脾脏切除对SD大鼠大脑中动脉闭塞模型的影响

发布时间：2018-02-13 08:10

本文关键词： 脑梗塞脾切除术免疫炎症大鼠　出处：《中山大学》2012年硕士论文　论文类型：学位论文

【摘要】：背景和目的卒中是最常见的神经科疾患，是由各种血管性病因引起的脑部疾病的总称。其中绝大部分（80％以上）为缺血性脑卒中，由于各种原因未能早期诊断和有效治疗，其中有10％的病人死亡，50％以上的病人留有永久性残疾，给患者本人、家庭和社会均造成沉重的负担。急性脑梗塞的治疗目前仍富有挑战，需要继续寻找新的治疗手段。寻找新的治疗方法，需要对脑梗塞的病理生理机制进行更深入的研究。然而，脑梗塞的病理生理机制非常复杂，主要包括以下几方面：酸中毒，细胞内钙离子超载，兴奋性氨基酸（excitatory amino acids，EAAs）毒性，，炎症细胞因子的损害。目前免疫调节治疗在动物实验中取得了一定的成果。免疫调节治疗可能是脑梗塞治疗的一个突破口。脾脏作为主要的免疫器官，在脑梗塞后的脑损伤机制中发挥重要作用。研究发现，脑梗塞之后，脾脏会出现明显萎缩，脾脏的功能会发生改变。有学者认为，脾脏可能是脑梗塞治疗的一个潜在靶点。目前国际上关于脾脏与脑梗塞的关系的研究并不多见，国内未见相关研究。因此有必要对脾与脑梗塞的关系进行更深一步的研究。本研究对脾切除大鼠采用线栓法建立大脑中动脉闭塞（middle cerebral arteryocclusion，MCAO）模型，探讨脾切除对大鼠脑梗塞的影响及其可能机制。方法（1）动物分组：将健康雄性Sprague-Dawley（SD）大鼠随机分为三组：MCAO对照组、假脾切除组、脾切除组，每组12只。（2）脾切除模型的制作：按大鼠腹部手术常规开腹，分离结扎血管，切除脾脏，彻底止血后关腹。假脾切除组找到脾脏后游离脾脏，余同脾切除组。（3）MCAO模型的制作：采用线栓法建立MCAO大鼠模型。（4）尼氏小体染色测定脑梗塞的体积。（5）免疫荧光法检测脑缺血区T细胞、中性粒细胞、巨噬细胞。（6）ELISA方法测定缺血侧脑匀浆和血清的IL-1β、TNF-α、IL-10水平。结果（1）脾切除组大鼠脑梗塞体积为34.93%±3.23%，比假脾切除组减少53.01%（74.33%±2.36%，P 0.001），比MCAO对照组减少54.81%（77.30%±2.62%，P 0.001）。（2）在脑缺血区，MCAO对照组（52.15±10.85/mm~2，p 0.001）和假脾切除组（46.46±13.44/mm~2，p 0.001）的T细胞数量较脾切除组（17.44±7.33/mm~2）明显增多。（3）在脑缺血区，MCAO对照组（56.46±11.25/mm~2，p=0.001）和假脾切除组（47.56±9.04/mm~2，p=0.032）的中性粒细胞较脾切除组（35.56±4.83/mm~2）明显增多。（4）在脑缺血区，MCAO对照组（39.31±8.69/mm~2，p 0.001）和假脾切除组（43.13±9.10/mm~2，p 0.001）的巨噬细胞数量较脾切除组（7.08±2.23/mm~2）明显增多。（5）缺血侧脑匀浆炎症因子：MCAO对照组（23.06±7.01pg/mg，p=0.04）和假脾切除组（24.04±4.81pg/mg，p=0.02）的脑匀浆IL-1β浓度较脾切除组（16.63±1.09pg/mg）明显增高。MCAO对照组（2.20±0.29pg/mg，p=0.01）和假脾切除组（2.09±0.39pg/mg，p=0.034）的脑匀浆TNF-α浓度较脾切除组（1.71±0.10pg/mg）明显增高。对于IL-10来说，趋势刚好相反，脾切除组脑匀浆IL-10的浓度（1.14±0.28pg/mg）较MCAO对照组（0.67±0.22pg/mg，p=0.002）和假脾切除增高（0.58±0.09pg/mg，p 0.001）。（6）血清炎症因子：IL-1β, TNF-α的ELISA结果与脑匀浆的结果相符，具有相同的趋势。IL-10各个组之间的差异没有统计学意义。结论（1）脾脏切除能明显减少MCAO大鼠模型的脑梗塞体积。（2）脾脏切除改善脑梗塞可能与炎症细胞（T细胞、中性粒细胞、巨噬细胞）的减少有关。（3）脾脏切除改善脑梗塞可能与促炎因子（IL-1β和TNF-α）的减少有关。（4）脾脏切除改善脑梗塞可能与抗炎因子（IL-10）的增多有关。
[Abstract]:Background and Purpose Stroke is the most common neurological disease , which is the general name of brain disease caused by various vascular aetiologies . Most of the patients ( more than 80 % ) are ischemic stroke , with 10 % of the patients died and more than 50 % of the patients left permanent disability , resulting in a heavy burden on the patients themselves , the family and society . However , the pathological and physiological mechanism of cerebral infarction is very complex , including the following aspects : acidosis , overload of intracellular calcium ion , excitatory amino acids ( EAAs ) toxicity , and damage of inflammatory cytokines . Immunoregulatory treatment may be a breakthrough in the treatment of cerebral infarction . The spleen , as the main immune organ , plays an important role in the mechanism of brain damage after cerebral infarction . It has been found that spleen may be a potential target for the treatment of cerebral infarction after cerebral infarction . Some scholars believe that the spleen may be a potential target for cerebral infarction treatment . At present , the study of the relationship between spleen and cerebral infarction is not uncommon . There is no correlation study in China . Therefore , it is necessary to study the relationship between spleen and cerebral infarction . In order to study the effect of splenic resection on cerebral infarction in rats and its possible mechanism , the rat model of middle cerebral artery occlusion was established . method ( 1 ) Group of animals : healthy male Sprague - Dawley ( SD ) rats were randomly divided into three groups : the control group , sham - splenic resection group and the splenic resection group , 12 rats in each group . ( 2 ) The spleen resection model was made : the abdominal operation of the rats was performed routinely , the ligation vessel was separated , the spleen was removed , the spleen was removed , and the spleen was removed completely . The spleen was removed from the spleen after the spleen was removed , and the remainder was removed from the spleen . ( 3 ) Making rat model by line plug method . ( 4 ) The volume of cerebral infarction was determined by Nissella staining . ( 5 ) Immunofluorescence assay was used to detect T cells , neutrophils and macrophages in cerebral ischemia area . ( 6 ) The levels of IL - 1尾 , TNF - 伪 and IL - 10 were determined by ELISA . Results ( 1 ) The volume of cerebral infarction in the spleen resection group was 34.93 % 卤 3.23 % , which was 53.01 % ( 74.33 % 卤 2.36 % , P 0.001 ) than the sham - splenic resection group , which was 54.81 % ( 77.30 % 卤 2.62 % , P 0.001 ) than the control group . ( 2 ) In the cerebral ischemic area , the number of T cells ( 52.15 卤 10.85 / mm ~ 2 , p 0.001 ) and sham - splenic resection group ( 46.46 卤 13.44 / mm ~ 2 , p 0.001 ) were significantly higher than those in the spleen resection group ( 17.44 卤 7.33 / mm ~ 2 ) . ( 3 ) In the focal cerebral ischemic area , the neutropenia group ( 56.46 卤 11.25 / mm ~ 2 , p = 0.001 ) and sham - splenic resection group ( 47.56 卤 9.04 / mm ~ 2 , p = 0.032 ) were significantly higher than those in the spleen resection group ( 35.56 卤 4.83 / mm ~ 2 ) . ( 4 ) In the cerebral ischemic area , the number of macrophages ( 39.31 卤 8.69 / mm ~ 2 , p 0.001 ) and sham - splenic resection group ( 43.13 卤 9.10 / mm ~ 2 , p 0.001 ) were significantly higher than those in the splenic resection group ( 7.08 卤 2.23 / mm ~ 2 ) . ( 5 ) The level of IL - 1尾 in the brain homogenate was significantly higher than that in the spleen resection group ( 16.63 卤 1.09pg / mg , p = 0.04 ) and sham - splenic resection group ( 24.04 卤 4.81pg / mg , p = 0.034 ) . The concentration of IL - 10 ( 1.14 卤 0.28pg / mg , p = 0.002 ) and sham - spleen resection group increased significantly ( 0 . 58 卤 0 . 09pg / mg , p 0.001 ) . ( 6 ) Serum inflammatory factors : IL - 1尾 , TNF - 伪 ELISA results were consistent with the results of brain homogenates , with the same trend . There was no statistical significance between IL - 10 and IL - 10 . mg,p= ( 1 ) Splenectomy can significantly reduce the volume of cerebral infarction in the rat model . ( 2 ) Splenectomy improving cerebral infarction may be associated with a decrease in inflammatory cells ( T cells , neutrophils , macrophages ) . ( 3 ) The improvement of cerebral infarction with spleen resection may be related to the reduction of pro - inflammatory factors ( IL - 1尾 and TNF - 伪 ) . ( 4 ) The improvement of cerebral infarction with spleen resection may be related to the increase of anti - inflammatory factor ( IL - 10 ) .

【学位授予单位】：中山大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R-332

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