比较差异均有统计学意义(P0.05)。将指标进行二分类Logistic回归分析显示：纹状体区ArKtrans值,皮层区△rVe值是预后不良的危险因素,其OR值分别为35.42及19.13,进一步结合ROC曲线,取Youden指数最大值,为判断预后的最佳临界点,纹状体ArKtrans值为1.020时,诊断敏感性93.3%,特异性为88.9%；皮层区△rVe为1.036,诊断敏感性86.7%,特异性为88.9%。结论：结合治疗前后纹状体区ArKtrans值及皮层区△rVe值,将有助于早期判断再灌注损伤,合理安排进一步干预治疗。
[Abstract]:The first part of the study on the dynamic changes of BBB permeability in rats with acute ischemic stroke. The experiment was conducted by using the Middle cerebral artery occlusion (MCAO) model in rats and the dynamic contrast enhancement (Dynamic contrast enhanced, DCE) magnetic resonance imaging (Magnetic resonance) technique to study the acute ischemia. The dynamic changes in the permeability of Blood brain barrier (BBB). Materials and methods: 60 MCAO models of SD rats were prepared by thread embolism method. According to the ischemia time (3 h, 6 h, permanent embolism) and the time of re passage (2 h, 6 h, 12 h, 24 h), 12 groups, 5 in each group. After treatment by SIEMENS workstation, BBB permeability parameters were measured: Ktrans? Ve, Kep and rKtrans, rVe, rKep, observe the changes of BBB permeability, and compare with the pathological "gold standard" Evans blue (Evans blue, EB) stain. Results: 3 hours of ischemia, 6 hours after reperfusion, BBB permeability is biphasic change, that is, after repass The value of rKtrans and rVe increased at 2 hours and 6 hours, the rKep value decreased, the permeability of BBB increased, the BBB permeability was reduced for 12 hours, and the 24 hour permeability increased again. All the 6 hour BBB permeability was the largest. The time points of the 6 hour ischemia group were all more BBB permeability than the 3 hour ischemia group. The BBB permeability in the permanent embolism group showed single-phase permeability. The change, that is, with the prolonged BBB permeability prolonged slightly increased with the duration of ischemia, the results of.MRI observation were consistent with the amount of EB dyed leaking. In each group and between groups, the rKtrans value, rVe value and rKep value were statistically different (P0.05), and Ktrans value was positively correlated with the Ve value (the correlation coefficient was 0.93, P0.01); Ktrans values were negatively correlated with Kep values (correlation coefficient). -0.84, P0.01); Ve value is negatively correlated with Kep value (correlation coefficient is -0.8, P0.01). Conclusion: BBB permeability in ischemia reperfusion group is dual phase change, BBB permeability in permanent embolism group is single-phase change of.DCE-MRI technique can accurately reflect the change of BBB permeability after cerebral ischemia, BBB permeability is affected by two factors of ischemia time and repassage time. With the prolongation of ischemic time, the shorter the time required for the increase of BBB permeability, the earlier time of BBB destruction and a significant increase in damage. The objective of the experimental study on the prediction of acute ischemic stroke HT by the second part of DCE-MRI technique: this experiment was conducted by observing the hemorrhagic transformation (Hemorrhagic transformation, HT) induced by ischemia-reperfusion injury. To further explore the occurrence rate, and further explore the feasibility of early prediction of HT by DCE-MRI technology. Materials and methods: 50 healthy male SD rats were randomly divided into 5 groups, 10 rats in each group, 3 h, 4.5 h, 6h, 7.5h, and 9h ischemia-reperfusion group. All experimental animals were established the MCAO ischemia reperfusion model, and were performed MRI before reperfusion immediately and 6 hours after perfusion, respectively. The scanning sequence includes: DWI (Diffusion weighted imaging, DWI), DCE-MRI and magnetic sensitive weighted imaging (Susceptibility weighted imaging, SWI). According to SWI images and final pathological results, the difference between HT group and no group before reperfusion is comparatively analyzed, and the parallel two classification regression and curve analysis are carried out. To determine the risk factors and sensitivity of HT before reperfusion, and the sensitivity and specificity. Results: 3 h, 4.5H ischemia reperfusion group did not have HT, 6h, 7.5h, 9h ischemia reperfusion group were 1, 3 and 4 respectively, and the incidence of HT was 26.7%. retrospective analysis of HT group and no group before reperfusion, before reperfusion, before reperfusion, before reperfusion, before reperfusion, reperfusion before reperfusion, before reperfusion, before reperfusion before reperfusion The rKtrans and rVe value HT groups in the striatum and cortical areas were higher than those in the non HT group (P0.05), but there was no statistical difference between the two groups in the striatum and the cortical areas before reperfusion (P0.05). The two classification Logistic regression analysis showed that the rKtrans value of the striatum was the risk factor for predicting HT, and the OR value was 263.614. The value is the best critical point for predicting bleeding. When the rKtrans value of the striatum is 1.565, the sensitivity of HT is predicted and the specificity is 73.9%. conclusion: the rKtrans value in the striatum can be used to predict the secondary HT in the acute ischemic stroke. Besides the time point (greater than 6 hours) and the rKtrans value of the striatum, it will be helpful to predict HT. early. The third part of the experimental study on the prediction of ischemia-reperfusion injury by DCE-MRI technique. The experiment observed the dynamic changes of posterior image and behavior before ischemia reperfusion in MCAO model of rats with different ischemic time points, and analyzed the relationship between the DCE.MRI parameters and the prognosis of acute ischemic stroke, and explored the effective prediction and monitoring of ischemia reperfusion injury. Materials and methods: 50 healthy male SD rats were randomly divided into 5 groups, 10 rats in each group, which were 3h, 4.5H, 6h, 7.5h, and 9h ischemia reperfusion group. All the experimental animals established the MCAO ischemia-reperfusion model. Each group was examined immediately before reperfusion and 6 hours after reperfusion, respectively, according to the SWI image and pathological results. The experimental animals were eliminated with HT, and the rest of the experimental animals were included in the experimental animals. The DCE original data of the experimental animals were treated by the workstation, and the rKtrans, rVe and rKep values of the cortex and striatum of the ischemic brain tissue were measured, and the difference values of the parameters (delta rKtrans, Delta rVe and delta rKep) before the perfusion were further obtained. Nerve functional behavioral score was scored before reperfusion immediately before reperfusion and 24 hours after reperfusion. The difference between the neurological functional behavior score and the neurological functional behavior score was positive, the difference between the neurological functional behavior score was negative or 0 thought the prognosis was bad, and the difference of the parameters representing the BBB permeability was poor. The value and the difference between the neurobehavioral behavioral score and the final prognosis were statistically analyzed to find the DCE index that could predict the ischemic reperfusion injury of cerebral apoplexy, and determine the best predictor and its best critical point, sensitivity and specificity. Results: there were significant differences in the rKtrans value of the striatum in different ischemic time points (F=64.94, P=0 .000), with multiple comparison, the difference of ArKtrans values between groups had statistical significance, and the ArKtrans value increased gradually with the prolongation of ischemic time. There was a significant difference in the rKtrans value of the cortical region before and after the treatment of different ischemic points (F=6.973, P=0.000), multiple comparison, and there was no statistically significant difference in ArKtrans values between the ischemic 113h, 4.5H and 6h groups (P0.05), but all higher than those of the ischemic 7.5h. And ArKtrans (P0.05) of group 9h, there was no significant difference in ArKtrans between ischemic 7.5h and 9h group (P0.05). There was significant difference between rVe (F=67.935, P=0.000) in the striatum after different ischemic time points (F=67.935, P=0.000), and there was no significant difference between 4.5 h and 6h groups. P0.05). There were significant differences (F=70.352, P=0.000) between different ischemic time points (F=70.352, P=0.000). There was no statistically significant difference in ArVe values between 4.5 h and 6 h in ischemia (P0.05), and there were significant differences in ArVe values between the remaining groups (P0.05). There was significant difference between the rKep values of the striatum after the treatment of different ischemic points (F=46.808, 000), multiple comparison, there was no significant difference in the value of delta rKep between 3 h and 4.5 h of ischemia (P0.05), and there was significant difference in the value of delta rKep between the remaining groups (P0.05). There was significant difference between the rKep values of the cortical region after the different ischemic time points (F=143.317, P=0.000), the multiplicity ratio, the ArKep value difference between the 3H and 4.5H ischemia, the remaining group The difference of ArKep value was statistically significant (P0.05). The two classification Logistic regression analysis showed that the ArKtrans value of the striatum, the rVe value of the cortical region was the risk factor of poor prognosis, the OR value was 35.42 and 19.13 respectively, and the maximum value of the Youden index was further combined with the ROC curve, which was the best critical point to judge the prognosis. When the body ArKtrans value is 1.020, the diagnostic sensitivity is 93.3%, the specificity is 88.9%, the cortical area Delta rVe is 1.036, the diagnostic sensitivity is 86.7%, the specificity is 88.9%. conclusion: the ArKtrans value of the striatum and the delta rVe value of the cortex area before and after the treatment will be helpful to the early judgment of reperfusion injury and the rational arrangement of the intervention treatment.
【学位授予单位】：复旦大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R445.2;R743.3

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