缓激肽及其类似物RMP-7对血脑屏障通透性的影响

缓激肽及其类似物RMP-7能选择性增加血肿瘤屏障的通透性,而对正常脑组织的血脑屏障几乎无影响。研究表明,缓激肽选择性开放血脑屏障是由于缓激肽与大脑毛细血管内皮细胞膜上的B2受体结合,引起细胞内Ca2+水平升高,从而引发包括NO和cGMP在内的一系列的信号传递反应,使胞饮小泡数量增加或紧密连接开放。本文就缓激肽及其类似物RMP-7选择性开放血脑屏障的新进展做一综述。     

缓激肽选择性增加局部脑缺血大鼠血脑屏障的通透性

目的　研究颈动脉灌注小剂量缓激肽对缺血后血脑屏障通透性的影响及机制。方法　免疫组化分析正常脑组织的缓激肽B2受体所在。大鼠大脑中动脉结扎 1h或 2h再灌流 1h。用放射自显影方法检测缓激肽对血脑屏障通透性的变化。WesternBlot方法检测bNOS ,iNOS和B2受体。NOS检测盒检测NOS的活性。结果　正常脑组织毛细血管内皮未见B2受体表达 ,在神经细胞上发现B2受体的表达。缺血 2h再灌流1h缓激肽灌注缺血区血脑屏障通透性显著增加。WesternBlot结果提示 ,在缓激肽灌注组和对照组间 ,缺血皮质区bNOS和B2受体没有明显变化 ,各组中均未检测出iNOS。缓激肽灌注组的NOS活性显著高于对照组。结论　正常脑组织毛细血管内皮未表达B2受体 ,神经细胞上可见B2受体的表达。灌注小剂量缓激肽能选择性增加局部脑缺血大鼠血脑屏障的通透性     

缓激肽对脑胶质瘤大鼠紧密连接影响的形态学观察

目的研究缓激肽(BK)对脑胶质瘤大鼠血肿瘤屏障紧密连接的影响。方法采用伊文氏兰(EB)法检测缓激肽作用后血肿瘤屏障(BTB)通透性的变化;应用透射电镜(TEM)观察BK作用后内皮细胞间紧密连接的变化,同时应用硝酸镧[La(NO3)3]和辣根过氧化物酶(HRP)作示踪剂,检测缓激肽作用后,小分子和大分子示踪剂通过紧密连接的情况。结果缓激肽可使血肿瘤屏障对伊文氏兰的通透性增加,在15min时达到高峰,以后逐渐下降。透射电镜显示缓激肽作用15min时,肿瘤组织毛细血管内皮细胞间紧密连接的完整性明显破坏,缝隙指数显著增加,同时可见硝酸镧和辣根过氧化物酶在紧密连接处沉积。结论缓激肽能够通过开放紧密连接选择性增加血肿瘤屏障的通透性。     

ATP敏感性钾通道蛋白在缓激肽选择性开放血肿瘤屏障中的表达

目的 探讨ATP敏感性钾通道蛋白在缓激肽选择性开放血肿瘤屏障中的作用.方法 建立大鼠脑胶质瘤模型, 颈内动脉灌注缓激肽后,采用免疫组化SABC法和 Western blot法测定肿瘤组织ATP敏感性钾通道蛋白的功能亚基Kir6.2的分布和表达的变化.结果 脑胶质瘤大鼠经颈内动脉灌注缓激肽后,其肿瘤内血管内皮细胞的 Kir6.2蛋白表达比对照组显著增多, 且以灌注后10 min 增加最为显著. 结论 ATP敏感性钾通道蛋白表达上调可能是缓激肽选择性开放血肿瘤屏障的重要机制之一.     

血管内皮细胞生长因子与实体肿瘤

血管内皮细胞生长因子（ＶＥＧＦ）具有选择性促血管内皮细胞生长和增强血管渗透性的作用，在多种实体肿瘤中有高水平合成，并与肿瘤的分级和转移相关，ＶＥＧＦ受体主要表达于血管内皮细胞，通过干预ＶＥＧＦ及其受体抑制肿瘤血管形成是阻遏肿瘤生长和转移的重要策略     

ROCK介导缓激肽开放SD大鼠血肿瘤屏障

目的利用ROCK的特异性抑制剂Y-27632,研究ROCK是否介导缓激肽开放血肿瘤屏障。方法应用ROCK的特异性抑制剂Y-27632预处理大鼠原代脑微血管内皮细胞后,用缓激肽诱导血肿瘤屏障开放,测量跨内皮阻抗值(TEER),辣根过氧化物酶(HRP)渗漏量,分析血肿瘤屏障的通透性的改变;应用Western-blot法检测紧密连接相关蛋白ZO-1的表达;应用免疫荧光方法观察原代大鼠脑微血管内皮细胞紧密连接相关蛋白ZO-1和丝状肌动蛋白结构和分布的改变。结果 Y-27632显著抑制缓激肽诱导TEER值的降低,HRP的升高;Y-27632显著抑制ZO-1的表达;Y-27632抑制ZO-1由内皮细胞的边缘向细胞质转移,抑制丝状肌动蛋白由细胞膜边缘向细胞中央区分布,应力纤维形成明显减少。结论 ROCK介导缓激肽开放血肿瘤屏障。     

低频超声开放大鼠血脑肿瘤屏障与TNF-α表达之间的关系

目的探讨肿瘤坏死因子-α(TNF-α)与低频超声开放大鼠血脑肿瘤屏障之间的关系。方法应用1MHz低频超声辐照C6胶质瘤大鼠,采用伊文思蓝(EB)法检测血脑肿瘤屏障通透性的变化,应用透射电镜观察脑微血管内皮细胞紧密连接的变化,应用酶联免疫吸附试验(ELISA)法检测脑组织中TNF-α含量的变化。结果低频超声辐照后大鼠血脑肿瘤屏障的通透性逐渐增加,在辐照后1.5h￣3h达高峰,12h恢复至正常水平;透射电镜显示低频超声辐照后,0.5h至9h血管内皮细胞的紧密连接均有不同程度的开放,至12h时关闭;低频超声辐照后大鼠胶质瘤侧脑组织中TNF-α含量逐渐增加,在1.5h￣3h达高峰,12h恢复至正常水平。结论低频超声辐照大鼠脑胶质瘤模型后引起TNF-α量的变化与血脑肿瘤屏障通透性变化的趋势及达到峰值的时间相一致。TNF-α的增加可能是低频超声辐照开放大鼠脑胶质瘤血肿瘤屏障的重要因素之一。     

神经胶质瘤缓激肽B2受体和nNOS表达水平与胶质瘤病理级别相关关系的研究

目的分析神经胶质瘤缓激肽B2受体和神经元型一氧化氮合酶(nNOS)表达水平与胶质瘤病理级别的相关关系。方法HE染色进行脑肿瘤的病理诊断及分级,免疫组化法测定不同病理级别神经胶质瘤缓激肽B2受体和nNOS的表达水平。结果神经胶质瘤B2受体和nNOS表达水平均与胶质瘤病理级别呈显著正相关,各病理级别神经胶质瘤B2受体和nNOS的表达水平均为:Ⅰ级(Ⅱ级(Ⅲ级。结论神经胶质瘤B2受体和nNOS表达水平均随着其病理级别的增加而升高,提示二者可作胶质瘤病理分级的标示物。     

缓激肽B2受体基因多态性对膝关节骨关节炎易感性的影响

背景：据报道在膝骨关节炎患者中应用缓激肽B2受体拮抗剂能产生持久的镇痛效果,缓激肽B2受体(BDKRB2)在骨关节炎的发展中具有重要作用。 目的：进一步明确缓激肽B2受体基因多态性与骨关节炎易感性的相关性。 方法：共有278例膝关节骨关节炎患者和291名健康志愿者参与试验。测定受试对象的缓激肽B2受体基因多态性,根据基因型和放射学分型进行统计学处理。 结果与结论：骨关节炎组和对照组中+9/-9 bp多态性的基因型分布和等位基因频率差异明显。-9/-9 bp 多态性和+9/+9 bp相比,与骨关节炎的罹患风险和严重程度存在明显相关性。-9 bp 等位基因的表达,与骨关节炎的Kellgren-Lawrence分型相关。结果提示缓激肽B2受体基因+9/-9 bp多态性可能成为骨关节炎易感性和严重性筛查的分子标记。  中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

caveolin-1对低频超声辐照选择性开放血肿瘤屏障中occludin表达及分布的影响

目的探讨低频超声辐照选择性开放血肿瘤屏障过程中,caveolin-1对occludin表达及分布的影响。方法 Western blot检测occludin的表达,测量跨内皮细胞电阻值检测血肿瘤屏障通透性,双重免疫荧光染色法检测caveolin-1及occludin在脑微血管及体外血肿瘤屏障内皮细胞内的定位表达。结果低频超声辐照后occludin表达及跨内皮细胞电阻随时间变化呈先下降后回升趋势,于低频超声辐照后1.5h,二者下降最为显著(P0.01)。以低频超声辐照后1.5h为时间点,抑制caveolin-1的表达,occludin的表达增加(P0.01),跨内皮电阻增高(P0.05)。双重免疫荧光染色显示caveolin-1和occludin在正常和脑胶质瘤大鼠脑微血管共定位,在体外血肿瘤屏障内皮细胞膜上共定位;低频超声辐照后1.5h,caveolin-1在胞质中表达显著增多,并诱导occludin从胞膜转移至胞质。结论低频超声辐照选择性开放血肿瘤屏障的过程中,caveolin-1增加血肿瘤屏障通透性与下调occludin的蛋白表达并诱导其内化相关。     

Nanoparticles functionalized with Pep-1 as potential glioma targeting delivery system via interleukin 13 receptor α2-mediated endocytosis

The treatment for glioma is one of the most challenging problems and therapeutic effect of glioma is often limited due to poor penetration into the tumor tissue. Interleukin 13 receptor α2 (IL-13Rα2) is over-expressed on tumor including established glioma cell lines and primary glioblastoma cell cultures. However, it will not cause activation of its signaling pathways. So it could be served as a promising targeted moiety for anti-glioma drug delivery. Pep-1, one specific ligand of IL-13Rα2, was identified to exhibit excellent capacity of crossing the blood tumor barrier (BTB) and homing to giloma. In this study, based on the IL-13Rα2-mediated endocytosis, Pep-1 was exploited as a potential ligand for effective glioma-targeting delivery. Pep-1 was functionalized to the surface of PEG-PLGA nanoparticles (Pep-NP) to evaluate its glioma homing, by taking advantage of the excessive expression of the IL-13Rα2 on the surface of glioma cells. Compared with non-targeting nanoparticles, Pep-NP exhibited a significantly enhanced cellular association in rat C6 glioma cells and improved penetration in 3D avascular C6 glioma spheroids. Following intravenous administration, Pep-NP could facilitate the distribution of the coumarin-6 in vivo glioma region, 2.21 times higher than that of NP for quantitative analysis. In conclusion, the Pep-NP could precisely target to the brain glioma, which was a potential targeting drug delivery system for glioma treatment.     

缓激肽对大鼠脑胶质瘤微血管中NF-κ B活性和表达的影响

目的 研究缓激肽(bradykinin,BK)对脑胶质瘤大鼠肿瘤微血管内皮细胞中转录因子核因子-κ B(nuclearfactor-kappaB,NF-κ B)的活性和表达的影响.方法 雌性Wistar大鼠96只,随机分为6组,即对照组、BK 5min组、BK 10min组、BK 15min组、BK 30min组和BK...     

Blockade of the kinin receptor B1 protects from autoimmune CNS disease by reducing leukocyte trafficking

Disruption of the blood brain barrier (BBB) and transendothelial trafficking of immune cells into the central nervous system (CNS) are pathophysiological hallmarks of Multiple Sclerosis (MS) and its animal model, Experimental Autoimmune Encephalomyelitis (EAE). Kinins are proinflammatory peptides which are released during tissue injury including EAE. They increase vascular permeability and enhance inflammation by acting on distinct bradykinin receptors, B1R and B2R. We studied the expression of B1R and B2R and the effect of their inhibition on the disease course, BBB integrity and T cell migration following myelin oligodendrocyte glycoprotein (MOG(35-55))-induced EAE. B1R, but not B2R expression was markedly enhanced in inflammatory CNS lesions in mice and humans. Brain endothelial cells could be identified as major source of B1R protein. The severity of EAE was significantly alleviated in B1R(-/-) mice compared with wild-type (WT) controls (P<0.05). Treatment of WT mice with the B1R antagonist R715 before and after disease onset was equally effective (P<0.05) while B1R activation by R838 promoted EAE (P<0.05). B1R inhibition was accompanied by a remarkable reduction of BBB disruption and tissue inflammation. In vitro analyses revealed that B1R suppression reverses the upregulation of ICAM-I and VCAM-I at the inflamed BBB thereby limiting T cell transmigration. In contrast, blocking B2R had no significant impact on EAE. We conclude that B1R inhibition can reduce BBB damage and cell invasion during autoimmune CNS disease and may offer a novel anti-inflammatory strategy for the treatment of MS.     

TNF-α在热疗降低胶质瘤侵袭性过程中的作用

 目的 探讨肿瘤坏死因子-α(TNF-α)在热疗抑制肿瘤侵袭性过程中的作用。方法 热处理大鼠恶性胶质瘤细胞(C6细胞)和胶质瘤大鼠后,放射免疫法监测培养液和脑胶质瘤组织内TNF-α的浓度;免疫组化法检测经热疗/ TNF-α/生理盐水处理过的胶质瘤组织内增殖细胞核抗原(PCNA)蛋白的表达。利用Transwell构建肿瘤侵袭模型,通过结晶紫染色法检测肿瘤侵袭性。电镜观察C6恶性胶质瘤大鼠肿瘤血管内皮细胞的凋亡。结果 热疗可增加C6细胞培养液和胶质瘤大鼠肿瘤组织内的TNF-α含量及降低胶质瘤侵袭性,均于热疗后120min时达高峰(P＜0.01)。热疗与TNF-α单独作用于胶质瘤大鼠后,均可引起胶质瘤大鼠肿瘤血管内皮细胞的凋亡。且TNF-α引起内皮细胞的凋亡水平与热处理后C6细胞培养液中TNF-α含量一致。结论 热疗可能是通过增加TNF-α引起肿瘤血管内皮细胞凋亡而抑制了肿瘤侵袭性。     

Reduced glioma growth following dexamethasone or anti-angiopoietin 2 treatment

All patients with glioblastoma, the most aggressive and common form of brain cancer, develop cerebral edema. This complication is routinely treated with dexamethasone, a steroidal anti-inflammatory drug whose effects on brain tumors are not fully understood. Here we show that dexamethasone can reduce glioma growth in mice, even though it depletes infiltrating T cells with potential antitumor activity. More precisely, T cells with helper or cytotoxic function were sensitive to dexamethasone, but not those that were negative for the CD4 and CD8 molecules, including gammadelta and natural killer (NK) T cells. The antineoplastic effect of dexamethasone was indirect, as it did not meaningfully affect the growth and gene expression profile of glioma cells in vitro. In contrast, hundreds of dexamethasone-modulated genes, notably angiopoietin 2 (Angpt2), were identified in cultured cerebral endothelial cells by microarray analysis. The ability of dexamethasone to attenuate Angpt2 expression was confirmed in vitro and in vivo. Selective neutralization of Angpt2 using a peptide-Fc fusion protein reduced glioma growth and vascular enlargement to a greater extent than dexamethasone, without affecting T cell infiltration. In conclusion, this study suggests a mechanism by which dexamethasone can slow glioma growth, providing a new therapeutic target for malignant brain tumors.     

热疗降低胶质瘤侵袭性的作用与TNF-α和Cx-43的关系

目的:探讨热疗诱发胶质瘤细胞释放的肿瘤坏死因子-α(TNF-α)对胶质瘤细胞侵袭性的影响机制。方法:热处理C6细胞后,应用放射免疫法动态监测培养液内TNF-α的含量;观察热处理C6细胞后的条件培养液(C6CM)对胶质瘤细胞内的丝裂素活化蛋白激酶(mitogen-activated protein kinase,MAPK)mRNA的转录水平、NF-κB含量及胶质瘤侵袭性的影响;利用免疫荧光技术检测C6CM对体外胶质瘤侵袭模型上连接蛋白Cx-43表达的影响。结果:热处理C6细胞后,培养液内TNF-α的含量增加,于120 min时达高峰。C6CM作用于体外胶质瘤侵袭模型后,胶质瘤细胞的MAPK mRNA转录水平及NF-κB含量增加,且均于第120 min时达最高水平。与此同时,体外胶质瘤细胞的连接蛋白Cx-43表达水平也呈相同的变化趋势。结论:热疗可诱发C6细胞释放TNF-α,其可能是通过增加胶质瘤细胞的MAPK mRNA转录水平和NF-κB的含量而导致胶质瘤细胞的连接蛋白Cx-43的表达增加,进而引起胶质瘤侵袭性降低的。