内皮 – 单核细胞激活多肽 – Ⅱ上调LINC00263增强血肿瘤屏障通透性

  目的  探讨缓激肽与罂粟碱（papaverine,PA）联合应用对血肿瘤屏障通透性和紧密连接（tight junctions,TJ）相关蛋白之闭锁小带蛋白1（ZO-1）表达的影响。  方法  应用大鼠胶质瘤模型,伊文思蓝（Evans blue,EB）渗透性实验检测血肿瘤屏障通透性,免疫组织化学法测定ZO-1蛋白表达,RT-PCR法测定ZO-1 mRNA表达。  结果  与对照组大鼠脑肿瘤组织中EB含量 [（0.182 ± 0.026）μg/g]比较,缓激肽、罂粟碱、缓激肽+罂粟碱组（联合组）大鼠脑肿瘤组织中EB含量[分别为（0.487 ± 0.031）、（0.503 ± 0.029）和（0.875 ± 0.040）μg/g]均明显升高（P < 0.01）;与缓激肽、罂粟碱组比较,联合组大鼠脑肿瘤组织中EB含量进一步增加（P < 0.01）。与对照组（0.379 ± 0.011）比较,缓激肽、罂粟碱、联合组大鼠脑肿瘤组织中ZO-1蛋白表达[分别为（0.259 ± 0.008）、（0.252 ± 0.010）和（0.135 ± 0.015）]均明显下降（P < 0.01）;与缓激肽、罂粟碱组比较,联合组大鼠脑肿瘤组织中ZO-1蛋白表达进一步降低（P < 0.01）。与对照组（0.876 ± 0.062）比较,缓激肽、罂粟碱、联合组大鼠肿瘤组织中ZO-1 mRNA 表达[分别为（0.735 ± 0.036）、（0.695 ± 0.050）和（0.420 ± 0.047）]均明显降低（P < 0.01）;与缓激肽、罂粟碱组比较,联合组大鼠脑肿瘤组织中ZO-1 mRNA表达进一步降低（P < 0.01）。  结论  缓激肽与罂粟碱联合应用对血肿瘤屏障的开放作用具有协同效应,此作用与紧密连接相关蛋白ZO-1表达水平下调有关。     

Function and structure of bradykinin receptor 2 for drug discovery

Type 2 bradykinin receptor (B2R) is an essential G protein-coupled receptor (GPCR) that regulates the cardiovascular system as a vasodepressor. Dysfunction of B2R is also closely related to cancers and hereditary angioedema (HAE). Although several B2R agonists and antagonists have been developed, icatibant is the only B2R antagonist clinically used for treating HAE. The recently determined structures of B2R have provided molecular insights into the functions and regulation of B2R, which shed light on structure-based drug design for the treatment of B2R-related diseases. In this review, we summarize the structure and function of B2R in relation to drug discovery and discuss future research directions to elucidate the remaining unknown functions of B2R dimerization.     

Effects of Bradykinin B2 Receptor Blockade on Infarct Size and Hemodynamics after Myocardial Infarction in Enalapril-treated Rats

Objectives To study the effects of bradykinin （BK） B2 receptor blockade on infarct size and hemodynamics after myocardial infarction （MI） in rats with angiotensin-converting enzyme （ACE） inhibition therapy. Methods MI was produced by ligating the left coronary artery. The effects of enalapril （ 500 μg/kg·day）, enalapril （ 500 μg/kg · day） with BK B2 receptor antagonist Hoe-140 （500 μg/kg · day）, angiotensin Ⅱ （Ang Ⅱ） type 1 （AT1） receptor antagonist losartan （3 mg/kg · day） on infarct size, left ventricular systolic pressure （ LVSP）, cardiac output index （CI） and stroke volume index （SVI） were observed in rats after MI. Treatments were started on the 2nd day after MI and continued for another 6 weeks. Results Enalapril reduced infarct size and improved CI and SVI compared with the untreated MI group （ P 〈 0. 05 ）, and these effects of enalapril were significantly blunted by concomitant treatment with Hoe-140 （P 〈 0. 05）. Losartan was less effective than enalapril. LVSP were unchanged in the three treatment groups. Conclusions BK can reduce infract size and improve hemodynamics in rats following MI. The cardioprotective effects of ACEI partly result from the action of BK exerted through the B2 receptor.     

雷米普利预适应对大鼠心脏延迟性的保护作用

目的探讨雷米普利预适应对大鼠离体心脏缺血-再灌注损伤的延迟性保护作用及机制。方法离体大鼠心脏采用Langendoff灌流法建立心肌缺血再灌注损伤模型。35只大鼠随机分为5组：（1）对照组；（2）缺血-再灌组（I／R组）；（3）雷米普利预处理组（Ramipril 0．05mg／kg）；（4）雷米普利预处理组（Ramipril 0．1mg／kg）：（5）HOE140＋雷米普利预处理组（HOE1400．1mg／kg＋Ramipril 0．1mg／kg）。每组7只。记录左室内压（LVP）、左室内压最大上升速率（＋dp／dtmax）、左室舒张末压（LVEDP）、心率（HR），定时收集冠脉流出液测量冠脉流量（CF）和肌酸激酶（CK）活性。再灌注结束后采用分光光度法测定心肌组织中丙二醛（MDA）含量。结果（1）与对照组比较，I／R组缺血-再灌注后10、20、30min可显著降低LVP和＋dp／dt max（P〈0.01），升高LVEDP（P〈0．01），降低CF（P〈0．01），缺血-再灌注后10、20min显著降低HR（P〈0．01）；（2）与I／R组比较，Ramipril 0．05mg／kg组缺血-再灌注后20、30min可显著升高LVP（P〈0．01）及增加CF（P〈0．05）；缺血-再灌注后10、20、30min可显著升高＋dp／dtmax（P〈0.01），降低LVEDP（P〈0.01）；（3）与I／R组比较，Ramipril 0．1mg／kg组缺血-再灌注后10、20、30min可显著升高LVP（P〈0．01），升高＋dp／dtmax（P〈0.01），降低LVEDP（P〈0.01）；增加CF（P〈0.05）；（4）与Ramipril 0．1mg／kg组比较，HOE1400．1mg／kg＋Ramipril 0．1mg／kg组缺血-再灌注后10、20、30min可显著降低LVP（P〈0.05）；降低＋dp／dtmax（P〈0．05）；升高LVEDP（P〈0.05）；降低CF（P〈0．05）；（5）I／R组缺血-再灌注后10、20、30minCK与对照组比较，均有显著性差异（P均〈0.01）；Ramipril 0．05mg／kg组及Ramipril 0．1mg／kg组缺血-再灌注后10、20、30minCK与I／R组比较，均有显著性差异（P均〈0.01）；HOE1400．1mg／kg＋Ramipril 0．1mg／kg组缺血-再灌注后10、20、30minCK与Ramipril 0．05mg／kg组及Ramipril 0．1mg／kg组比较，均有显著性差异（P均〈0.01）；（6）实验前24h舌下静脉注射雷米普利0．05mg／kg或0．1mg／kg均可显著降低缺血-再灌注心肌组织中MDA含量。结论雷米普利对缺血再灌注诱导离体大鼠心肌损伤具有延迟性保护作用，其机制可能是与激活缓激肽B2受体，减少缓激肽降解有关。     

Modulation of voltage-activated Ca currents by pain-inducing agents in a dorsal root ganglion neuronal line,F-11

Whole cell currents evoked by pain-inducing agents—bradykinin (Bk), capsaicin (Cap), and reciniferatoxin (RTX), and their modulation of voltage-activated Ca currents were examined in F-11 cells using a patch electrode voltage clamp technique. Most F-11 cells generated action potentials under current clamp if their membrane potentials were held sufficiently negative. Average peak inward Na current (I_Na) was 100 μA/cm² and the I_Na was abolished by 10^?6 M tetrodotoxin. At least two types of Ca currents could be clearly distinguished on the basis of voltage dependency and kinetics; a low threshold transient I_Ca(t) and a high threshold sustained I_Ca(I). In addition, another high threshold transient Ca current, presumably I_Ca(n), was observed. About 30% of the cells produced inward current for these pain-inducing agents, when activated at the membrane holding potential of ?70 mV. In some F-11 cells, the amplitude of action potential was observed to increase during 10^?6 M Cap-induced depolarization. Both low and high threshold Ca currents were reduced by 10^?6 M Bk in the majority of the cells. Similarly, both 10^?6 M Cap and 10^?9 M RTX reduced these Ca currents. However, a considerable number of cells showed an initial enhancement followed by reduction in the amplitude of these Ca currents. With higher concentrations of these ligands, all Ca currents were suppressed. Such modulation of voltage-activated Ca currents by pain-inducing agents occurred in both the presence and absence of apparent receptor-activated current flows in the cells. In pertussis toxin (PTX)-treated cells, the inhibitory modulation of Ca currents by pain-inducing agents was suppressed. In contrast, in cholera toxin (CTX)-treated cells, this inhibitory modulation appeared to be enhanced. These data indicate that the inhibitory modulation of Ca channel currents by Cap and RTX, similarly to that of Bk, involves a PTX-sensitive inhibitory G protein (G_i). © 1993 Wiley-Liss, Inc.     

高表达缓激肽B2受体的大鼠胶质瘤模型的研究

目的建立高表达缓激肽受体(B2R)的大鼠胶质瘤模型,为研究缓激肽选择性开放血脑屏障的机制及解决目前临床应用中存在的问题提供必要的模型。方法①大鼠B2R的真核细胞的表达和其载体(prB2R)侵染C6胶质瘤细胞株;②Real-timeRT-PCR测定B2R的转录;③WesternBlot法测定B2R的表达水平。结果①大鼠胶质瘤细胞株C6高表达B2R;②Real-timeRT-PCR测定C6-B2R1和C6-B2R2克隆的B2R分别比C6对照高7.6和6.9倍;③C6-B2R1和C6-B2R2克隆的蛋白表达水平高于C6对照克隆的3.8和3.78倍。移植C6-B2R1肿瘤1周后的B2R表达水平高于C6对照肿瘤的3.6倍。结论高表达B2R的大鼠胶质瘤模型已被成功建立。     

SYNERGISTIC POTENTIATION BY CAPTOPRIL AND PROPRANOLOL OF BRADYKININ-INDUCED BRONCHOCONSTRICTION IN THE GUINEA-PIG

1. Captopril (30 or 100 micrograms/kg intravenous (i.v.] in anaesthetized artificially ventilated guinea-pigs potentiated bronchoconstrictor responses to bradykinin, but not those to histamine or the thromboxane A2-mimetic U46619. 2. Propranolol (5 mg/kg, i.v.) potentiated bradykinin-induced broncho-constriction. The potentiated responses were further augmented by captopril. 3.The captopril-potentiated responses to bradykinin were inhibited during cyclo-oxygenase inhibition with indomethacin. Bronchoconstrictor responses to bradykinin, but not those to histamine or U46619, were reduced after thromboxane synthase inhibition with dazoxiben. The thromboxane A2 antagonist AH23848 inhibited bronchoconstrictor responses to bradykinin, arachidonic acid or U46619 whereas it did not affect those to histamine. 4. A kininase I inhibitor DL-2-mercaptomethyl-3-guanidinoethyl thiopropanoic acid caused no change in bronchoconstriction caused by bradykinin and did not alter the potentiated responses occurring after captopril. 5. Thus, confirmation has been obtained that bradykinin causes broncho-constriction in the guinea-pig indirectly, by release of eicosanoids. Thromboxane A2 is likely to be the major eicosanoid released, since the bronchoconstrictor effect of bradykinin was blocked by indomethacin, dazoxiben and AH23848. The intensity of the bronchoconstriction appears dependent on sympathetic influences mediated by beta-adrenoceptors. Kininase I, in contrast to kininase II apparently has little role in terminating the effects of bradykinin in the lung.     

Cultured astrocytes express proteins involved in vesicular glutamate release

Bradykinin induces receptor-mediated calcium-dependent release of glutamate from cultured astrocytes through a mechanism that is neither due to cell-swelling mechanism nor due to the reversal of the glutamate transporter. Astrocytes may thus release glutamate using a mechanism resembling the neuronal vesicular release of neurotransmitters. Synaptobrevin is a vesicular protein that together with plasma membrane proteins syntaxin and SNAP-25 participate in formation of the anchoring core complex required for initiation of exocytosis. Here, we demonstrate that synaptobrevin II is present in cultured astrocytes. Furthermore, we demonstrate that botulinus toxin type B and tetanus toxin cause a decrease in synaptobrevin II immunoreactivity and abolish bradykinin-induced release of glutamate from cultured astrocytes. While we were not able to demonstrate the presence of SNAP-25 or syntaxin immunoreactivity in cultured astrocytes, pretreatment with BoTx-A (which cleaves SNAP-25) and BoTx-C (which cleaves syntaxins) result in a decrease in the baseline release of glutamate and diminish the bradykinin-evoked release of glutamate from cultured astrocytes. These findings strongly support the notion that astrocytes may release neurotransmitters using a mechanism similar to the neuronal secretory process.     

糖皮质激素抑制缓激肽引起的PC12细胞外钙内流与蛋白激酶C有关

缓激肽（BK）可引起PC12细胞的[Ca2＋]i；升高，该作用不通过电压敏感性钙通道，而是由G蛋白转导，通过内钙释放和外钙内流引起的。BK的作用可受到皮质疏（B）的快速抑制。本研究利用无钙／再加钙方法，把BK对内钙释放和外钙内流作用分开来进行研究。结果发现：（1）B可以抑制BK引起的外钙内流，对内钙释放影响不明显；（2）B－BSA（牛血清白蛋自偶联的皮质疏）对BK所引起外钙内流的抑制作用与游离的B作用类似；（3）PKC激活剂（PMA）可以模拟，而PKC的抑制剂G66976可以逆转B的抑制作用；（4）百日咳毒素（PTX）预处理可以阻断B抑制BK引起的外钙内流。结果提示：B可能作用在膜受体上，通过PTX敏感的G蛋白-PKC途径抑制BK引起的外钙内流。