补阳还五汤及有效组分生物碱和苷对大鼠血小板聚集及血小板环核苷酸的影响

目的探讨补阳还五汤及其有效组分生物碱和苷抗血小板聚集机制。方法大鼠分别给予补阳还五汤原方、生物碱、苷和噻氯匹定，进行ADP诱导的血小板聚集实验。取聚集前、后的血小板提取cAMP、cGMP，采用放免法检测血小板cAMP、cGMP。结果各组血小板聚集比较，生物碱组、苷组和噻氯匹定组血小板聚集强度与空白组相比显著降低（P〈0．01）。原方组血小板聚集强度与空白组相比显著降低（P〈0．05）。血小板聚集后cAMP含量降低（P〈0．01）．而原方、生物碱、苷和噻氯匹定均可抑制ADP诱导的血小板cAMP下降（P〈0．05，P〈0．01）。血小板聚集后cGMP含量降低（P〈0．01），原方、生物碱、苷和噻氯匹定也可抑制聚集后血小板cGMP下降（P〈0．05，P〈0．01）。结论生物碱、苷、原方和噻氯匹定可抑制ADP诱导的大鼠血小板聚集，各药可抑制血小板聚集后血小板内cAMP、cGMP的下降，提示其抗血小板聚集作用是通过抑制聚集后血小板内环核苷酸降低而实现的。     

影响血小板聚集试验结果的因素探讨

本文应用北京世帝科学仪器公司生产的 L G- PABER- 型血小板聚集仪 ,三磷酸腺苷 (ADP)作聚集诱导剂 ,对影响血小板聚集结果的因素进行探讨。1　 ADP浓度对血小板聚集速度和强度将 ADP配成 0 .1、0 .5、1.0及 2 .0 U mol/L四种不同浓度 ,分别诱导同批 2 0份标本 ,测定 2分钟及 4分钟的聚集率。结果显示 ,ADP浓度在 0 .5～ 2 .0 μmol/L 范围内对血小板聚集结果无明显差别。2　 ADP配成实验浓度后的稳定性将配成 1.0 μmol/L ADP在 4℃保存第 3、 7、 10、 14天分别与鲜配制的 1.0 μmol/L 测定同一份标本 ,比较其聚集率结果 ,存…     

牛心内膜腺苷三磷酸双磷酸酶对血小板聚集的抑制作用

AIM: To study the anti-aggregatory effect of bovine endocardial endothelial cell (EEC)-associated apyrase. METHODS: Cultured bovine EEC was used. Adenosine diphosphate (ADP) was analyzed by reversed phase HPLC, and rabbit platelet aggregation was measured turbimetrically. RESULTS: Incubation of EEC with ADP 500 mumol.L-1 resulted in a progressive decrease in ADP concentration, which was paralleled by the decrease in platelet aggregating potential of the unmetabolized ADP. In the presence of aspirin (Asp 1 mmol.L-1)-treated EEC 1 x 10(9) cells.L-1, the aggregation of Asp (1 mmol.L-1) and methylene blue (10 mumol.L-1)-treated platelets in response to thrombin 500 U.L-1 and platelet activating factor (PAF 1 nmol.L-1) was markedly inhibited and was reversible, which was very similar to that in apyrase-treated platelets. The supernatants of EEC had no effect on platelet aggregation. EEC inhibited ADP (5 mumol.L-1)-induced platelet aggregation, but failed to inhibit adenosine 5'-O-(2-thiodiphosphate) (ADP-beta-S, an unmetabolizable structural analog of ADP, 15 mumol.L-1)-induced platelet aggregation. CONCLUSION: ADP hydrolysis by EEC-associated apyrase is a major anti-thrombotic mechanism of bovine EEC.     

普罗托品对家兔血小板功能的影响

普罗托品(protopine,Pro)体外(1—1000μmol·L~(-1))和体内(10和20mg·kg~(-1))均抑制ADP,胶原,花生四烯酸(AA)和烙铁头蛇毒血小板聚集素(TMVA)诱导的兔血小板聚集及血小板5-HT释放。Pro不抑制AA诱导的免血小板TXA_2生成。也不升高血小板内cAMP水平,但升高cGMP水平。提示其抗血小板作用的机制与升高血小板内cGMP水平,抑制血小板释放活性物质有关。     

MK-447对兔血小板中胶原、ADP及血栓素A_2稳定类似物诱导的血小板变形、聚集和腺苷三磷酸释放的影响

目的:研究MK-447对胶原、ADP及血栓素A_2稳定类似物(STA_2)诱导的血小板变形、聚集和释放反应的影响。方法:浊度法评价血小板变形和聚集反应,测定富含血小板血清中ATP的量确定释放反应。结果:(1)MK-447诱导血小板变形,不被吲哚美辛抑制。预置MK-447可使胶原、ADP及STA_2的血小板变形能力下降,时程延长。(2)MK-447抑制胶原的聚集反应,并使ADP和STA_2聚集增强。(3)胶原和STA_2的释放反应可被MK-447抑制和增强。MK-447对STA_2的作用与S-145无关。结论:血小板变形在其激活早期发挥重要作用。MK-447诱导血小板变形,并对不同聚集剂的作用表现为抑制和增强的双重影响。     

川芎嗪对人类血小板的药理作用

川芎嗪对收缩状态的Salganicoff′s人血小板条有松弛作用,ID₈₀约160μg/ml。明显抑制ADP、花生四烯酸和TXA₂同类物SQ26655所引起的收缩效应。能使血小板cAMP含量升高近1倍,随给药剂量增加,张力继续降低,但cAMP含量并不继续增加。用腺苷环化酶抑制剂SQ 22536后,川芎嗪对钙离子载体(calcium ionophore)A 23187所引起的血小板条收缩效应呈抑制作用,提示川芎嗪松弛血小板条的作用可能与抑制Ca²⁺作用有关。     

异钩藤碱对血小板聚集与血栓形成的抑制作用

目的研究异钩藤碱(isorhynchophylline,Isorhy)对血小板聚集与血栓形成的影响,并探讨其机制。方法以比浊法测定Isorhy体外给药对大鼠血小板聚集的影响;采用动-静脉旁路血栓形成法制作大鼠血栓模型,观察Isorhy对血栓形成的作用;以放免法测定Isorhy对ADP作用下cAMP含量的影响。结果Isorhy0.65mmol.L-1和1.30mmol.L-1对ADP(1.5×10-5mol.L-1)和凝血酶(thrombin,Thr,3U.ml-1)诱导的大鼠血小板聚集均有抑制作用(P<0.01)。静脉注射Isorhy10mg.kg-1和5mg.kg-1可明显降低大鼠血栓形成湿重(P<0.01)。Isorhy0.33~1.30mmol.L-1可升高ADP作用后的血小板cAMP浓度(P<0.01)。结论Isorhy明显抑制血小板聚集与大鼠血栓形成,其抗ADP所致血小板聚集的作用机制至少部分地与升高cAMP水平有关。     

丹七片对动物血小板聚集的抑制作用及其机制研究

目的研究丹七片对家兔和大鼠血小板聚集的影响,并探讨其作用机制。方法以阿魏酸钠为阳性对照,采用比浊法测定丹七片对凝血酶和胶原诱导的血小板聚集的影响,采用酶联免疫法测定丹七片对凝血酶作用下的血小板内环磷酸腺苷(cAMP)含量的影响。结果丹七片可明显抑制由凝血酶和胶原诱导的血小板聚集;丹七片可升高凝血酶作用下的血小板内cAMP含量。结论丹七片抑制由凝血酶和胶原诱导的血小板聚集的作用机制与升高血小板内cAMP的含量有关。     

5HT对STA2介导的血小板聚集和解释反应的双重影响

目的：研究５－ＨＴ对ＳＴＡ２血小板聚集和释放反应的影响及可能的分子机制。方法：以透光法，介质中ＡＴＰ含量及荧光图像法评介血小板变形，聚集反应和［Ｃａ＾］ｉ水平。结果：（１）５－ＨＴ预处理可消除ＳＴＡ２的血小板变形，ＳＴＡ２ ０．３μｍｏｌ·Ｌ＾－１的聚集增强，１－２ｍｏｌ·Ｌ＾－１的聚集不变，释放反应抑制。（２）５－ＴＨ预处理增加ＳＴＡ２ ０．３μｍｏｌ·Ｌ＾－１的［Ｃａ＾２＋］ｉ降低３μｍｏｌ·     

蛋白酪氨酸磷酸化在血小板激活因子诱导血小板信号传导中的调节 …

AIM: To study the role of protein tyrosine phosphorylation (PTP) in platelet activating factor (PAF)-induced platelet signal transduction cascade. METHODS: Washed rabbit platelets were used to test the inhibitory effect of genistein (Gen) on platelet aggregation and serotonin secretion. Intracellular Ca2+ ([Ca2+]i) and pH (pHi) were measured by a dual wavelength fluorophotometer with Fura 2-AM and BCECF-AM. PTP was determined with a specific anti-phosphotyrosine monoclonal antibody by Western blotting. RESULTS: Pretreatment with Gen (100 and 200 mumol.L-1) inhibited PAF (20 nmol.L-1)-stimulated platelet serotonin release by 23.7% +/- 2.0% and 41% +/- 8%, respectively. Similar inhibitory effects of Gen were observed on PAF-evoked increase of [Ca2+]i and intracellular alkalization. PAF also elicited a pronounced increase in PTP of several bands with M(r) 70,000, 60,000, 50,000, 42,000/40,000, and 34,000, which were suppressed markedly by Gen 200 and 400 mumol.L-1. Pretreatment with staurosporine (Sta) 20 nmol.L-1, BAPTA 200 mumol.L-1, and egtazic acid 2 mmol.L-1 to inhibit PKC activation, [Ca2+]i elevation, and Ca2+ influx respectively, also showed an inhibitory effects on the formation of PTP. CONCLUSION: PTP is involved in multiple signal transduction pathways induced by PAF, on which PKC activation and calcium mobilization play a regulatory role.     

普鲁托品对兔血小板内钙的影响

AIM: To study the influence of protopine (Pro) on the cytoplasmic free Ca2+ concentration ([Ca2+]i) in rabbit platelets. METHODS: Measurement of [Ca2+]i of platelets in vitro by Fura 2-AM fluorescence technique. RESULTS: In the presence of CaCl2 1 mmol.L-1, Pro 10, 20, and 40 mumol.L-1 attenuated the rise in [Ca2+]i evoked by ADP from (420 +/- 57) to (320 +/- 26), (264 +/- 21), and (180 +/- 14) nmol.L-1, respectively, by arachidonic acid (AA) from (280 +/- 36) to (210 +/- 17), (184 +/- 21), and (143 +/- 16) nmol.L-1, respectively, and by platelet-activating factor (PAF) from (350 +/- 42) to (282 +/- 31), (223 +/- 30), and (165 +/- 15) nmol.L-1, respectively. In the presence of egtazic acid 1 mmol.L-1, Pro 10, 20, and 40 mumol.L-1 reduced the Ca2+ release induced by ADP, AA, and PAF, respectively. Pro 10, 20, and 40 mumol.L-1 also decreased ADP-, AA-, and PAF-induced Ca2+ influx. CONCLUSION: Pro inhibited not only Ca2+ release but also the influx of Ca2+.     

Effect of 3,6-dimethamidodibenzopyriodonium citrate on Ca2+ in rabbit platelet in vitro.

AIM: To study the effect of 3,6-dimethamidodibenzopyriodonium citrate (I-65) on the cytoplasmic free Ca2+ ([Ca2+]i) concentration in rabbit platelet. METHODS: Measurement of the cytosolic Ca2+ of platelets in vitro by using Quin 2-AM fluorescence technique. RESULTS: In the presence of CaCl2 1 mmol.L-1, I-65 (10, 20, and 30 mumol.L-1) reduced the rise in [Ca2+]i induced by thrombin and calcimycin from 142 +/- 22 nmol.L-1 and 124 +/- 18 nmol.L-1 to 118 +/- 20, 78 +/- 12, 40 +/- 10 nmol.L-1, respectively and 108 +/- 15, 77 +/- 14, 37 +/- 14 nmol.L-1, respectively. In the presence of egtazic acid 2 mmol.L-1, I-65 (10, 20, and 30 mumol.L-1), reduced the Ca2+ release induced by thrombin from 52 +/- 11 nmol.L-1 to 34 +/- 9, 19 +/- 6, and 11 +/- 5 nmol.L-1, respectively. In addition, I-65 (10, 20, and 30 mumol.L-1) also reduced the Ca2+ influx induced by thrombin from 91 +/- 13 nmol.L-1 to 84 +/- 15, 58 +/- 15, and 28 +/- 19 nmol.L-1, respectively. CONCLUSION: I-65 inhibited not only the Ca2+ release, but also the influx of Ca2+ in activation platelet.     

5—HT增强家兔ADP介导的血小板聚集反应

AIM: To study the enhanced effects of 5-hydroxytryptamine (5-HT) on ADP-induced aggregation. METHODS: Platelet aggregation was quantified by the light transmission, the cytosolic-free calcium ([Ca2+]i) was measured by digital fluorescent microscopy, and inositol 1,4,5-triphosphate (IP3) was determined by receptor binding assay. RESULTS: In rabbit platelet-rich plasma (PRP), 5-HT 0.03-3 mumol.L-1 induced a decrease in light transmission (DLT) in a concentration-dependent manner with centralization of granules, as revealed by electron microscopy. The DLT was accompanied with neither platelet aggregation nor a release reaction. In single washed platelets loaded with Fura-2, 5-HT caused a concentration-dependent elevation of [Ca2+]i, and IP3 level was also transiently increased in washed platelets at 15 s after stimulation by 5-HT. Adenosine diphosphate (ADP) also caused DLT transiently in PRP before its own aggregation without a release reaction. Pretreatment of PRP or washed platelets with 5-HT, the DLT by ADP was reduced concentration-dependently and ADP-induced aggregation and [Ca2+]i mobilization were enhanced. CONCLUSION: The enhancement of ADP-induced aggregation was attributed to the superimposition of the calcium release from the storage sites and calcium influx induced by ADP over the calcium release from the storage sites by 5-HT.     

内皮细胞衍生的一氧化氮抑制凝血酶激活的兔血以Na^＋／H^＋交换

AIM: To study the effect of nitric oxide (NO) derived from endothelial cells on Na+/H+ exchange in rabbit platelets activated by thrombin. METHODS: Intracellular Ca2+ ([Ca2+]i) and intracellular pH (pHi) were measured by the dual-wavelength fluorophotometer with the fluorescent probes Fura-2 and 2',7'-biscarboxyethyl-5,6-carboxyfluorescein (BCECF). Effects of NO on rabbit platelets were tested by cultured bovine endothelial cells (BAEC). RESULTS: BAEC (0.1-1 x 10(9).L-1) inhibited thrombin (100 U.L-1)-induced platelet aggregation in a concentration-dependent manner. This inhibiting effect was abolished by preincubating BAEC with NG-nitro-L-arginine 1 mmol.L-1. When the [Ca2+]i store was depleted with ionomycin in the presence of egtazic acid (EGTA), the increase in pHi induced by thrombin was inhibited. Refilling intracellular Ca2+ store partially reversed this effect. BAEC 2 x 10(8).L-1 inhibited thrombin (100 U.L-1)-induced elevation of pHi and mobilization of intracellular Ca2+ store (P < 0.01). No direct effect of endothelial cells on unstimulated rabbit platelets was observed. CONCLUSION: NO derived from endothelial cells inhibited thrombin-induced rabbit platelet activation by inhibiting thrombin-induced [Ca2+]i mobilization and then inhibiting the consequent Na+/H+ exchange in rabbit platelets.     

槲皮素单硫酸酯钠盐对凝血酶诱导的猪血小板聚集的抑制作用

研究槲皮素单硫酸酯钠盐对凝血酶诱导的猪血小板聚集的抑制作用。方法：用比浊法测定血小板聚集,Ｆｕｒａ ２－ＡＭ荧光法检测胞浆游离钙浓度（「Ｃａ＾２＋）ｉ」。用组蛋白ⅢＳ,「γ＾３２Ｐ」ＡＴＰ与蛋白激酶Ｃ酶液一起温育的方法测定ＰＫＣ的活性。用ＳＤＳ－ＰＡＧＥ分离骨架蛋白。     

TMB-8对去甲肾上腺素和BHQ引起新生大鼠单个脑细胞内游离钙增高的影响

应用AR-CM-MIC阳离子测定系统,研究TMB-8对体外新生SD大鼠单个脑细胞内游离钙的抑制作用及其机制。结果表明,在无细胞外钙情况下,静息[Ca²⁺]i为79±13nmol·L^-1。TMB-810,30μmol·L^-1能明显降低静息[Ca²⁺]i。TMB-8100μmol·L^-1对高钾去极化引起的[Ca²⁺]i显著增高无明显影响。在细胞外钙为1.3mmol·L^-1时,去甲肾上腺素诱导的细胞内[Ca²⁺]i升高可部分被TMB-8抑制;TMB-8(30μmol·L^-1)对BHQ引起的[Ca²⁺]i的升高无明显抑制作用。而当细胞外液[Ca²⁺]i为0时,TMB-8几乎完全抑制了去甲肾上腺素和BHQ的作用。提示TMB-8降低脑细胞内游离钙的作用机制是通过促使细胞内钙进入肌浆网以抑制内钙的释放,并通过饱和肌浆网内Ca²⁺间接地阻滞细胞膜钙通道。     

Activation of rabbit platelets by Ca2+ influx and thromboxane A2 release in an external Ca(2+)-dependent manner by zooxanthellatoxin-A, a novel polyol.

1. Zooxanthellatoxin-A (ZT-A), a novel polyhydroxylated long chain compound, isolated from a symbiotic marine alga Simbiodinium sp., caused aggregation in rabbit washed platelets in a concentration-dependent manner (1-4 microM), accompanied by an increase in cytosolic Ca2+ concentration ([Ca2+]i). 2. ZT-A did not cause platelet aggregation or increase [Ca2+]i in a Ca(2+)-free solution, and Cd2+ (0.1-1 mM), Co2+ (1-10 mM) and Mn2+ (1-10 mM) inhibited ZT-A-induced aggregation. SK&F96365 (1-100 microM), a receptor operated Ca2+ channel antagonist, and mefenamic acid (0.1-10 microM), a non-specific divalent cation channel antagonist, inhibited platelet aggregation and the increase in [Ca2+]i induced by ZT-A. 3. Indomethacin (0.1-10 microM), a cyclo-oxygenase inhibitor, and SQ-29548 (0.1-10 microM), a thromboxane A2 (TXA2) receptor antagonist, inhibited platelet aggregation and the increase in [Ca2+]i induced by ZT-A. 4. Methysergide (0.01-1 microM), a 5-HT2 receptor antagonist, inhibited ZT-A-induced platelet aggregation but did not affect the increase in [Ca2+]i induced by ZT-A. 5. Tetrodotoxin (1 microM), a Na+ channel blocker and chlorpheniramine (1 microM), a H1-histamine receptor antagonist, neither affected ZT-A-induced platelet aggregation nor the increase in [Ca2+]i induced by ZT-A. 6. Genistein (1-100 microM), a protein tyrosine kinase inhibitor, and staurosporine (0.01-1 microM), a protein kinase C inhibitor, also inhibited ZT-A-induced platelet aggregation. 7. The present results suggest that ZT-A elicits Ca(2+)-influx from platelet plasma membranes. The resulting increase in [Ca2+]i subsequently stimulates the secondary release of TXA2 from platelets.(ABSTRACT TRUNCATED AT 250 WORDS)