实验性脑出血大鼠下丘脑组织蛋白质组学研究
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摘要
目的:脑出血(intracerebral hemorrhage ,ICH)是神经系统的常见疾病,它是指脑动脉、静脉或毛细血管破裂所致脑实质内的出血,占急性脑血管病的20%-30%,致残率和病死率均较高,严重危及人类的生命和身体健康。脑出血并发下丘脑植物神经功能紊乱近年来备受国内外研究者的关注,其发生严重影响了患者的预后。较常见的植物神经功能紊乱的症状包括应激性溃疡、中枢性高热以及水电解质紊乱等。应激性溃疡是急性卒中最严重的并发症之一,对其积极预防及治疗在一定程度上可以改善患者的预后。脑出血后合并中枢性高热则常常提示病情危重,死亡率大大增加。下丘脑作为整合中枢和垂体、脑干等其他部分发生纤维联系涉及神经内分泌、体温调节、摄食、饮水、睡眠觉醒周期及生殖等自主功能的调节。由此可见,直接对下丘脑组织进行研究必定会更加直观的揭示ICH后并发上述常见的植物神经功能紊乱的病理生理机制。
蛋白质是基因功能活动的最终执行者,是生命现象复杂性和多样性的直接体现者。蛋白质组学技术的兴起是生命进入后基因组时代的标志。它是从整体的角度研究蛋白质的特征,包括蛋白质的表达水平、翻译后修饰、蛋白与蛋白之间的相互作用等,揭示蛋白质功能与细胞生命现象之间的规律。差异蛋白质组学则是分析不同生物体在不同时刻或者状态下蛋白质表达上的差异,为探索细胞分子机制及寻找治疗药物靶点提供了可能性的依据。
本研究通过对大鼠脑出血模型的制作,对正常下丘脑组织和脑出血72小时下丘脑组织进行差异蛋白质组学研究,找出有差异表达的蛋白质,以期探寻脑出血后并发下丘脑植物神经功能紊乱的病理生理机制,为进一步筛选特异性功能相关蛋白和药物干预靶位提供线索。
方法:
1.制作大鼠脑出血模型,于72小时处死动物,取下丘脑组织。空白对照组取相应部位脑组织。2.提取下丘脑组织的总蛋白,利用双向电泳(2-DE)技术比较大鼠脑出血72小时下丘脑组织和正常对照组下丘脑组织蛋白的差异表达。第一向等电聚焦电泳、第二向垂直SDS-PAGE电泳后,以硝酸银或考马斯亮蓝R250染色。投射扫描双向电泳凝胶图,得到图谱后用Image Master 2D Elite软件进行图像分析。对感兴趣的蛋白点从凝胶上切下,胶内蛋白质酶解。应用基质辅助激光解析/电离飞行时间串联质谱(MALDI-TOF/TOF MS)技术结合数据库检索进行蛋白质功能鉴定及分析。3.以Western blot技术验证了两个蛋白(电压依赖性阴离子选择通道蛋白1和二氢嘧啶酶相关蛋白2)的表达变化。
结果:成功制作大鼠脑出血模型,运用蛋白质组学技术,比较脑出血72小时组与正常对照组的下丘脑组织,发现29种蛋白的表达量发生了变化:上调蛋白14个,包括电压依赖性阴离子选择通道蛋白1,Rad GDP分解抑制剂a,谷氨酰胺合成酶,鸟嘌呤核苷酸结合蛋白G分子亚基β-2,血影蛋白α链,二氢硫辛酸脱氢酶,4-氨基丁酸氨基转移酶,α-互联蛋白,细胞色素(b-c1)复合体亚基1,丙酮酸激酶同功酶M1/M2,泛素蛋白C末端水解酶L1,14-3-3蛋白E,神经丝轻链多肽,琥珀酸脱氢酶(泛醌)黄素蛋白亚基。下调蛋白15个,包括肌酸激酶B型,苹果酸酶,过氧化物酶1,髓鞘碱性蛋白S,磷酸甘油酸变位酶-1,果糖二磷酸醛缩酶C,磷酸丙糖异构酶,3-磷酸甘油醛脱氢酶,二氢嘧啶酶相关蛋白2,丙酮酸脱氢酶(E1)组成亚基β,N,N-二甲基精氨酸二甲氨水解酶,磷脂酰乙醇胺结合蛋白1,α-烯醇化酶,ATP合酶亚基α,谷氨酸脱氢酶1等。
结论: 1.获得了脑出血72小时下丘脑组织和正常空白对照组下丘脑组织的全蛋白表达双向电泳图谱。2.鉴定出的脑出血72小时和正常对照组的下丘脑组织差异表达蛋白涉及细胞骨架蛋白、能量代谢酶类、抗氧化作用的酶、细胞凋亡相关蛋白以及神经递质酶等方面,我们认为它们可能与脑出血后并发下丘脑植物神经功能紊乱的发病机制相关,为进一步筛选特异性功能相关蛋白和药物干预靶位奠定了基础。3.以Western blot技术验证了两个差异表达蛋白(电压依赖性阴离子选择通道蛋白1和二氢嘧啶酶相关蛋白2),为进一步研究此两种蛋白参与的功能网络调控奠定基础。
nervous system diseases,including hemorrhage in parenchyma, which is caused by parenchyma of arteries, veins and capillaries.It takes up 20 percent to 30 percent of the acute cerebral-vascular disorders, with high rates of mortality and disability,severely threatening the quality of life and human health. Recently, hypothalamus vegetative nerve dysfunction,complicated by cerebral hemorrhage,has drawn worldwide attention,since it has a serious effect on the prognosis of ICH patients. Generally,the vegetative nerve dysfunction includes stress ulcer,central hyperpyrexia,fluid electrolyte imbalance, and et al.The stress ulcer is one of the most serious complications of acute stroke.Thus,to a certain degree,effective prevention and treatment will improve the patients′prognosis.And the central hyperpyrexia caused by cerebral hemorrhage always indicates the aggravation of the illness and thus increasing dramatically the mortality rate.Hypothalamus,acting as an integration center, regulates the function of the autonomic nervous system by contacting nerve fibres in hypophysis and brainstem,involving nervous incretion,the regulation of temperature,eating and drinking,the sleep-wakefulness cycle,and reproduction. So the research done on hypothalamus will clarify the pathophysiological mechanisms of the ICH-complicated hypothalamus vegetative nerve dysfunction.
As everyone knows,proteins are the final executor of gene expression,as well as the embodiment of vital complexity and diversity.The technology of proteomics is a sign that life has entered into post-genome era.It studies the feature of the proteins as a whole,including the expression of proteins,the modification of the post-translation of proteins,the interaction between proteins,and et al,in order to explore the rules between protein functions and life.Differential proteomics,a specialized subdivision of the technology,is used to analyze the differences of protein expression in different organisms at different moments and under different states,which can provide the possibility for the research on molecular mechanism in cell and for tracing the treatment drones.
In this paper,in order to find the metabolic proteins and come up with an understanding of the mechanism for ICH-complicated vegetative nerve functional disturbance in hypothalamus,we focused the differential proteomics on normal hypothalamus and abnormal hypothalamus which had suffered from cerebral hemorrhage for 72 hours,by preparing rat models with ICH.In this way,we expected to explore the pathological and physiological mechanisms of ICH-complicated vegetative nerve functional disturbance,and to provide further evidence for screening the disease-specific markers and intervention targets.
Methods:
1.We made models of rat ICH.Kill the rats at 72 hour and got the tissues of the hypothalamus.Got the corresponding part in controls.
2. We extracted the whole tissue proteins, then compared the differences in the protein expressions of normal hypothalamus and abnormal hypothalamus, using 2-dimension electrophoresis(2-DE):isoelectric focusing electrophoresis being the first dimension,and vertical SDS-PAGE electrophoresis being the second dimension.The gels were visualized by colloidal coomassive blue staining and analysed with ImageMaster 2D Elite software.The proteins of interest were in-gel digested and identified,using MALDI-TOF/TOF tandem mass spectrometry.
3. We used Western blot technology to verify the alteration of protein VDAC1 and DPYL2.
Results:
We made models of ICH in rat successfully.Our data showed that the levels of 29 proteins in hypothalamus tissue were significantly altered in ICH group compared with controls.14 up-regulated protein spots were found which were identified as Succinate dehydrogenase [ubiquinone] flavoprotein subunit, Ubiquitin carboxyl-terminal hydrolase isozyme L1,Voltage-dependent anion-selective channel protein 1,14-3-3 protein epsilon,Pyruvate kinase isozymes M1/M2,Cytochrome b-c1 complex subunit 1, Spectrin alpha chain(brain), 4-aminobutyrate aminotransferase(mitochondrial),Dihydrolipoyl dehydrogenase(mitochondrial),Rab GDP dissociation inhibitor alpha, Alpha-internexin,Neurofilament light polypeptide,Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-2,Glutamine synthetase. While 15 down-regulated proteins were identified as Creatine kinase B-type, Malate dehydrogenase(mitochondrial),Peroxiredoxin-2,Myelin basic protein S,Phosp- hoglycerate mutase 1,Fructose-bisphosphate aldolase C,Triosephosphate isomerase,Glyceraldehyde-3-phosphate dehydrogenase,N(G),N(G)-dimethyla- rginine dimethylaminohydrolase 1,Dihydropyrimidinase-related protein 2,Pyr- uvate dehydrogenase E1 component subunitbeta,(mitochondrial),Phosphatidy- lethanolamine-binding protein 1,ATP synthase subunit alpha,Glutamate dehydrogenase 1(mitochondrial), Alpha-enolase.
Conclusion:
1.We got 2-DE maps of the whole hypothalamus tissue proteins from both the ICH group and the control group.
2.The identified proteins involved cytoskeleton proteins, energy metabolic enzymes, antioxidant proteins and neurotransmitter enzyme. We concluded that these proteins may be involved in the pathogenesis of ICH-complicated vegetative nerve functional disturbance, laying the foundation for further screening of the disease-specific markers and intervention targets.
3.We used Western blot technology to verify the alteration of VDAC1 and DPYL2 , which makes it possible to study the participation of these two proteins in function network.
蛋白质是基因功能活动的最终执行者,是生命现象复杂性和多样性的直接体现者。蛋白质组学技术的兴起是生命进入后基因组时代的标志。它是从整体的角度研究蛋白质的特征,包括蛋白质的表达水平、翻译后修饰、蛋白与蛋白之间的相互作用等,揭示蛋白质功能与细胞生命现象之间的规律。差异蛋白质组学则是分析不同生物体在不同时刻或者状态下蛋白质表达上的差异,为探索细胞分子机制及寻找治疗药物靶点提供了可能性的依据。
本研究通过对大鼠脑出血模型的制作,对正常下丘脑组织和脑出血72小时下丘脑组织进行差异蛋白质组学研究,找出有差异表达的蛋白质,以期探寻脑出血后并发下丘脑植物神经功能紊乱的病理生理机制,为进一步筛选特异性功能相关蛋白和药物干预靶位提供线索。
方法:
1.制作大鼠脑出血模型,于72小时处死动物,取下丘脑组织。空白对照组取相应部位脑组织。2.提取下丘脑组织的总蛋白,利用双向电泳(2-DE)技术比较大鼠脑出血72小时下丘脑组织和正常对照组下丘脑组织蛋白的差异表达。第一向等电聚焦电泳、第二向垂直SDS-PAGE电泳后,以硝酸银或考马斯亮蓝R250染色。投射扫描双向电泳凝胶图,得到图谱后用Image Master 2D Elite软件进行图像分析。对感兴趣的蛋白点从凝胶上切下,胶内蛋白质酶解。应用基质辅助激光解析/电离飞行时间串联质谱(MALDI-TOF/TOF MS)技术结合数据库检索进行蛋白质功能鉴定及分析。3.以Western blot技术验证了两个蛋白(电压依赖性阴离子选择通道蛋白1和二氢嘧啶酶相关蛋白2)的表达变化。
结果:成功制作大鼠脑出血模型,运用蛋白质组学技术,比较脑出血72小时组与正常对照组的下丘脑组织,发现29种蛋白的表达量发生了变化:上调蛋白14个,包括电压依赖性阴离子选择通道蛋白1,Rad GDP分解抑制剂a,谷氨酰胺合成酶,鸟嘌呤核苷酸结合蛋白G分子亚基β-2,血影蛋白α链,二氢硫辛酸脱氢酶,4-氨基丁酸氨基转移酶,α-互联蛋白,细胞色素(b-c1)复合体亚基1,丙酮酸激酶同功酶M1/M2,泛素蛋白C末端水解酶L1,14-3-3蛋白E,神经丝轻链多肽,琥珀酸脱氢酶(泛醌)黄素蛋白亚基。下调蛋白15个,包括肌酸激酶B型,苹果酸酶,过氧化物酶1,髓鞘碱性蛋白S,磷酸甘油酸变位酶-1,果糖二磷酸醛缩酶C,磷酸丙糖异构酶,3-磷酸甘油醛脱氢酶,二氢嘧啶酶相关蛋白2,丙酮酸脱氢酶(E1)组成亚基β,N,N-二甲基精氨酸二甲氨水解酶,磷脂酰乙醇胺结合蛋白1,α-烯醇化酶,ATP合酶亚基α,谷氨酸脱氢酶1等。
结论: 1.获得了脑出血72小时下丘脑组织和正常空白对照组下丘脑组织的全蛋白表达双向电泳图谱。2.鉴定出的脑出血72小时和正常对照组的下丘脑组织差异表达蛋白涉及细胞骨架蛋白、能量代谢酶类、抗氧化作用的酶、细胞凋亡相关蛋白以及神经递质酶等方面,我们认为它们可能与脑出血后并发下丘脑植物神经功能紊乱的发病机制相关,为进一步筛选特异性功能相关蛋白和药物干预靶位奠定了基础。3.以Western blot技术验证了两个差异表达蛋白(电压依赖性阴离子选择通道蛋白1和二氢嘧啶酶相关蛋白2),为进一步研究此两种蛋白参与的功能网络调控奠定基础。
nervous system diseases,including hemorrhage in parenchyma, which is caused by parenchyma of arteries, veins and capillaries.It takes up 20 percent to 30 percent of the acute cerebral-vascular disorders, with high rates of mortality and disability,severely threatening the quality of life and human health. Recently, hypothalamus vegetative nerve dysfunction,complicated by cerebral hemorrhage,has drawn worldwide attention,since it has a serious effect on the prognosis of ICH patients. Generally,the vegetative nerve dysfunction includes stress ulcer,central hyperpyrexia,fluid electrolyte imbalance, and et al.The stress ulcer is one of the most serious complications of acute stroke.Thus,to a certain degree,effective prevention and treatment will improve the patients′prognosis.And the central hyperpyrexia caused by cerebral hemorrhage always indicates the aggravation of the illness and thus increasing dramatically the mortality rate.Hypothalamus,acting as an integration center, regulates the function of the autonomic nervous system by contacting nerve fibres in hypophysis and brainstem,involving nervous incretion,the regulation of temperature,eating and drinking,the sleep-wakefulness cycle,and reproduction. So the research done on hypothalamus will clarify the pathophysiological mechanisms of the ICH-complicated hypothalamus vegetative nerve dysfunction.
As everyone knows,proteins are the final executor of gene expression,as well as the embodiment of vital complexity and diversity.The technology of proteomics is a sign that life has entered into post-genome era.It studies the feature of the proteins as a whole,including the expression of proteins,the modification of the post-translation of proteins,the interaction between proteins,and et al,in order to explore the rules between protein functions and life.Differential proteomics,a specialized subdivision of the technology,is used to analyze the differences of protein expression in different organisms at different moments and under different states,which can provide the possibility for the research on molecular mechanism in cell and for tracing the treatment drones.
In this paper,in order to find the metabolic proteins and come up with an understanding of the mechanism for ICH-complicated vegetative nerve functional disturbance in hypothalamus,we focused the differential proteomics on normal hypothalamus and abnormal hypothalamus which had suffered from cerebral hemorrhage for 72 hours,by preparing rat models with ICH.In this way,we expected to explore the pathological and physiological mechanisms of ICH-complicated vegetative nerve functional disturbance,and to provide further evidence for screening the disease-specific markers and intervention targets.
Methods:
1.We made models of rat ICH.Kill the rats at 72 hour and got the tissues of the hypothalamus.Got the corresponding part in controls.
2. We extracted the whole tissue proteins, then compared the differences in the protein expressions of normal hypothalamus and abnormal hypothalamus, using 2-dimension electrophoresis(2-DE):isoelectric focusing electrophoresis being the first dimension,and vertical SDS-PAGE electrophoresis being the second dimension.The gels were visualized by colloidal coomassive blue staining and analysed with ImageMaster 2D Elite software.The proteins of interest were in-gel digested and identified,using MALDI-TOF/TOF tandem mass spectrometry.
3. We used Western blot technology to verify the alteration of protein VDAC1 and DPYL2.
Results:
We made models of ICH in rat successfully.Our data showed that the levels of 29 proteins in hypothalamus tissue were significantly altered in ICH group compared with controls.14 up-regulated protein spots were found which were identified as Succinate dehydrogenase [ubiquinone] flavoprotein subunit, Ubiquitin carboxyl-terminal hydrolase isozyme L1,Voltage-dependent anion-selective channel protein 1,14-3-3 protein epsilon,Pyruvate kinase isozymes M1/M2,Cytochrome b-c1 complex subunit 1, Spectrin alpha chain(brain), 4-aminobutyrate aminotransferase(mitochondrial),Dihydrolipoyl dehydrogenase(mitochondrial),Rab GDP dissociation inhibitor alpha, Alpha-internexin,Neurofilament light polypeptide,Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-2,Glutamine synthetase. While 15 down-regulated proteins were identified as Creatine kinase B-type, Malate dehydrogenase(mitochondrial),Peroxiredoxin-2,Myelin basic protein S,Phosp- hoglycerate mutase 1,Fructose-bisphosphate aldolase C,Triosephosphate isomerase,Glyceraldehyde-3-phosphate dehydrogenase,N(G),N(G)-dimethyla- rginine dimethylaminohydrolase 1,Dihydropyrimidinase-related protein 2,Pyr- uvate dehydrogenase E1 component subunitbeta,(mitochondrial),Phosphatidy- lethanolamine-binding protein 1,ATP synthase subunit alpha,Glutamate dehydrogenase 1(mitochondrial), Alpha-enolase.
Conclusion:
1.We got 2-DE maps of the whole hypothalamus tissue proteins from both the ICH group and the control group.
2.The identified proteins involved cytoskeleton proteins, energy metabolic enzymes, antioxidant proteins and neurotransmitter enzyme. We concluded that these proteins may be involved in the pathogenesis of ICH-complicated vegetative nerve functional disturbance, laying the foundation for further screening of the disease-specific markers and intervention targets.
3.We used Western blot technology to verify the alteration of VDAC1 and DPYL2 , which makes it possible to study the participation of these two proteins in function network.
引文
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25 MurrayRust J,Leiper J,McAlister M,et al.Structural insights into the hydrolysis of cellular nitric-oxide synthase inhibitors by dimethylarginine dimethylaminohydrelase.Nat Struct BIO,2001,8(8):679～683
26 Tran CT,Fox MF,Vallance P,et al.Chromosomal localization,gene structure,and expression pattern of DDAH1;comparison with DDAH2 and implications for evolutionary origins.Genomics,2000,68:101～105
27 Wang S,Paton JF,Kasparov S,et al.The challenge of real-time measurements of nitric oxide release in the brain .Auton Neurosci,2006,126:59～67
28 Simona Renia,Vanessa Palermo, Andrea Guarnera,et al.Swapping of the N-terminus of VDAC1 with VDAC3 restores full activity of the channel and confers anti-aging features to the cell.FEBS Letters, 2010, 584:2837～2844
29 Zaid H, Abu-Hamad S, Israelson A,et al. The voltage-dependent anion channel-1 modulates apoptotic cell death. Cell Death Differ,2005, 12: 751～760
30 Xu X, Forbes JG, Colombini M,et al. Actin modulates the gating of Neurospora crassa VDAC. Membr. Biol, 2001,180:73～81
31 Cnops L,Plas B,Arcken L,et al.Age-dependent expression of collapsing response mediator proteins(CRMPs) in cat visual cortex.Eur.J.Neurosci,2004,19:2345～2351
32 Quinn CC,Gray GE,Hockfield S,et al.A family of proteins implicated in axon guidance and outgrowth.NEUROBIOL,1999,41:158～164
33 Gu Y,Ihara Y,Cunsolo V,et al.Evidence that collapsing response mediator protein-2 is involved in the dynamics of microtubules.J Bio Chem,2000,275:17917～17920
34 Veyrac A, Giannetti N, Charrier E, et al. Expression of collapsin response mediator proteins 1, 2 and 5 is differentially regulated in newly generated and mature neurons of the adult olfactorysystem.Eur.J.Neurosci,2005,21:2635～2648
35 Fukata Y, Itoh TJ, Kimura T, et al. CRMP-2 binds to tubulin heterodimers to promote microtubule assembly. Nat. Cell Biol, 2002,4: 583～591
36 Yoshimura T, Kawano Y, Arimura N,et al.GSK-3b regulates phosphorylation of CRMP-2 and neuronal polarity. Cell,2005,120:137～149
37 Sultana R,Boyd-Kimball D, Poon HF,et al. Redox proteomics identification of oxidized proteins in Alzheimer’s disease hippocampus and cerebellum: An approach to understand pathological and biochemical alterations in AD. Neurobiol. Aging ,2006,27:1564～1576
38 Castegna A, Aksenov M, Thongboonkerd V, et al. Proteomic identification of oxidatively modified proteins in Alzheimer’s disease brain. Part II. Dihydropyrimidinase-related protein 2, alpha-enolase and heat shock cognate 71. Neurochem, 2002,82: 1524～1532
39 Suzuki Y, Nakagomi S, Namikawa K, et al. Collapsin response mediator protein-2 accelerates axon regeneration of nerve-injured motor neurons of rat. Neurochem,2003, 86:1042～1050
40 Weitzdoerfer R, Fountoulakis M, Lubec G,et al. Aberrant expression of dihydropyrimidinase related protein-2, -3, and -4 in fetal Down syndrome brain. Neural Transm Suppl,2001,61:95～107
41 Robert J Ferl, Michael S Manak, Matthew F Reyes,et al. The 14-3-3s. Genome Biol , 2002, 3(7): 3010～3017
42 Lau, Xiaohua Jin, Jie Ren, et al . The 14-3-3τphosphoserine binding protein is required for cardiomyocyte survival. Mol Cell Biol,2007, 27( 4) : 1455～1466
43 Astrid Bruckmann H, Yde Steensma M, Joost Teixeirade Mattos,et al . Regulation of transcription by Saccharomyces cerevisiae 14-3-3 proteins. Biochem J, 2004, 382( 3) : 867～875
44 B Van Everbroeck,S Quoilin, J Boons, et al . A prospective study of CSF markers in 250 patients with possible Creutzf eldtJakob disease. J Neurol Neurosurg Psychiatry, 2003,74(9):1210～1214
45 Elena Garca Martn, P Ayuso,Antonio Luengo, et al .Genetic variability of histamine receptors in patients with Parkinson's disease . BMC Medical Genetics, 2008, 9: 15～21
46 H Tschampa,M Neumann, I Zerr, et al . Patients with Alzheimer's disease and dementia with Lewy bodies mistaken for Creutzfeldt Jakob disease. J Neurol Neurosurg Psychiatry, 2001, 71( 1) : 33～39
47 Mitsuko Hashiguchi, Kazuya Sobue, Hemant K,et al. 14-3-3 is an effector of tau protein phosphorylation. J Biol Chem, 2000,275( 33) : 25247～25254
48 Gjumrakch Aliev, Eldar Gasimov, Mark E Obrenovich, et al .Atherosclerotic lesions and mitochondria DNA deletions in brain microvessels: Implication in the pathogenesis of Alzheimer's disease. Vasc Health Risk Manag, 2008, 4( 3) : 721～730
1 Juvela S,Kase CS,Angela MS,et al.Advances in intracerebral hemorrhage management.Stroke,2006,37(2):301～304
2 Paul T,Rdiger K,Cristina MZ,et al.Classification and pathogenesis of cerebral hemorrhages after thrombolysis in ischemic stroke.Stroke,2006,37(2):556～561
3 Servadei F,Murray GD,TeasdMe GM,et al.Traumatic subarachnoid hemorrhage:demographic and clinical study of 750 patients from the European brain injury consortium survey of head injuries.Neurosurgery,2002,50(2):261～267
4 Udvardy M,Palatka K,Court DA,et al.Acute gastrointestinal bleeding in patients with antithrombotic therapy.Orv Hetil,2002,143(4):183～187
5刘海峰,李武英.重型颅脑损伤后应激性溃疡出血的原因分析与护理方法探讨.山西医药杂志,2010,39(1):94～95
6康凌.脑卒中应激性溃疡的治疗及预防.中西医结合心脑血管病杂志,2009,7(12):1506～1508
7张元仁,秦向华.急性脑卒中与植物神经功能紊乱的临床研究.中医临床研究,2010,1(2):63～65
8郭迎霞.急性脑卒中后脑心综合征104例临床观察.中国实用神经疾病杂志,2010,13(9):81～82
9雷建明,黎朝茂,江先娣.急性脑血管意外并发多器官功能衰竭临床分析.河北医学,2006,12(4):321～323
10陈德昌,刘大为.多器官功能衰竭综合征的诊断与发病机理探讨.中国医学杂志,1988,4(68):22～26
11许贤豪.多发性硬化研究进展.中华神经科杂志,2004,37(1):3～6
12姜剩勇,秦新月,胡梅等.170例多发性硬化患者的临床特征及误诊分析.重庆医学,2009,38(13):1637～1639
13柳真裕,郑金瓯.以勃起障碍为首发症状的多发性硬化2例报告.Guangxi Medical Journal,2006,28(11):1852～1853
14 Padma-Nathan H.Neurologic evaluation of ereclile dysfunction.UrolClin North Am,1988,15(1):77～80
15 Betts CD,Jones SJ,Fowler CG,et a1.Erectile dysfunction in multiple sclerosis:associaled neurological and neurophyiological deficits,and treatment of the condition.Brain,1994,117(6):1303～1310
16 Zesiewicz TA,Baker MJ,Wahba M, et al.Autonomic nervous system dysfunction in Parkinson's disease.Curr Treat Options Neurol,2003,5(2):149～160
17尹伟华,刘春风,罗蔚峰等.帕金森病患者抑郁的相关因素分析.苏州大学学报(医学版),2003,23(1):60～62
18 Bronner G,Royter V,Korczyn AD,et al.Sexual dysfunction in Parkinson's disease.Sex Marital Ther ,2004,30(2):95～105
19何锦照,刘卫彬,李才明等.伴自主神经功能障碍的重症肌无力(附6例报告).Chin J Nerv Dis,2004,30(5):368～370
2林婉丽,龚丽娜,彭莉.亚低温治疗仪治疗脑卒中并发中枢性高热的观察与护理.实用临床医药杂志,2010,14(18):35～36
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22任泽光,吴建中,尹可.高血压ICH后脑血流及自由基变化规律及其与脑水肿的关系.中华神经外科杂志,1995,11(3):209～212
23 Tymianski M,Tator CH,Keinan N,et al. Normal and abnormal calcium homeostasis in neurons:A basis for the pathophysiology of traumatic and ischemic central nervous system injury.Neurosurg, 1996,38:1176～1195
24 Duffy S,Nguyen,Aries VA,et al.A novel GABA-transaminase inhibitor,reduces epileption activity in rat hippocampal Slices.Neuroscience,2004,126:423～432
25 MurrayRust J,Leiper J,McAlister M,et al.Structural insights into the hydrolysis of cellular nitric-oxide synthase inhibitors by dimethylarginine dimethylaminohydrelase.Nat Struct BIO,2001,8(8):679～683
26 Tran CT,Fox MF,Vallance P,et al.Chromosomal localization,gene structure,and expression pattern of DDAH1;comparison with DDAH2 and implications for evolutionary origins.Genomics,2000,68:101～105
27 Wang S,Paton JF,Kasparov S,et al.The challenge of real-time measurements of nitric oxide release in the brain .Auton Neurosci,2006,126:59～67
28 Simona Renia,Vanessa Palermo, Andrea Guarnera,et al.Swapping of the N-terminus of VDAC1 with VDAC3 restores full activity of the channel and confers anti-aging features to the cell.FEBS Letters, 2010, 584:2837～2844
29 Zaid H, Abu-Hamad S, Israelson A,et al. The voltage-dependent anion channel-1 modulates apoptotic cell death. Cell Death Differ,2005, 12: 751～760
30 Xu X, Forbes JG, Colombini M,et al. Actin modulates the gating of Neurospora crassa VDAC. Membr. Biol, 2001,180:73～81
31 Cnops L,Plas B,Arcken L,et al.Age-dependent expression of collapsing response mediator proteins(CRMPs) in cat visual cortex.Eur.J.Neurosci,2004,19:2345～2351
32 Quinn CC,Gray GE,Hockfield S,et al.A family of proteins implicated in axon guidance and outgrowth.NEUROBIOL,1999,41:158～164
33 Gu Y,Ihara Y,Cunsolo V,et al.Evidence that collapsing response mediator protein-2 is involved in the dynamics of microtubules.J Bio Chem,2000,275:17917～17920
34 Veyrac A, Giannetti N, Charrier E, et al. Expression of collapsin response mediator proteins 1, 2 and 5 is differentially regulated in newly generated and mature neurons of the adult olfactorysystem.Eur.J.Neurosci,2005,21:2635～2648
35 Fukata Y, Itoh TJ, Kimura T, et al. CRMP-2 binds to tubulin heterodimers to promote microtubule assembly. Nat. Cell Biol, 2002,4: 583～591
36 Yoshimura T, Kawano Y, Arimura N,et al.GSK-3b regulates phosphorylation of CRMP-2 and neuronal polarity. Cell,2005,120:137～149
37 Sultana R,Boyd-Kimball D, Poon HF,et al. Redox proteomics identification of oxidized proteins in Alzheimer’s disease hippocampus and cerebellum: An approach to understand pathological and biochemical alterations in AD. Neurobiol. Aging ,2006,27:1564～1576
38 Castegna A, Aksenov M, Thongboonkerd V, et al. Proteomic identification of oxidatively modified proteins in Alzheimer’s disease brain. Part II. Dihydropyrimidinase-related protein 2, alpha-enolase and heat shock cognate 71. Neurochem, 2002,82: 1524～1532
39 Suzuki Y, Nakagomi S, Namikawa K, et al. Collapsin response mediator protein-2 accelerates axon regeneration of nerve-injured motor neurons of rat. Neurochem,2003, 86:1042～1050
40 Weitzdoerfer R, Fountoulakis M, Lubec G,et al. Aberrant expression of dihydropyrimidinase related protein-2, -3, and -4 in fetal Down syndrome brain. Neural Transm Suppl,2001,61:95～107
41 Robert J Ferl, Michael S Manak, Matthew F Reyes,et al. The 14-3-3s. Genome Biol , 2002, 3(7): 3010～3017
42 Lau, Xiaohua Jin, Jie Ren, et al . The 14-3-3τphosphoserine binding protein is required for cardiomyocyte survival. Mol Cell Biol,2007, 27( 4) : 1455～1466
43 Astrid Bruckmann H, Yde Steensma M, Joost Teixeirade Mattos,et al . Regulation of transcription by Saccharomyces cerevisiae 14-3-3 proteins. Biochem J, 2004, 382( 3) : 867～875
44 B Van Everbroeck,S Quoilin, J Boons, et al . A prospective study of CSF markers in 250 patients with possible Creutzf eldtJakob disease. J Neurol Neurosurg Psychiatry, 2003,74(9):1210～1214
45 Elena Garca Martn, P Ayuso,Antonio Luengo, et al .Genetic variability of histamine receptors in patients with Parkinson's disease . BMC Medical Genetics, 2008, 9: 15～21
46 H Tschampa,M Neumann, I Zerr, et al . Patients with Alzheimer's disease and dementia with Lewy bodies mistaken for Creutzfeldt Jakob disease. J Neurol Neurosurg Psychiatry, 2001, 71( 1) : 33～39
47 Mitsuko Hashiguchi, Kazuya Sobue, Hemant K,et al. 14-3-3 is an effector of tau protein phosphorylation. J Biol Chem, 2000,275( 33) : 25247～25254
48 Gjumrakch Aliev, Eldar Gasimov, Mark E Obrenovich, et al .Atherosclerotic lesions and mitochondria DNA deletions in brain microvessels: Implication in the pathogenesis of Alzheimer's disease. Vasc Health Risk Manag, 2008, 4( 3) : 721～730
1 Juvela S,Kase CS,Angela MS,et al.Advances in intracerebral hemorrhage management.Stroke,2006,37(2):301～304
2 Paul T,Rdiger K,Cristina MZ,et al.Classification and pathogenesis of cerebral hemorrhages after thrombolysis in ischemic stroke.Stroke,2006,37(2):556～561
3 Servadei F,Murray GD,TeasdMe GM,et al.Traumatic subarachnoid hemorrhage:demographic and clinical study of 750 patients from the European brain injury consortium survey of head injuries.Neurosurgery,2002,50(2):261～267
4 Udvardy M,Palatka K,Court DA,et al.Acute gastrointestinal bleeding in patients with antithrombotic therapy.Orv Hetil,2002,143(4):183～187
5刘海峰,李武英.重型颅脑损伤后应激性溃疡出血的原因分析与护理方法探讨.山西医药杂志,2010,39(1):94～95
6康凌.脑卒中应激性溃疡的治疗及预防.中西医结合心脑血管病杂志,2009,7(12):1506～1508
7张元仁,秦向华.急性脑卒中与植物神经功能紊乱的临床研究.中医临床研究,2010,1(2):63～65
8郭迎霞.急性脑卒中后脑心综合征104例临床观察.中国实用神经疾病杂志,2010,13(9):81～82
9雷建明,黎朝茂,江先娣.急性脑血管意外并发多器官功能衰竭临床分析.河北医学,2006,12(4):321～323
10陈德昌,刘大为.多器官功能衰竭综合征的诊断与发病机理探讨.中国医学杂志,1988,4(68):22～26
11许贤豪.多发性硬化研究进展.中华神经科杂志,2004,37(1):3～6
12姜剩勇,秦新月,胡梅等.170例多发性硬化患者的临床特征及误诊分析.重庆医学,2009,38(13):1637～1639
13柳真裕,郑金瓯.以勃起障碍为首发症状的多发性硬化2例报告.Guangxi Medical Journal,2006,28(11):1852～1853
14 Padma-Nathan H.Neurologic evaluation of ereclile dysfunction.UrolClin North Am,1988,15(1):77～80
15 Betts CD,Jones SJ,Fowler CG,et a1.Erectile dysfunction in multiple sclerosis:associaled neurological and neurophyiological deficits,and treatment of the condition.Brain,1994,117(6):1303～1310
16 Zesiewicz TA,Baker MJ,Wahba M, et al.Autonomic nervous system dysfunction in Parkinson's disease.Curr Treat Options Neurol,2003,5(2):149～160
17尹伟华,刘春风,罗蔚峰等.帕金森病患者抑郁的相关因素分析.苏州大学学报(医学版),2003,23(1):60～62
18 Bronner G,Royter V,Korczyn AD,et al.Sexual dysfunction in Parkinson's disease.Sex Marital Ther ,2004,30(2):95～105
19何锦照,刘卫彬,李才明等.伴自主神经功能障碍的重症肌无力(附6例报告).Chin J Nerv Dis,2004,30(5):368～370