HPLC法测定灯盏花素片中灯盏花乙素的含量

目的:建立灯盏花素片中灯盏花乙素的含量测定方法;方法:采用高效液相色谱法,使用C18柱,流动相为甲醇-水-冰醋酸(30:70:1),检测波长335nm;结果:该制剂中灯盏花乙素在0.2～4.0μg范围内线性良好,平均回收率为100.21%,RSD为1.48%;结论:该方法简便、快速、准确,可用于灯盏花素片的含量测定和质量控制.     

HPLC法测定灯盏花素片中灯盏花乙素的含量

目的建立灯盏花素片中灯盏花乙素的含量测定方法。方法采用高效液相色谱法,使用C18柱,流动相为甲醇-水-冰醋酸(30∶70∶1),检测波长335nm。结果该制剂中灯盏花乙素在0.0624～0.312μg范围内线性良好,平均回收率为99.92%,RSD为0.24%。结论该方法简便、快速、准确,可用于灯盏花素片的含量测定和质量控制。     

灯盏花素对豚鼠单一心室肌细胞ICa的抑制作用

目的：观察灯盏花素对豚鼠单一心室肌细胞钙离子电流（ＩＣａ）的影响。方法：应用全细胞膜片钳制技术。结果：灯盏花素能明显抑制心室肌细胞的Ｃａ＾２＋通道,使ＩＣａ减小。此作用有明显的电压依赖性。在峰电流电压下作用最明显,而对其反转电位无明显影响。在指令电位０ｍＶ时,０．５ｍｇ％灯盏花素使ＩＣａ减小５．４％,１ｍｇ％灯盏花素使ＩＣａ减小２２．９％（Ｐ〈０．０１）,２ｍｇ％灯盏花素使ＩＣａ减小４５．０％（Ｐ     

灯盏花素对大鼠脑微血管内皮细胞OGD损伤的保护作用及机制研究

<正>灯盏花素(breviscapine,Bre)是从短亭飞蓬中提取的黄酮类化合物,具有保护缺血性脑血管疾病、调节血管内皮功能、减轻炎性反应等作用[1]。本实验通过分离培养大鼠脑微血管内皮细胞(BMECs),建立氧糖剥夺(OGD)BMECs模型,研究Bre对BMECs损伤是否具有保护作用,并探讨其可能的机制。相关研究尚未见报道。1材料与方法     

灯盏花素抗肾上腺素实验性心律失常的作用

目的观察并比较灯盏花素、普萘洛尔及利多卡因对心律失常的保护作用.方法以静脉缓慢恒速注射肾上腺素诱发大鼠发生多种心律失常.结果灯盏花素(8mg/kg)和普萘洛尔(1mg/kg)均可使肾上腺素诱发的大鼠心律失常发生时间延迟、严重程度减轻、死亡率降低(与生理盐水组比较,P＜0.05),灯盏花素与普萘洛尔的效果相近,优于利多卡因.结论灯盏花素可有效地预防肾上腺素过量所导致的心律失常.     

灯盏乙素和灯盏花素对急性心肌梗死的保护作用

目的研究灯盏乙素和灯盏花素对急性心肌梗死的保护作用。方法建立大鼠急性心肌梗死模型后,腹腔注射不同剂量灯盏乙素和灯盏花素,4h后取出心脏行氯化硝基四氮唑蓝（NBT）染色,测量心肌梗死面积,比较两种药物对心肌梗死面积影响的量效关系;建立犬急性心肌梗死模型,股静脉注射50mg/kg灯盏花素或灯盏乙素,记录不同时间点的心外膜电图,比较两种药物对心肌缺血程度和范围的影响。结果灯盏乙素单体能够降低心肌梗死面积,抑制梗死区心肌细胞的凋亡,50mg/kg灯盏乙素能够降低左冠状动脉前降支结扎后犬心外膜电图抬高的∑ST段。结论灯盏乙素单体是灯盏花素的主要药理活性成分,较灯盏花素具有更好的量效关系。     

灯盏花素的研究进展

灯盏花素及其制剂在心脑血管疾病治疗领域展现了良好的疗效,本文综述了近年来灯盏花素在药理学、药动学、制剂、制备、结构改造及临床应用等方面的最新研究进展。     

灯盏花素静脉给药抗LPS致大鼠急性肾损伤作用的研究

目的:探讨灯盏花素对脓毒症大鼠常见肾功能指标如肾组织TNFα和IL-1β水平的影响。方法:90只SD大鼠随机等分为脓毒症组、灯盏花素组、对照组。脓毒症的建模采用盲肠结扎穿孔(CLP)构建;对照组则予除CLP外的相同手术。灯盏花素组制模前1h尾静脉注射灯盏花素10mg/kg。对照组和脓毒症组则分别肌注平衡液5ml/kg,术后每隔12小时同样给药一次。术后24小时取肾组织,ELISA检测肾组织中TNFα及IL-1β表达水平。结果:灯盏花素组大鼠肾脏中TNFα及IL-1β表达水平较脓毒症组显著降低。结论:灯盏花素可能是调节TNFα及IL-1β表达而起到对脓毒症肾组织的保护作用。     

注射用灯盏花素治疗脑梗塞106例的疗效验证观察

自１９９４年８月～１９９５年３月我省临床应用注射用灯盏花素（冻干粉针剂）,取得了较好的效果,现报告如下。临床资料　病例选择　本组病例均为住院患者,经临床、ＣＴ、ＭＲＩ检查确诊为脑梗塞的患者。采用单盲对照,随机选择发病在１月内的病例分为３组：Ａ组为注谢用灯盏花素治疗组１０６例,男５５例,女５１例,年龄４４～８２岁,平均６３－４１±１２－６８岁。Ｂ组为云南灯盏花注射液对照组５０例,男２２例,女２８例,年龄５２～７９岁,平均６５－５±１０－２５岁。Ｃ组为复方丹参对照组４４例,男２６例,女１８例,年龄４…     

Activation of large‐conductance Ca2+‐activated K+ channels by pinacidil in human umbilical vascular endothelial cells

The effect of pinacidil, an opener of ATP‐sensitive K⁺ (K_ATP) channels, on large‐conductance Ca²⁺‐activated K+ (BK_Ca) channels was investigated in cultured endothelial cells of human umbilical veins. In whole cell configuration, pinacidil (30 μM) increased the amplitude of K⁺ outward currents (I_K). Charybdotoxin (100 nM), but not glibenclamide (10 μM), suppressed pinacidil‐induced increase in I_K. Neither carbonyl cyanide m‐chlorophenyl hydrazone (CCCP; 10 μM), an inhibitor of mitochondrial Ca²⁺‐uniporter, nor cyclosporin A (200 nM), an inhibitor of the mitochondrial permeability transition pore, affected pinacidil‐induced increase in I_K. In inside‐out patch configuration, bath application of pinacidil (30 μM) did not change single channel conductance but increased the activity of BK_Ca channels. Pinacidil (30 μM) shifted the activation curve of BK_Ca channels to less positive membrane potential by approximately 15 mV. Pinacidil stimulated the activity of these channels in a concentration‐dependent manner. The EC₅₀ value for pinacidil‐induced channel activity was 20 μM. After BK_Ca channels had been enhanced by Evans blue (100 μM), subsequent application of pinacidil (100 μM) did not further increase the channel activity. These results clearly indicate that in addition to the activation of K_ATP channels, pinacidil can also stimulate BK_Ca channels in endothelial cells. These effects could contribute to the regulation of vascular tone if similar results were found in endothelial cells in vivo. Drug Dev. Res. 48:6–16, 1999. © 1999 Wiley‐Liss, Inc.     

溴氰菊酯对大鼠神经细胞内游离钙的影响

目的 研究溴氰菊酯对大鼠神经细胞内游离钙([Ca^2 ]i)的影响,并探讨其可能机制。方法 用溴氰菊酯处理分离的大鼠脑细胞,用Fura-2/AM为细胞内钙离子荧光指示剂,测定神经细胞内游离钙浓度;并利用NMDA受体竞争性拮抗剂AP5、非竞争性拮抗剂MK－801、Na^ 离子通道阻断剂TTX、电压依赖性钙通道阻断剂尼莫地平,探讨了溴氰菊酯影响胞内钙的可能机制;另外还观察了去除胞外钙时溴氰菊酯对胞内钙的影响情况。结果 溴氰菊酯浓度在0－200nmol/L范围内,可以显著提高大鼠皮层和海马细胞内游离钙的浓度（P＜0．01）,并有良好的剂量－反应关系（相关系数分别为r=0.964,r=0.981);AP5、MK－801和TTX可以有效阻断溴氰菊酯的升钙作用;而尼莫地平则无影响;去除胞外钙时,溴氰菊酯的升钙作用也消失。结论 溴氰菊酯导致的胞内钙升高,主要是由谷氨酸激活NMDA受体门控钙通道引起的胞外钙内流,和电压依赖性钙通道及胞内钙库释放无关。     

异丙酚对PC12细胞膜流动性及[Ca~(2+)]_i的影响

目的　研究异丙酚对PC12细胞膜流动性及 [Ca2 + ]i的影响。方法　以PC12细胞作为神经细胞模型 ,加入不同浓度的异丙酚 ,用荧光偏振法测定细胞膜微粘度 η的动态变化 ,用激光扫描共聚焦显微镜检测 [Ca2 + ]i 随时间变化曲线。结果　①异丙酚各剂量组的 η值均降低 ,并呈现剂量依赖性 ;② 30、10 0mg·L-1的异丙酚在加药后 2 0～ 30s使[Ca2 + ]i 短暂升高 ,[Ca2 + ]i的峰值分别比加药前增加了119%和 14 0 % ,但在 5 0s内均恢复到加药前水平。结论　异丙酚的全麻作用机制可能与细胞膜流动性增高有关     

脉络宁注射液对人血管内皮细胞缺氧损伤的保护作用

目的:研究脉络宁注射液对人脐静脉血管内皮细胞缺氧损伤的保护作用及其机制。方法:常规进行人血管内皮细胞(HUVECs)培养,将细胞随机分为正常对照组、缺氧组和脉络宁20 mg.L-1组。采用CCK-8法检测细胞存活率;Hochest33258荧光染料检测细胞凋亡率;RT-PCR法检测各组细胞中血管内皮生长因子(VEGF),血管生成素-2(Ang-2)mRNA表达水平。结果:脉络宁注射液可显著提高缺氧损伤细胞的存活率(P<0.05);与正常对照组比较,缺氧组细胞内VEGF和Ang-2 mRNA表达水平明显增高。与缺氧组比较,脉络宁组VEGF和Ang-2的表达进一步增强,各组间比较均有差异(P<0.05)。结论:脉络宁注射液可减轻血管内皮细胞缺氧损伤,上调缺氧血管内皮细胞中VEGF和Ang-2的表达,对血管内皮细胞缺氧损伤有一定的保护作用。     

High glucose-induced human umbilical vein endothelial cell hyperpermeability is dependent on protein kinase C activation and independent of the Ca2+-nitric oxide signalling pathway

1. Endothelial barrier dysfunction plays a pivotal role in the pathogenesis of diabetic vascular complications. The precise molecular mechanisms by which hyperglycaemia causes the increased permeability in endothelial cells are not yet well understood. In the present study, we investigated whether high concentrations of glucose induce endothelial permeability through the activation of protein kinase C (PKC) and/or the calcium-nitric oxide (NO) signalling pathway in human umbilical vein endothelial cells (HUVEC). 2. Endothelial permeability was measured by albumin diffusion across endothelial monolayers under the stimuli of high glucose (HG; 20 mmol/L), 100 nmol/L phorbol-myristate-acetate (PMA) or 100 nmol/L histamine. The intracellular calcium concentration ([Ca2+]i) was detected in HUVEC using the fluorescent probe fura-2 AM. The effects of PKC inhibitors (LY379196 and hypocrellin A) and the NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA) on endothelial permeability and [Ca2+]i were determined. 3. High glucose and PMA increased endothelial permeability associated with decreased [Ca2+]i, whereas histamine triggered significant increases in endothelial permeability, accompanied by increases in [Ca2+]i in HUVEC. Hypocrellin A (HA) and LY379196 reversed both HG- and histamine-induced endothelial permeability. The NOS inhibitor L-NMMA only abolished histamine- and not HG-induced endothelial permeability. Neither LY379196, HA nor L-NMMA had any significant effects on alterations in [Ca2+]i caused by HG and histamine. 4. These results indicate that increased endothelial permeability in HUVEC induced by HG is dependent on PKC activity and is independent of the [Ca2+]i-NO pathway. Increased endothelial permeability due to other inflammatory factors, such as histamine, may also be mediated by the PKC pathway. Thus, PKC inhibitors would be a potential therapeutic approach to endothelial dysfunction induced by hyperglycaemia, as well as other inflammatory factors, in diabetes.     

辛伐他汀对脐静脉内皮细胞金属基质蛋白酶9表达的影响

目的观察辛伐他汀对脐静脉内皮细胞（human umbilical vein endothelial cell,HUVEC）金属基质蛋白酶9（matrix metalloproteinase-9,MMP-9）表达的影响。方法采用逆转录聚合酶链反应及蛋白质免疫印迹分析检测MMP-9mRNA转录和蛋白水平表达,观察辛伐他汀不同浓度及不同孵育时间HUVECMMP-9表达的影响。结果辛伐他汀呈浓度和时间依赖性减低HU-VEC的MMP-9mRNA转录和蛋白水平的表达。结论辛伐他汀可抑制HUVE CMMP-9表达,防治动脉粥样硬化。     

Apoptosis of human umbilical vein endothelial cells induced by artesunate

Artesunate (ART), a semi-synthetic derivative of artemisinin isolated from the traditional Chinese herb Artemisia annua, is an effective novel antimalarial drug. The present study investigated the apoptotic activity of artesunate in cultured human umbilical vein endothelial cell (HUVEC) by means of nuclear staining, DNA agarose gel electrophoresis, and flow cytometry. The observations also indicated that artesunate induced apoptosis of HUVEC in a concentration-dependent and time-dependent manner. A Western immunoblot analysis showed down-regulation of the bcl-2 protein and up-regulation of the bax protein in the artesunate-treated HUVEC. Ca2+ in cells was evaluated by fluorescent spectrophotometer using Fura 2-AM as probe. These results suggest that artesunate may be a potential apoptosis-inducing agent for endothelial cells.     

CYP1A1对人血管内皮细胞内钙的影响

目的探讨CYP1A1对人血管内皮细胞HVEC304细胞内钙的影响,探索CYP1A1的内源性作用。方法构建CYP1A1的红色荧光表达载体,转染人血管内皮细胞,激光共聚焦检测细胞内的钙离子浓度。结果CYP1A1表达载体在人血管内皮细胞中成功表达,定位于细胞质。过表达CYP1A1的HVEC304细胞(HVEC304-CYP1A1)中游离钙的浓度明显低于未转染的HVEC304细胞(P<0.01)。随着veratridine浓度的上升,HVEC304-CYP1A1细胞中游离钙的水平逐渐上升。结论过表达CYP1A1人血管内皮细胞中,钙离子基础水平明显降低,veratridine可以剂量依赖性地逆转这一作用。     

Kaempferol stimulates large conductance Ca2+ -activated K+ (BKCa) channels in human umbilical vein endothelial cells via a cAMP/PKA-dependent pathway

BACKGROUND AND PURPOSE: Kaempferol has been shown to possess a vasodilator effect but its mechanism of action remains unclear. In this study, experiments were carried out to study the effect of kaempferol on K+ channels in endothelial cells. EXPERIMENTAL APPROACH: K+ channel activities in human umbilical vein endothelial cells (HUVECs) were studied by conventional whole cell and cell-attached patch-clamp electrophysiology. KEY RESULTS: Kaempferol stimulated an outward-rectifying current in HUVECs in a dose-dependent manner with an EC50 value of 2.5+/-0.02 microM. This kaempferol-induced current was abolished by large conductance Ca2+ -activated K+ (BKCa) channel blockers, such as iberiotoxin (IbTX) and charybdotoxin (ChTX), whereas the small conductance Ca2+ -activated K+ (SKCa) channel blocker, apamin, and the voltage-dependent K+ (KV) channel blocker, 4-aminopyridine, had no effect. Cell-attached patches demonstrated that kaempferol increased the open probability of BkCa channels in HUVECs. Clamping intracellular Ca2+ did not prevent kaempferol-induced increases in outward current. In addition, the kaempferol-induced current was diminished by the adenylyl cyclase inhibitor SQ22536, the cAMP antagonist Rp-8-Br-cAMP and the PKA inhibitor KT5720, but was not affected by the guanylyl cyclase inhibitor ODQ, the cGMP antagonist Rp-8-Br-cGMP and the PKG inhibitor KT5823. The activation of BKCa channels by kaempferol caused membrane hyperpolarization of HUVECs. CONCLUSION AND IMPLICATIONS: These results demonstrate that kaempferol activates the opening of BKCa channels in HUVECs via a cAMP/PKA-dependent pathway, resulting in membrane hyperpolarization. This mechanism may partly account for the vasodilator effects of kaempferol.     

溶血卵磷脂对血管内皮细胞增殖及凋亡的影响

目的观测溶血卵磷脂(LPC)对血管内皮细胞增殖及凋亡的影响,从而探讨动脉硬化发生的机制。方法取体外培养的人脐静脉内皮细胞(HUVECs),在含不同浓度LPC(5、10、20mg/L)的培养基中分别培养6、12、24、48h,用四甲基偶氮唑蓝(MTT)比色法、流式细胞仪、荧光显微镜观测血管内皮细胞增殖及凋亡的变化。结果与正常对照组比较,LPC抑制内皮细胞增殖,促进细胞凋亡,且其作用呈时间-效应、浓度-效应依赖关系,但随着LPC浓度增加,细胞凋亡增加的同时细胞死亡比例也增加。结论LPC抑制血管内皮细胞增殖,促进细胞凋亡,可能是其致动脉粥样硬化机制之一。