针刺治疗脑缺血再灌注损伤：与减轻炎症反应有关？

摘要 背景：脑缺血己成为严重影响人类健康的主要疾病之一。针刺治疗急性脑缺血的疗效是公认的但其治疗机制不明。目的：探讨脑缺血状态下核转录因子-kB（NF-kB）、细胞间黏附分子-1 (ICAM-1)、血管细胞黏附分子-1（VCAM-1）的变化及针刺的干预作用。 设计及时间：分子生物学实验于2008年12月至2009年10月在中南大学湘雅医院神经外科实验室完成。 研究对象/材料/对象：健康SD大鼠46只随机分成假手术组、脑缺血组、针刺预处理组,针刺干预组。方法：以线栓法制作大鼠大脑中动脉缺血2h再灌注模型,并用针刺干预,然后应用RT-PCR逆转录反应、蛋白免疫印迹法分析观察各组大鼠脑组织NF-kB ,ICAM-1、VCAM-1 mRNA及蛋白的表达。结果：脑缺血大鼠动物模型被成功复制,其神经功能评分表明,针刺明显改善上述指标,尤其对再灌注70h组的影响显著;针刺组脑缺血再灌注后NF-kB、ICAM-1、 VCAM-1mRNA及蛋白质的表达明显下降。结论：针刺可明显改善脑缺血再灌注后的神经功能评分指标,并且该作用可能与抑制脑缺血再灌注后NF-kB、ICAM-1、VCAM-1的表达,下调脑缺血区域组织的NF-kB、ICAM-1、VCAM-1 mRNA及蛋白质表达水平有关。     

炎症信号受体TLR4在脑出血大鼠血肿周围脑组织的表达

目的探讨TLR4在脑出血大鼠血肿周围脑组织中的表达变化及其作用。方法采用自体尾动脉血注入法制作脑出血大鼠模型,采用免疫组织化学方法观察大鼠脑组织TLR4蛋白表达,反转录多聚酶链反应(RT-PCR)方法检测大鼠脑组织TLR4mRNA的相对表达。结果脑出血大鼠脑组织TLR4免疫阳性产物较对照组明显增加(P<0.01),同时伴有脑组织TLR4mRNA相对表达上调(P<0.05)。结论脑出血大鼠模型TLR4表达增高,提示TLR4在脑出血炎症损伤中具有一定作用。     

老年脑白质疏松合并高血压对患者认知功能的影响

目的探讨老年脑白质疏松(LA)合并高血压对患者认知功能的影响。方法选取2014年1月~2015年1月期间于我院神经内科门诊就诊或病房收治的42例老年LA合并高血压患者为研究对象,归为老年LA伴高血压组,并选择同期40例无脑白质病变的单纯老年高血压患者归为老年高血压组,以及来我院体检的30例健康老年人作为老年健康对照组。以蒙特利尔认知评估量表(Mo CA)评估3组的认知功能。结果老年LA伴高血压组的视空间与执行功能、命名、注意力、抽象及延迟回忆的评分及总分均显著低于老年健康对照组,差异均有统计学意义(P0.05)。且视空间与执行功能、抽象的评分及总分均显著低于老年高血压组,差异有统计学意义(P0.05)。老年LA伴高血压组的认知功能损害发生率显著高于老年高血压组及对照组,差异均有统计学意义(P0.05)。结论与老年高血压和健康老人相比,老年LA合并高血压患者的认知功能明显减退,认知功能障碍率明显升高,故需要对这些老年患者定期检测认知功能,对认知功能障碍进行早期干预。     

高血压脑出血患者急性期血肿周围组织的炎症反应特征

目的探讨高血压脑出血患者血肿周围组织的炎症反应特征。方法29例基底节区脑出血患者行微创血肿清除术,按出血后手术时间分为6—24h组、24～48h组及〉48h组,将手术中引流的血肿周围脑组织行白细胞共同抗原（LCA）和细胞间黏附分子-1（ICAM-1）免疫组化染色,并与对照组（非脑病尸检患者6例）进行比较。结果脑出血组脑组织标本免疫组化染色可见LCA、ICAM-1免疫阳性微血管及细胞,与对照组相比,表达量显著提高（均P〈0．01）;各时段组间LCA的表达差异无统计学意义;ICAM-1的表达24～48h组及〉48h组显著高于6—24h组（均P〈0．01）,24～48h组与〉48h组间差异无统计学意义。结论脑出血急性期血肿周围脑组织存在急性炎症反应;ICAM-1的表达在出血后48h内逐渐增多。     

脑白质疏松与胰岛素抵抗和单核细胞趋化蛋白-1的关系

目的探讨脑白质疏松患者胰岛素抵抗(IR)与血浆单核细胞趋化蛋白-1(MCP-1)水平的关系。方法选择48例脑白质疏松患者,以30例健康者作为对照组。采用放射免疫法测定两组的空腹血胰岛素(FINS)水平,同时检测空腹血糖(FBG)、血脂,以稳态模型评估法(HOMA)评价胰岛素抵抗(HOMA-IR)。采用ELISA法测定血清MCP-1水平。结果脑白质疏松组的MCP-1水平高于健康对照组(P<0.01);脑白质疏松组FINS、HOMA-IR均高于健康对照组(P<0.01)。相关性分析显示脑白质疏松患者血清MCP-1水平与FPG、TC、LDL-C、血清胰岛素、HOMA-IR呈明显正相关(P<0.01)。结论脑白质疏松患者存在胰岛素抵抗,MCP-1与胰岛素抵抗密切相关。     

肝细胞生长因子对自发性高血压大鼠血压的影响

背景：肝细胞生长因子是一种多功能生长因子，它能促进多种细胞生长与移行及各种组织器官的发生。在心血管系统，它具有抗凋亡、抗纤维化、促进内皮细胞损伤后修复作用，推测其可能具有降压效应。 目的：观察外源性肝细胞生长因子对自发性高血压大鼠血压、血管内皮系统和肾素-血管紧张素系统的影响并探讨其调节血压的可能机制。 设计、时间及地点：随机对照动物实验，于2007-03/07在安徽医科大学第一附属医院心内科完成。 材料：外源性肝细胞生长因子粉剂购于美国Peprotech公司，成年自发性高血压大鼠组和WKY大鼠，均14周龄，体质量200~ 250 g。自发性高血压大鼠随机分为实验组和单纯自发性高血压大鼠组，WKY大鼠为正常对照组，每组12只。 方法：实验组每间隔24 h从尾静脉依次给予肝细胞生长因子5，10，15，20，25 μg/kg，自发性高血压大鼠组和正常对照组同时给予等量生理盐水。每次注射安抚大鼠5 min后测血压与心率，最后一次注射后观察血压降至最低值时(约注射后30 min)，麻醉后处死，各取右心室血2 mL。 主要观察指标：①观察肝细胞生长因子对自发性高血压大鼠组收缩压及心率的影响。②分别用比色法测血清一氧化氮水平、放射免疫法测血浆内皮素、血管紧张素Ⅱ水平。 结果：实验组注射肝细胞生长因子5 μg/kg血压下降不明显，注射10 μg/kg约5 min后血压开始下降，30 min降至最低，1 h后血压开始逐渐回升，5 h后血压基本回到原先水平。注射20 μg/kg达最大降压幅度，收缩压下降达40~50 mm Hg，再增加剂量最大降压幅度及持续时间不变。整过程心率无明显变化。两个对照组血压无明显变化。实验组较自发性高血压大鼠组内皮素、血管紧张素Ⅱ含量下降，一氧化氮含量上升（P < 0.05）。 结论：从静脉给予外源性肝细胞生长因子能快速降低自发性高血压大鼠组的血压，在一定剂量范围内，呈剂量-效应与时间-效应关系。肝细胞生长因子系统、血管内皮系统、肾素-血管紧张素系统可能共同参与血压的调节。     

脑白质疏松患者血浆单核细胞趋化蛋白-1和组织因子的变化

目的探讨脑白质疏松患者血浆单核细胞趋化蛋白-1和组织因子的变化及临床意义。方法对56例脑白质疏松患者,56例脑梗死患者和30例健康对照组应用酶联免疫吸附双抗体夹心法（ELISA法）定量测定血浆单核细胞趋化蛋白-1和组织因子的水平。结果脑梗死和正常对照组相比,脑白质疏松组血浆单核细胞趋化蛋白-1含量（263.76±92.36）pg/ml明显高于脑梗死组（128.49±76.97）pg/ml和对照组（112.11±60.36）pg/ml统计学处理差异具有显著性（P〈0.01）。随脑白质疏松程度的加重,血浆单核细胞趋化蛋白-1的含量越高。结论脑白质疏松患者血浆单核细胞趋化蛋白-1和组织因子的水平明显升高,并且与脑白质疏松的程度密切相关。     

氯沙坦对卒中易感型自发性高血压大鼠的脑保护作用

目的 研究氯沙坦对卒中易感型自发性高血压大鼠(SHRsp)脑保护作用的机制。方法 6周龄雄性SHRsp随机分为生理盐水组、小剂量氯沙坦组(10mg·kg-1·d-1)和大剂量氯沙坦组(30mg·kg-1·d-1),记录血压和脑卒中临床表现评分,18周后处死,光镜观察脑卒中的发生率和脑血管结构;电镜观察脑组织的超微结构;TUNEL法检测神经细胞凋亡。结果 大剂量氯沙坦组血压明显低于未用药组(PO.05)。对照组SHRsp死亡率、脑动脉中膜厚度/管腔半径的比值和神经细胞凋亡率均高于小剂量和大剂量氯沙坦组,差异均有显著性意义(P相似文献   

Toll样受体信号通路与缺血性中风炎症反应机制的研究进展

Toll样受体既是机体天然免疫信号受体,同时也是炎症信号受体,在脑缺血发生后通过MyD88依赖性途径和TRIF依赖性途径参与炎症级联反应。阻断或者激活相应的Toll样受体信号传导通路,可有效抑制脑缺血急性期炎症级联反应的发生、发展,减轻炎性损伤。干预Toll样受体信号传导通路是脑缺血后抗炎治疗的途径之一。     

白芍总甙对局灶性脑缺血大鼠脑组织炎症信号表达的影响

目的观察白芍总甙对局灶性脑缺血(Focal Cerebral ischemic,FCI)大鼠脑组织Toll样受体4(TLR4)基因表达的作用。方法参照longa等方法制成大鼠局灶性大脑中动脉梗死(MCAO)模型,将80只SD雄性大鼠随机分为假手术组,溶剂组,白芍总甙低、高两个剂量组,采用RT-PCR法检测缺血灶周围脑组织TLR4基因表达水平。结果生理盐水组TLR4基因相对表达均明显高于假手术组(P<0.01);白芍总甙中高剂量组TLR4相对表达量明显低于生理盐水组(P<0.01)。结论生理盐水组大鼠缺血灶周围脑组织TLR4基因高表达,白芍总甙对TLR4表达有抑制作用,这可能是其抗炎作用的机理之一。     

Morphological changes in spontaneously hypertensive rats

Summary Our previous studies of angiotension-induced acute hypertension showed increased intracerebral arteriolar permeability associated with markedly enhanced pinocytosis. This study was performed to determine whether similar findings occurred in spontaneous non-pharmacologically induced chronic hypertension.Cerebrovascular permeability to horseradish peroxidase (HRP) was studied over an 82-week period in spontaneously hypertensive rats (SHR) derived from a strain that originated from Japan. In a few animals increased cerebrovascular permeability to HRP was observed, associated with enhanced pinocytosis. Quantitatively, the number of pinocytotic vesicles in permeable arteriolar segments was significantly increased suggesting that enhanced pinocytosis is the principal mechanism of early cerebrovascular changes in SHR.Light microscopy of renal, ocular and cerebral vessels revealed medial hyperplasia affecting renal vessels at 16 weeks and occurring later in ocular and cerebral vessels. Deposition of fibrin in renal vessels was observed from 64 weeks onwards but was not associated with renal failure.This work was supported by the Medical Research Council of Canada, Grant No. MT 1647     

Effect of treatment with choline alphoscerate on hippocampus microanatomy and glial reaction in spontaneously hypertensive rats

The influence of long term treatment with choline alphoscerate on microanatomy of hippocampus and glial reaction was assessed in spontaneously hypertensive rats (SHR) used as an animal model of cerebrovascular disease. Choline alphoscerate is a cholinergic precursor, which has shown to be effective in countering cognitive symptoms in forms of dementia disorders of degenerative, vascular or combined origin. Male spontaneously hypertensive rats (SHR) aged 6 months and age-matched normotensive Wistar-Kyoto (WKY) rats were treated for 8 weeks with an oral daily dose of 100 mg/kg of choline alphoscerate, 285 mg/kg of phosphatidylcholine (lecithin) or vehicle. On the hippocampus of different animal groups, nerve cell number and GFAP-immunoreactive astrocytes were assessed by neuroanatomical, immunochemical and immunohistochemical techniques associated with quantitative analysis. Treatment with choline alphoscerate countered nerve cell loss and glial reaction primarily in the CA1 subfields and in the dentate gyrus of the hippocampus of SHR. Phosphatidylcholine did not affect hypertension-dependent changes in hippocampal microanatomy. Both compounds did not affect blood pressure values in SHR. These data suggest that choline alphoscerate may play a role in the countering hippocampal changes induced by cerebrovascular involvement. The observation that treatment with choline alphoscerate attenuates the extent of glial reaction in the hippocampus of SHR suggests also that the compound may afford neuroprotection in this animal model of vascular brain damage.     

Cerebroventricular dilation in spontaneously hypertensive rats (SHRs) is not attenuated by reduction of blood pressure

In previous studies, we found that spontaneously hypertensive rats (Okamoto-Aoki SHRs) suffer progressive postnatal dilation of the brain ventricles. In the present study we examined intracerebroventricular pressure and blood pressure as possible mechanisms of ventricular dilation in SHRs. We found that intracerebroventricular pressure was not elevated in SHRs. The role of blood pressure was examined in SHRs treated chronically with the antihypertensive drug, captopril, beginning in utero, and in renal hypertensive Sprague-Dawley rats (SDs). Although our experimental treatments produced significant changes in mean arterial pressures, they did not alter brain ventricular size: SDs with experimental hypertension had normal-sized brain ventricles and SHRs with pharmacologically reduced blood pressure had enlarged ventricles. These results suggest that neither increased intraventricular pressure nor high blood pressure is the sole cause of hydrocephalus in SHRs.     

Renin antisense injected intraventricularly decreases blood pressure in spontaneously hypertensive rats

Brain renin-angiotensin system plays an important role in blood pressure regulation and is suggested to play a role in the development and maintenance of hypertension. To test the hypothesis that brain renin may play a significant role in hypertension in spontaneously hypertensive rats (SHR), phosphorothioated antisense oligodeoxynucleotides targeted to renin mRNA were administered intracerebroventricularly in SHR. Administration of an antisense but not its sense oligodeoxynucleotide produced a prolonged duration of decrease in blood pressure. Intra-arterial administration of the antisense oligodeoxynucleotide at the same dose that decreased blood pressure when administered intraventricularly did not affect blood pressure. Furthermore, renin mRNA but not angiotensin AT1 receptor mRNA levels were decreased in the hypothalamus of the antisense oligodeoxynucleotide-treated rats. These results suggest that brain renin may play a significant role in hypertension in SHR.     

An angiotensin II type 1 receptor blocker can preserve endothelial function and attenuate brain ischemic damage in spontaneously hypertensive rats

Hypertension reduces endothelial nitric oxide synthase (eNOS) expression and leads to endothelial dysfunction. However, few studies have demonstrated the influences of hypertension on eNOS function in the cerebral cortex. The present study investigates the influences of hypertension on endothelial function in the cerebral cortex and the protective effects of antihypertensive agents against brain ischemia through the preservation of endothelial function. Five‐ and ten‐week‐old male Wistar rats and spontaneously hypertensive rats (SHR) were used for experiments. Five‐week‐old SHR received olmesartan, hydralazine, or vehicle for 5 weeks in drinking water. eNOS activation in the cerebral cortex was evaluated by analyzing levels of total and Ser¹¹⁷⁷‐phosphorylated eNOS protein by Western blot. Blood pressure of 10‐week‐old SHR without treatment was clearly high, and the ratio of phospho‐eNOS/total eNOS protein was significantly low. Five‐week treatment with olmesartan or hydralazine suppressed the elevation of blood pressure and the reduction of phosphorylated eNOS‐Ser¹¹⁷⁷ in SHR, and olmesartan was more effective in maintaining phosphorylation of eNOS‐Ser¹¹⁷⁷ than hydralazine. To assess the contribution of eNOS to maintaining cerebral blood flow (CBF), we monitored CBF by laser‐Doppler flowmetry after L‐N⁵‐(1‐iminoethyl)ornithine (L‐NIO) infusion. CBF response to L‐NIO was preserved in olmesartan‐treated SHR but not in hydralazine‐treated SHR. Furthermore, infarct volume 48 hr after transient focal brain ischemia in olmesartan‐treated SHR was significantly reduced compared with vehicle‐treated SHR. These findings indicate that chronic prehypertensive treatment with olmesartan could attenuate brain ischemic injury through the maintenance of endothelial function in the cerebral cortex in SHR. © 2010 Wiley‐Liss, Inc.     

Autoradiographic comparison of muscarinic M1 and M2 binding sites in the CNS of spontaneously hypertensive and normotensive rats

Spontaneously hypertensive rats (SHR) respond with exaggerated pressor responses of central origin in response to pharmacologic stimulation of brain muscarinic receptors when compared with those to normotensive Wistar Kyoto (WKY) rats. At least part of the enhanced response to central muscarinic stimulation may be due to alterations in the expression of one or more of the five subtypes of muscarinic receptors. SHR are also known to exhibit regional alterations in the levels of mRNA encoding the M1, M2 and M4 receptors. In this study, we estimated the number of specific muscarinic receptor binding sites in 12-week-old SHR and WKY by measuring the binding of M1- and M2-selective ligands. Using standard autoradiographic techniques, coronal sections obtained from 12-week-old SHR and WKY were incubated with [³H]pirenzepine or [³H]AFDX 384 to label M1 and M2 receptors, respectively. Although both strains exhibited similar distribution patterns for both binding sites, sections derived from SHR expressed a significant increase in the number of [³H]pirenzepine binding sites compared to normotensive WKY in caudate putamen, CA3 region of the hippocampus, cingulate cortex, substantia nigra, posterior hypothalamic area and tuberomammillary nucleus. An increased number of [³H]AFDX 384 binding sites in SHR were observed in the olfactory tubercle, nucleus accumbens, basolateral amygdaloid nucleus, rostroventrolateral medulla and nucleus paragigantocellularis. Decreases in the number of [³H]AFDX 384 binding sites in SHR were also observed in the parietal cortex, medial geniculate, and lateral hypothalamic area. Statistically significant site-selective differences in binding densities between strains ranged from 4.0% to 35.5% of WKY means. These alterations in the expression of M1 and M2 binding sites in cardiovascular regions may contribute to the strain's hyper-responsiveness to cholinergic drugs and possibly to the appearance of other autonomic or behavioral phenotypes exhibited by SHR, including the hypertensive state itself.     

Chronic unpredictable stress accelerates atherosclerosis through promoting inflammation in apolipoprotein E knockout mice

Introduction

Chronic unpredictable stress (CUS) has been suggested to accelerate atherosclerosis. However, the underlying mechanism of this adverse effect is not fully understood. Since chronic stress can promote or even initiate inflammation response, which is thought to be a major contributor to atherogenesis, we postulated that stress-induced inflammatory response might be one important reason for CUS-promoted atherosclerotic disease.

Materials and methods

We used the CUS treated apolipoprotein E (ApoE)-deficient mice, which have been shown to spontaneously develop atherosclerosis with features similar to those seen in humans, as an animal model. Haematoxylin and eosin staining and immunohistostaining were used to analyze the plaque formation and composition.

Results

Histological analysis clearly demonstrated that CUS treatment promoted the development of atherosclerotic lesions, such as triggering plaque rupture, increasing plaque size and plaque-to-surface ratio, and also led to profound changes in plaque composition, as evidenced by increased macrophage and T cell infiltration and decreased smooth muscle cell mass, all reflecting an unstable plaque phenotype. Moreover, adhesion molecular vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1), acute phase reactant C-reactive protein (CRP), and proinflammatory cytokine interleukin-6 (IL-6) were significantly enhanced in CUS treated ApoE^-/- mice compared with untreated control animals (P < 0.01).

Conclusion

The involvement of CUS in the pathogenesis of atherosclerosis is at least partially attributable to its acceleration of inflammation.     

小檗碱对颈动脉粥样硬化家兔NF—kB和VCAM-1表达的影响

目的探讨小檗碱预防颈动脉粥样硬化形成的作用机制。方法将24只雄性新西兰大白兔随机分为3组:正常组、颈动脉粥样硬化组(模型组)、小檗碱预防组(小檗碱组),每组各8只。正常组予普通饮食5周;模型组和小檗碱组给予高脂饲料喂养1周后行右侧颈动脉内膜空气干燥术,术后继续高脂饲料喂养4周;小檗碱组在给予高脂饲料喂养的同时每日灌服小檗碱。第5周取右侧颈动脉组织行HE染色观察颈动脉病理改变、行核因子-κB(Nuclear factor kappa B,NF-κB)免疫组织化学染色半定量检测NF-κB的活性,RT-PCR法检测NF-κBp65-mRNA、VCAM-1 mRNA。结果模型组颈动脉有明显的动脉粥样硬化斑块形成;小檗碱组的内膜轻度增厚,内膜下有少量泡沫细胞形成。小檗碱组NF-κBp65阳性细胞数密度高于正常组(P<0.01),明显低于模型组(P<0.01)。小檗碱组NF-κBp65/β-actin、VCAM-1/β-actin扩增带吸光度值比值高于正常组(0.42±0.05比0.18±0.04;0.61±0.11比0.20±0.04,P<0.01),而明显低模型组(0.42±0.05比0.66±0.10;0.61±0.11比0.81±0.11,P<0.01)。NF-кB活性与VCAM-1呈显著正相关(r=0.926P<0.01)。结论小檗碱预防家兔颈动脉粥样硬化可能通过抑制NF-κB活性,降低VCAM-1表达而起作用。