论文标题:血小板活化因子与脑缺血的相关性研究
Study on correlation between platelet activating factor and cerebrovascular ischemia
论文作者 徐彬
论文导师 黄鉴政,论文学位 硕士,论文专业 神经病学
论文单位 浙江大学,点击次数 101,论文页数 38页File Size996k
2002-03-08论文网 http://www.lw23.com/lunwen_962362552/ 心肌梗塞,血管生长,碱性成纤维细胞生长因子,血管内皮细胞生长因子,腺苷,贝那普利,缬沙坦
myocardial infarction,angiogenesis,basic fibroblast growth factor,vascular endothelial growth factor,adenosine,benazepril,valsartan
血小板活化因子(Platelet activating factor，PAF)是近年研究较活跃的细胞因子之一，它具有广泛的生物学活性，可诱导脑血栓形成，造成脑细胞缺血，促发脑细胞Ca内流，改变脑血流以及造成缺血区内的多种生理病理过程，包括兴奋性毒性、自由基、炎症反应及线粒体和DNA损伤可以触发细胞凋亡。随着我国经济发展和人们生活水平的提高以及人口的老龄化，脑血管病的发生率有增高的趋势。血小板活化因子(PAF)在缺血性心脏病、血栓形成等疾病中的作用国内外学者已有所研究。探讨PAF在脑血管病，尤其在脑缺血疾病中的生理学意义，可为脑血管病的防治开辟一个新的领域。为此，我们观察了 研究生学位论文 脑血管病患者PAF水平的动态变化，以期了解PAF在脑缺 血患者中的病理生理学意义。 资料和方法 一、研究对象 1、病例组：120例脑血管病患者均系1999年1月至 ”“年8 月间在浙江大学医学院附属第二医院神经内科 急诊及住院的病人，诊断标准按第四次全国脑血管病会议 制订的诊断标准并经头颅 CT或 MR证实，并删除了可能 影啊际F的其他同素，如台卉患有支气管哮7’克 糖尿病、 缺血性心脏病 旧有心绞痛、心肌梗死等病史）等，入选 脑缺血组病人43例，均在起病72小时内（急性期）和半 月后 广恢复期）各采静脉血1 次，其中男性25例，女性 旧 例，年龄抡 ～钻 岁，平均币 ＊8上＊．7） 岁，另外对 其中7例（男4例、女3例）患者分别于起病的第1、3。 7、15、21、ZS R各采静脉血 1次并检测血中叩F的变化， 以其丛到动态观察的目的。脑出血组21例，男性12例， 女性9例，年龄41～79岁，平均仿1．3门．2）岁。 研究生学位论文 2、疾病对照组：即非脑血管病的其他神经系统疾病 的患者32例，其包括坐骨神经痛7例，颈椎病6例，急 性脊髓炎4例，吉兰－巴雷综合征6例，癫痈5例，原发 性帕金森氏病2例，痉挛性斜颈2例，其中男性16例， 女性16例，年龄41～77岁，平均“4．it10．9）岁。 3、健康对照组：均系健康体检者，排除其他可能影 响PAF的因素后入选46例，其中男性27例、女性19例． 年龄41～SO 岁，平均（6O．3士门．5）岁。 t 以上各组年龄经统计全检验，无显著性差异 （刘）．05）。 二、实验方法 除健康对照组为相隔15天各采静脉血1次、其余各 组均在起病 72小时内（急性期）和 15天（恢复期）各采 静脉血l次。检测过程均在浙江大学医学院附属第二医院 消化实验室内进行，所有标本均能及时处理。PAF标准品 由美国 S i gma公司购得：硅胶 G由上海华美生物工程公司 主产。 PAF采用薄层层析法测定：首先采静脉血2毫升，立 研究生学位论文 即加入预冷的 3毫升氯仿和 6毫升甲醇试管中，混匀，12009 离心15分钟，取上清液与氯仿6毫升和甲醇9．6毫升混 匀，室温静置1小时，同上离心，取出下层氯仿相，37oC 水浴并氮气吹干（通风柜中操作八 取15克硅胶G加水调 至糊状，倾注平板（s－c20cm），120℃ 烘千120分钟，冷 却至室温备用。最后在离底部约11．scm处点样及标准品 （各 10VI），用氯仿：甲醇：水（65：35：6，V／V／V） 展开，展距约10、5cm，展毕凉干，以10％磷钥酸均匀喷 雾，80”C X 30分钟，直至斑点显蓝色，将与标准品盯值 ？目司的层祈带刮下，加，入0．＊湖“土OUL g解，弃仇淀， 上清液比色定量。 结 果 一、脑缺血组、脑出血组的急性期和恢复期PAF水平 比非脑血管疾病组和健康对照组显著升高，而非脑血管疾 病组与健康对照组PAF水平无显著性差别。脑缺血组的急 性期PAF水平比恢复期明显升高，而脑出血和非脑血管疾 病组、健康对照组则无此改变。
Platelet activating factor(PAF), a widely studied cytokine, has extensive biological activities. It can induce cerebral thrombosis, cerebral ischemia, calcium ion influx into brain cells, excitatory toxicity, DNA damage and neuron apoptosis. Role of PAP in the pathogenesis of ischemic hear* disease and thiombosis Ha< hecn elucidated. To discuss the role of PAF in the physiology of cerebral vascular disease is very important. Thus, we observed the dynamic change of PAF in the patients with cerebral vascular disease.Subject and Method1. Subjects(1). Patients: We examined 120 patients with cerebral vascular disease(from Jan 1999 to Aug 2000) hospitalized in Neurological department of Zhejiang University Medical School. Patients were diagnosed by the standard from 4th National Congress for Cerebral Vascular Disease. Patients with bronchial asthma, diabetes mellitus, ischemic heart disease were excluded. There were 43 patients withischemic cerebral vascular disease in acute stage (<72hrs) or in stage of recovery (>15 days), the average age is 65.8?10.7 yrs(42-83), 25 was male, and 18 was female. In 7 patients (4 males and 3 females), PAF was detected on the 1* 3rd, 7th , 15th ,21th, and 28* day after onset for dynamic observation. There were 21 patients(12 male and 9 female ) with cerebral hemorrhage, the aberage age was 61.3 + 12.2yrs(41-79yrs). (2).Disease controls: Totally there were 32 non-cerebrovascular disease patients(16 male and 16 female) including 7 sciatica, 6 cervical syndrome, 4 acute myelitis, 6 GBS, 5 epilepsy, 2 Parkinson"s disease, 2 spasmodic torticollis, the average age was 54.1 + 10.9yrs(41-77yrs). (3).Healthy controls: There were 46 healthy controls( 27 male and 19 female), the average age was 60.3 ? 11.5yrs(41-80yrs).2. Method.Got the venous biood at 72 hrs and 15 days after the onset, and deteci the PAF. The standard PAF was bought from Sigma company, silica ge G was produced by Shanghai Huamei bioengineering company.PAF was detected by thin-layer chromatography. The 2 ml venous blood was immediately put into the test tube with 3ml Chloroform and 6 ml methyl alcohol, then centrifugation(1200g, 15min), got the supernatant, adding 6ml Chloroform and 9.6 ml methyl alcohol, left in room temperature for 1 hrs, then centrifugation(1200g, 15min), got the Chloroform phase, then water bath(37癈). Prepare the plate using silica ge G and water, baking(120癈,120min)> and cooling to room temperature. Adding samples and standard PAF. Outspreaded them by chloroform : methyl alcohol:water(65:35:6,v/v/v). Cooling it. Mist spray using 10% phosphomolybdate(80*C, 30min) until the blot appearing blue. Scrapingdown the chromatography zone with same Rf value as standard PAP, resolve it by 200ml 0.25%BSA, get rid of the sendimentation, color matching and quantu.Results1. In the ischemic cerebral vasculardisease group, PAP in the acute stage was higher than that of recovery stage. But PAP in the hemorrhagic and non-cerebral vascular disease group had same PAP value in the acute and recovery stages. In the acute stage, there were significant difference among the group of cerebral ischemia, cerebral hemorrhage, non-cerebrovascular disease and healthy control, except between healthy control and non-cerebrovascular disease. In the recovery stage, PAP of cerebral ischemia and cerebral hemorrhage group was higher than that of non-cerebrovascular disease and healthy control group.2. In 7 patients with cerebral ischemia, PAP level was observed dynamically, PAP didn"t changed significantly at the 1-3 days after the onset of disease, and did decrease at 3-28 days after onset, but it was still higher than that of healthy control and non-cerebrovascular disease.ConclusionFrom the results above, PAP level was significantly elevated in cerebral ischmic disease, it was highest at 72 hrs after onset, and could persist for a long period of time. Detection of PAP is helpful for understanding the pathogenesis and diagnosis of stro
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