Characterization of Platelet Activating Factor (PAF)Binding Sites in Rabbit Platelet Membranes

本文用放射配基受体结合实验方法研究了兔血小板膜上血小板活化因子（PAF）受体的性质。[^3H]-PAF与膜的特异结合率达70－80％，在25℃测得PAF与膜结合的平衡解离常数（Kd）为40.8nmol/L，最大结合位点为5.4pmol/mg蛋白。特异的PAF受体拮抗剂海风藤酮对结合有较强的抑制作用，IC50为30.0μmol/L。（－）－denudatin B，樟叶素(polysyphorin）和南藤素（wallichinine）是从中草药中分离的化合物，均能抑制这一结合，其IC50分别为0.6μmol/L、10μmol/L和2.5μmol/L。

Effect of Xiaoyu Zhixue Tablet (消瘀止血片) on Expression ofPlatelet Membrane Glycoproteins in Patients withHemorrhagic Thrombopathy

Objective: To observe the effect of Xiaoyu Zhixue tablet (消瘀止血片,XYZXT) on the expression of platelet membrane glycoproteins in patients with hemorrhagic thrombopathy, and to explore its possible mechanism. Methods: The total of 148 patients with hemorrhagic thrombopathy were randomly divided into two groups, the traditional Chinese medicicne (TCM) group (n=98) treated with XYZXT and the Western medicine (WM) group (n=50) treated with adrenosin, vitamins C, K and P, both for 6 months. The therapeutic effect and the recovery rate of platelet aggregation in the two groups were observed. And platelet membrane glycoprotein (GP) Ⅰb/Ⅸ, GPⅡb/Ⅲa complexes, GPⅠb, GPⅡb, GP Ⅲa and P-selectin were analyzed by flow cytometry in both groups before and after treatment and also in 34 normal healthy subjects. Results: The total effective rate of hemostasis was 89. 8% in TCM group and 54. 0% in the WM group (x2=45.83, P<0.01), and the recovery rate of platelet aggregation was 72.4% and 4.0% respectively (x2=62.06, P<0.01). The fluorescence intensity of GP Ⅰ b/Ⅸ, GPⅡb/Ⅲa complexes, GPⅠb, GPⅢa and P-selectin were lower in both groups before treatment than those in the healthy subjects. Expression of above-mentioned marks was elevated in TCM group after 6 months' therapy, which was insignificantly different as compared with the healthy subjects (P>0.05) and higher than those in the WM group (P<0.05). Conclusion: One of the mechanisms in treating hemorrhagic thrombopathy with XYZXT is that it could regulate the expression of GP Ⅰb/Ⅸ, GPⅡ b/Ⅲa complexes, GPⅠb, GPⅢa and P-selectin at the level of receptor protein.

Expression of Platelet Membrane Glycoprotein Ib/Ⅸ/Ⅴ Complex,a Receptor of Thrombin,in Patients with Hemorrhagic Thrombopathy

To investigate the role of platelet membrane glycoprotein （GP） Ib/Ⅸ/Ⅴ complex and its subunit GP Ibα in patients with hemorrhagic thrombopathy （HT）, the expressions of GP Ib/Ⅸ/Ⅴ complex and GP Ibα,defined as mean fluorescence intensity （MFI）, were assessed by flow cytometry. The maximum aggregation of platelet was determined by turbidity method. These indicators were compared among 68 HT patients with the presenting complaint of hemorrhage, 33 well-controlled HT patients and 32 normal healthy subjects. The results showed that the MFI of GP Ib/Ⅸ /Ⅴ complex and GP Ibα was markedly lower in HT patients with current hemorrhage than that in the healthy subjects, with difference being statistically significant （P〈0.05）. There was no significant difference in the expressions of GP Ib/ Ⅸ/ Ⅴ complex and GP Ibα between well-controlled HT patients and normal healthy subjects （P〉0.05）. It was concluded that the expression of GP Ib/Ⅸ /Ⅴ complex, the receptor of thrombin and von Willebrand factor, was down-regulated in HT patients with current hemorrhage, which might result in the dysfunction of platelet aggregation and recurrence of HT.

Association of the Platelet Membrane Glycoprotein Ⅰa C807T Gene Polymorphism with Aspirin Resistance

To explore the correlation between the C807T polymorphism of platelet membrane glycoprotein Ⅰa （GP Ⅰa） gene and aspirin resistance in Chinese people, 200 patients with high-risk of atherosclerosis took aspirin （100 mg/d） for 7 days. Platelet aggregation function was detected using adenosine diphosphate （ADP） and arachidonic acid （AA） before and after the administration of aspirin. Then the subjects were divided into three groups according to the results of platelet aggregation function： an aspirin resistant （AR） group, an aspirin semi-responder （ASR） group and an aspirin-sensitive （AS） group. Platelet GP Ⅰa gene 807CT polymorphism was examined by means of polymerase chain reaction-sequence specific primers （PCR-SSP）. The results showed that T allelic frequency in AR group and ASR group were higher that of AS group （P〈0.005）, and the prevalence of genotypes （TT＋TC） of these two groups was significantly higher than that in AS group （P〈0.05）. Platelet GP Ⅰa T allele was significantly associated with aspirin resistance as revealed by multiple logistic regression （OR=3.76, 95% CI： 2.87-9.58）. The results suggest that inherited platelet GP Ⅰa variations may have an important impact on aspirin resistance and the presence of GP Ⅰa T allele may be a marker of genetic susceptibility to aspirin resistance.

Redistribution of Platelet Membrane Glycoprotein IV and Release of Intracellular α-granule Thrombospondin in Patients with Chronic Myelogenous Leukemia

The redistribution of platelet membrane glycoprotein IV (GPIV) and the release of intracellular Q-granule thrombospondin (TSP) were examined and the inhibition of 5-thromboglobulin (&TG) and platelet factor 4 (PF4) in patients with chronic myelogenous leukemia (CML) was observed and quantitation of β-TG and PF4 in sera was conducted. GPIV in inactive platelet from CML was 36080±17010 molecules/platelet as compared with 13190±4810 from the controls (P<0,01), No abnormality was found in the distribution of platelet membrane GPIb and GPIIb/III.(P>0. 05). The GPIV redistribution on active platelet membrane induced thrombin (1U/ml) from CML and healthy donors was 44320132310 and 228001 12700 molecules/platelet respectively (P<0. 01 ). The difference in the release of intracellular Q-granule TSP between CML and the control group was not found (P>0.05). There was no direct correlation between GPIV expression and TSP binding after platelet activation. The high leveIs of β-TG and PF4 in sera inhibited release of intracellular a-granule TSP in vitro. These results indicate that the abnormality of platelet membrane GPIV is a common marker in CML, therefore the specific increase of platelet GPIV in patients with CML may be a useful tool for the diagnosis and monitoring of the platelet dysfunction. The release of interna1 TSP pools is hindered by either β-TG or PF4 in sera.

Effect of Xiaoyu Zhixue Tablet(消瘀止血片) on the Expression of Platelet Membrane Glycoprotein Ⅰb/Ⅸ/Ⅴ Complex in Patients with Chronic Renal Failure

Objective： To investigate the effect of Xiaoyu Zhixue Tablet （消瘀止血片, XYZXT） on the expression of platelet membrane glycoprotein （GP） Ⅰ b/Ⅸ/Ⅴ complex and GP Ⅰ b α in patients with chronic renal failure （CRF） in early metaphase. Methods： Fifty-one patients with CRF in early metaphase （treated group） were treated with XYZXT, 3 months as the course of treatment for 2 courses. The previous therapies remained unchanged. Flow cytometry was used to assess the expression of platelet GP Ⅰb/Ⅸ/Ⅴ complex and GP Ⅰb α in patients with CRF, and turbidity method was used to determine the platelet maximum aggregation rate （MAR）, meanwhile the renal function was measured. The final data were compared with those before the treatment, and with those in the normal control group （31 healthy subjects）. Results： Compared with the normal control group, expressions of GP Ⅰ b/Ⅸ/Ⅴ complex and GPⅠb α, and platelet MAR in CRF patients were significantly lower （P=0.007,P=0.001,P=0.009） before the treatment; after the treatment with XYZXT, the above indexes in CRF patients were remarkably increased （P=0.033,P=0.026, P=0.045）, but still lower than those in the normal control group, however, it was not statistically significant. Conclusion： （1） The expression of GP Ⅰ b/Ⅸ/Ⅴ complex in CRF patients of early metaphase was decreased, which lead to platelet aggregation dysfunction. This might be one of the reasons for the hemorrhagic trend in CRF. （2） XYZXT was able to upgrade expressions of GP Ⅰb/Ⅸ/Ⅴ complex and GPⅠb α in CRF patients, improve platelet function and clown-regulate platelet activation in patients with CRF.

Effect of Dauricine on Redistribution of Glycoprotein Ⅳ in Platelet Membrane of Patients with Mitral Stenosis

Objective: To investigate the effect of dauricine on the irreversible platelet aggregability of patients with mitral stenosis (MS).Methods: Glycoprotein Ⅳ (GPⅣ ) and thrombospondin (TSP) levels on the membrane surface of the stationary platelet or platelet activated by thrombin (0. 05 U/ml, 0. 1 U/ml,0. 5 U/ml, 1. 0 U/ml) in 16 patients with MS were measured with flow cytometric method and compared with those of the healthy (14 subjects). Results: The GPⅣ level of stationary platelet, the GPⅣ and TSP level of activated platelet in MS patients were higher than those in the healthy significantly (P < 0. 01,< 0. 05, < 0. 005 ), while the TSP level of stationary platelet in the patients was not different to the healthy (P > 0. 05). The GPⅣ redistribution on the activated platelet surface was apparently inhibited by dauricine (50 μmol/L, P < 0. 05 - 0. 005) and the release of TSP from intracellular α-granules was inhibited by dauricine only in the activated platelets induced by thrombin of low concentration (0. 05 U/ml and 0. 1 U/ml, P < 0. 05 - 0. 01 ), inhibiting effect was not found in those activated with high concentration of thrombin. Conclusion: The activity and reactivity to thrombin of platelets increased in MS patients, and dauricine was able to reduce the occurrence of the irreversible platelet aggregation in MS patients.Original article on CJIM(Chin) 1998; 18(8): 461

The Relationship Between P-Selectin on Platelet Membrane and Platelet Function in Chronic Cor Pulmonale.

Platelet plays an importent role in thromboembolic diseases. It is demonstrated that recurrent pulmonary embolism may cause pulmonary hypertention. In recent years，P-selectin（granule membrane protein，GMP40）was found to be a surface marker for platelet activation.

Platelet activation and the protective effect of aprotinin in hepatolithiasis patients

OBJECTIVE: To explore platelet activation and the protective effect of aprotinin in patients with hepatolithiasis. METHODS: The count of plaletets and levels of CD_(62P) and CD_(63) were measured by flow cytometry in 38 patients with hepatolithiasis. Several measurements were carried out after treatment with aprotinin. RESULTS: The levels of CD_(62P), CD_(63) in patients with hepalolithiasis were higher than those in patients with cholecystolithiasis (P<0.05), but the count of platelets was lower (P<0.05). After operation, the levels of CD_(62P), CD_(63) were significantly increased in patients with hepatolithiasis, but the count of platelets was lower (P<0.05). Postoperative levels of CD_(62P), CD_(63) were significantly lower in patients treated with aprotinin than in normal controls (P<0.05); but there was no significant change in the count of platelets in the two groups. CONCLUSION: Platelet activation occurs in patients with hepatolithiasis, and may be inhibited by aprotinin.

Diagnosis of Disorders of Platelet Function by Flow Cytometry

The platelet membrane glycoprotein (GP)Ⅱ b/Ⅲ a, GPIX were detected by immunogold assay and flow cytometry, respectively, in two cases of Glannzmann's thrombasthenia (GT). The immunogold assay showed that the GPⅡ b/Ⅲ a gold granules in GT were decreased obviously, compared with that in normal controls. Flow cytometry results showed that GPⅡ b/Ⅲ a in two cases of GT was 0.1 % and 0.5 % of normal control, respectively. While GPIX showed no difference between the GT and normal control. The patients' father showed no bleeding symptoms, whose GPⅡ b/Ⅲ a accounted for 36.99 % of normal control. The experiment results suggested that flow cytometry is a quick, simple and sensitive method for the diagnosis of GP disorders.

Effects of Xuezhikang Capsule(血脂康胶囊) on Blood Lipids,Platelet Activation and Coagulation-Fibrinolysis Activity in Patients with Hyperlipidemia

Objective: To investigate the effects of Xuezhikang capsule (XZK,血脂康胶囊) on blood lipids level, platelet activation and coagulation-fibrinolysis activity in patients with hyerlipidemia. Methods: Seventy-six patients of hyperlipidemia were randomly divided into two groups, the XZK group (n=38) treated with XZK 600mg, taken two times per day and the Simvastatin (SIM) group (n = 38) treated with SIM 20mg per day, with the treatment lasting 8 weeks for both groups. Levels of fasting serum lipids, including total cholesterol (TC), triglyceride (TG), high and low density lipoprotein cholesterol (HDL-C and LDL-C), plasma GMP-140, fibrinogen (FIB), tissue plasminogen activator (t-PA), plasminogen activator inhibitor type-1 (PAI-) and prothrombin time (PT) were all measured before and 8 weeks after treatment. Data were compared before and after treatment and also compared with those measured in 20 healthy subjects of control. Results: Before treantment the levels of TC, TG and LDL-C were obviously higher and HDL-C level was significantly lower in hyperlipidemia patients than those in healthy subjects (P<0.05 or P<0.01). After 4-8 weeks of XZK treatment, the levels of TC, TG, LDL-C and FIB and activities of GMP-140 and PAI-1 were obviously lowered (P<0. 05 or P<0. 01). But in the SIM group there was no obvious change in FIB (P>0.05), instead it showed obvious increase of HDL-C and decrease of TC and LDL-C after treatment (P<0.05 or P<0.01). Conclusion: XZK could inhibit platelet activity and improve coagulation-fibrinolysis function, besides its lipids lowering effect.

Clinical Observation on Shu Xin Yi Mai Capsules for Prevention of Recurrent Stricture after Coronary Artery Introducing Treatment

Shu Xin Yi Mai Capsules (舒心益脉胶囊) combined with western medicine was used in the routine treatment of 22 cases who successfully received coronary artery introducing therapy (Chinese-western medicine group),and the results was compared with the 26 cases treated routinely with simple western medicine in the control group (western medicine group).It was found that both the recurrence rate of angina pectoris and the incidence rate of recurrent stricture in the Chinese-western medicine group were significantly lower than that in the control group (both P<0.05).There was no significant difference between the two groups in expression of platelet activating molecules CD62P (a-granular membrane protein),CD63 (lysosome intact membrane protein) and CD41 (glucoprotein IIb) before the treatment,but with a significant difference after the treatment (P<0.05).

NIR light-induced tumor phototherapy using ICG delivery system based on platelet-membrane-camouflaged hollow bismuth selenide nanoparticles

Near-infrared(NIR)light-triggered photothermal therapy(PTT)is a promising treatment strategy for treating cancer.The combination of nanotechnology and NIR has been widely applied.However,the therapeutic efficacy of the drug-delivery system depends on their ability to avoid phagocytosis of endothelial system,cross the biological barriers,prolong circulation life,localize and rapidly release the therapeutic at target sites.In this work,we designed a platelet membrane(PM)-camouflaged hollow mesoporous bismuth selenide nanoparticles(BS NPs)loading with indocyanine green(ICG)(PM@BS-ICG NPs)to achieve the above advantages.PM-coating has active tumor-targe ting ability which could preve nt drug leakage and provide drug long circulation,causing drug delivery systems to accumulate in tumor sites effectively.Moreover,as a type of the photothermal sensitizers,BS NPs are used as the inner cores to improve ICG stability and are served as scaffolds to enhance the hardness of this drug delivery system.For one hand,the thermal vibration of BS NPs under NIR laser irradiation causes tumor inhibition through hyperthermia.For another hand,this hyperthermia process could damage PM and let ICG rapid release from PM@BS-ICG NPs.The in vitro and in vivo results showed that this biomimetic nano-drug delivery system exhibits obvious antitumor activity which has good application prospect.

Effect of the Principle of Activating Blood Circulation to Break Stasis on GMP-140 and D_2 Dimer in Patients with Acute Cerebral Infarction

Objective: To explore the clinical efficacy of the principle of activating blood circulation to break stasis (ABCBS) and its influence on platelet membranous protein particle (GMP-140) and D 2 dimer (D-dimer) before and after treatment. Methods: Eighty-eight patients with blood stasis syndrome (BSS) of acute cerebral infarction (ACI) were randomly divided into two groups, both of which were treated with conventional treatment, i.e. with western medicine (WM), with Salvia injection added through intravenously dripping.One of the two groups was used as the control and the other group as the treated group who had ABCBS herbs orally taken in addition. The duration of treatment course for both groups was 3 weeks. Results: There were changes in both groups over clinical symptoms, nerve function deficit scoring and GMP-140, D-dimer, but the treated group showed significantly better than that of the control group, ( P <0.05). Conclusion: ABCBS principle could serve as an important auxiliary treating method for BSS of ACI, as it can effectively alter the blood of ACI patients which was viscous, condense, coagulant and aggregating.

Leukocyte/platelet hybrid membrane-camouflaged dendritic large pore mesoporous silica nanoparticles co-loaded with photo/chemotherapeutic agents for triple negative breast cancer combination treatment

Triple-negative breast cancer(TNBC)is an aggressive subset of breast cancer and currently lacks effective therapeutic targets.As two main phototherapeutic methods,photothermal therapy(PTT)and photodynamic therapy(PDT)show many advantages in TNBC treatment,and their combination with chemotherapy can achieve synergistic therapeutic effects.In the present study,a biomimetic nanoplatform was developed based on leukocyte/platelet hybrid membrane(LPHM)and dendritic large pore mesoporous silicon nanoparticles(DLMSNs).A near infrared(NIR)fluorescent dye IR780 and a chemotherapeutic drug doxorubicin(DOX)were co-loaded into the large pores of DLMSNs to prepare DLMSN@DOX/IR780(DDI)nanoparticles(NPs),followed by camouflage with LPHM to obtain LPHM@DDI NPs.Through the mediation of LPHM,LPHM@DDI NPs showed an excellent TNBC-targeting ability and very high PTT/PDT performances in vitro and in vivo.Upon NIR laser irradiation,LPHM@DDI NPs exhibited synergistic cytotoxicity and apoptosis-inducing activity in TNBC cells,and effectively suppressed tumor growth and recurrence in TNBC mice through tumor ablation and anti-angiogenesis.These synergistic effects were sourced from the combination of PTT/PDT and chemotherapy.Altogether,this study offers a promising biomimetic nanoplatform for efficient co-loading and targeted delivery of photo/chemotherapeutic agents for TNBC combination treatment.

妇宝胶囊止血机制的实验研究

目的:研究妇宝胶囊的止血机制。方法:大鼠给药后测定组织纤溶酶原激活物(t-PA)和纤溶酶原激活物抑制物(PAI-1)的活性;家兔给药后测定其血小板聚集率和血小板α-颗粒膜蛋白-140(GMP-140)的含量。结果:给予大鼠妇宝胶囊内容物4.0、8.0g/(kg·d),家兔3.0、6.0g/(kg·d),共6d,后测定大鼠t-PA的活性降低(P<0.01)、PAI-1的活性提高(P<0.01),但家兔血小板聚集和血小板α-颗粒膜蛋白-140的含量未产生明显的影响(P>0.05)。结论:妇宝胶囊的止血机制之一是通过抑制纤维蛋白溶解系统达到目的。

缺血性脑血管疾病患者血小板膜糖蛋白变化及临床意义

目的探讨血小板活化与缺血性脑血管疾病的关系及血小板活化因子在脑梗死中的重要意义。方法对2003年9月至2004年11月石家庄市第二医院神经内科收治的71例缺血性脑血管病患者和20名健康体检者,采用酶联免疫吸附双抗体夹心法测定循环中血小板α颗粒膜糖蛋白-140(GMP-140)、血小板膜糖蛋白(GP)Ⅱb/Ⅲa、GPⅠb的表达水平。结果GMP-140、GPⅡb/Ⅲa在脑供血不足、轻度脑梗死、中重度脑梗死各组间依次递增,中重度脑梗死组GPⅠb低于正常对照组(P<0.05),脑供血不足组GPⅠb高于正常对照组(P<0.05)。结论在缺血性脑血管疾病的不同阶段均存在血小板活化,GMP-140、GPⅡb/Ⅲa、GPⅠb均可作为血小板活化的分子标志物,为预防、诊断脑梗死及监测脑梗死的严重程度提供重要参考指标。

重型颅脑损伤患者凝血纤溶标志物动态观察(英文)

Objective To investigate the dynamic changes and the clinic significance of coagulation and fibrinolysis markers in patients with severe craniocerebral trauma.Methods Thirty three consecutive patients [Glasgow coma scale (GCS) ≤8 and APACHE Ⅱ≥10] including 10 women and 23 men with age of (46.3±16.3) years old and with severe craniocerebral trauma were enrolled in this study.Endothelin(ET),alpha-granular membrane protein-140(GMP-140) and D-dimer(DD) were measured at 1st day,3rd day and 7th day of admission,head computerized tomography(CT) was performed on each patient.Twenty healthy people with similar age were as control.ET was determined by the reagent kit which from Shanghai Shenjia company and GC-1200 γ radiatory counter.GMP-140 was determined by the reagent kit which from Suzhou university and PEKINEINMER WIZARD-1470 γ radiatory counter.DD was determined by the reagent kit Shanghai sung biological product company limited and measured by latex agglutination.Results ET,GMP-140 and DD levels were significantly increased at the time of admission[(75.24±26.44)pg/ml,(26.43±13.94)ng/ml,(3.20±0.97)μg/ml,respectively],then gradually decreased.ET and DD levels were significantly higher than control group even after 7 days[(44.66±15.25)pg/ml,(1.35±0.55) μg/ml,respectively],but not for GMP-140.All of the coagulation and fibrinolysis abnormalities markers were negatively correlated with GCS,P<0.05.There were 18 survivors including 8 vegetables and 15 dead including 1 case suffered from disseminated intravascular coagulation(DIC) in this investigation.Conclusions Coagulation and fibrinolysis abnormalities markers were occurred at the very early stage in the patients with severe craniocerebral trauma.The levels of ET,GMP-140 and DD levels were negatively correlated with GCS.