心痛方对心肌缺血模型大鼠MPO 活性 及PS 表达的干预作用

目的探讨心痛方对心肌缺血大鼠模型心肌梗死面积及髓过氧化物酶（MPO）、P 选择素（PS）的干预作 用。方法80 只清洁级SD 大鼠,随机分成假手术组、模型组、心痛方组、欣康组,采用结扎法造成心肌缺血模型,NBT 染色法测心肌梗死面积,比色法检测MPO 活性,免疫组化法检测PS 的表达。结果1.模型组、心痛方组、欣康组心肌梗 死面积、MPO 活性及PS 表达均显著高于假手术组,差异有统计学意义（P<0.01）;2.心肌梗死面积比较,心痛方组、欣康 组分别小于模型组（P<0.05）,差异均有统计学意义;3.MPO 活性比较,心痛方组、欣康组分别低于模型组（P<0.05）,而 心痛方组低于欣康组（P<0.05）,差异均有统计学意义;4.PS 表达比较,心痛方组、欣康组分别低于模型组（P<0.05）,差 异均有统计学意义。结论心痛方能缩小心肌梗死面积,抑制缺血心肌MPO 活性及PS 表达。在抑制MPO 活性方面心 痛方优于欣康。     

FGF-21对大鼠血小板聚集和活化的抑制作用

临床上大部分抗血小板药物存在继发性出血等副作用,亟需寻找一种安全有效的抗血小板药物。二磷酸腺苷(ADP)和花生四烯酸(AA)是介导血小板聚集的主要物质,文章探讨成纤维细胞生长因子-21(FGF-21)是否抑制ADP或AA诱导大鼠血小板聚集和活化。将大鼠分为正常对照组、正常鼠血小板活化组、阿司匹林干预后血小板活化组、FGF-21高、中、低剂量干预后血小板活化组。给药干预后提取各组血小板,分别用ADP或AA处理,观察处理后血小板聚集情况以及P选择素和血栓素(TXB2)表达水平。与正常对照组相比,正常鼠血小板经ADP或AA处理活化后,血小板聚集率显著升高,血浆中P选择素和TXB2含量明显上升;与正常鼠血小板经ADP或AA处理活化后相比,经阿司匹林和FGF-21干预后分别经ADP或AA处理活化后的血小板聚集率显著下降,血浆中P选择素和TXB2含量显著下降;FGF-21干预组经ADP或AA活化后,血小板聚集率、P选择素和TXB2含量下降水平呈明显剂量依赖性。目前国内外尚未发现FGF-21对血小板聚集与活化作用的相关报道,研究首次证明FGF-21具有抑制ADP和AA诱导血小板聚集和活化作用及明显的抗血凝作用,填补FGF-21在抗血凝研究领域空白,为开发安全有效的抗血凝药物提供理论依据。     

Cryopreservation of Canine Platelets

Background: Platelet cryopreservation allows long-term storage and immediate availability of transfusion products.
Hypothesis: The addition of a preparation inhibiting platelet activation (Thrombosol, in 2% dimethyl sulfoxide [DMSO]) will enhance in vitro function and prolong in vivo survival of cryopreserved platelets compared with those preserved in 6% DMSO.
Animals: Thirty-three research dogs.
Methods: Prospective study. Eleven fresh canine apheresis platelet concentrates (PCs) were each split into 3 units: fresh and cryopreserved in 6% DMSO or Thrombosol. Platelet analysis, performed 1–10 weeks postfreezing, included in vitro functional testing and in vivo survival assessed by administration of biotinylated platelets.
Results: Platelet aggregation was diminished in cryopreserved PC. Cryopreserved platelets could be activated, as based on mean thrombin-stimulated P-selectin expression (6% DMSO, 23.0%; Thrombosol, 18.4%), although to a lesser extent than fresh PC (49.1%) ( P < .0001). The mean maximum in vivo platelet recovery for fresh PC was 80.3%, significantly greater than recovery for 6% DMSO (49.2%) and Thrombosol PC (43.7%) ( P ≤ .001). The half-life (days) of fresh PC (3.8 ± 0.4) was significantly ( P < .002) greater than that of 6% DMSO (1.9 ± 1.0) and Thrombosol (2.4 ± 1.1) PC, with no difference ( P = .3) between cryopreserved PC.
Conclusions and Clinical Importance: Cryopreservation of canine platelets using Thrombosol did not provide any advantage over preservation using 6% DMSO. Cryopreserved platelets can be activated in vitro and provide therapeutic benefit when fresh platelets are unavailable. Further studies are needed to assess their in vivo hemostatic function.     

复方丹参注射液对急性水肿型胰腺炎患者可溶性粘附分子水平的影响

目的:研究复方丹参注射液对急性水肿型胰腺炎患者外周血粘附分子水平的影响,探讨复方丹参注射液治疗急性胰腺炎的机制。方法:采用复方丹参注射液结合西药治疗32例急性水肿型胰腺炎患者(治疗组),取常规西药治疗的35例急性水肿型胰腺炎患者(治疗对照组)作为对比观察,并于治疗前后检测患者血液中粘附分子P-选择素(P-sel)和细胞间粘附分子-1(ICAM-1)水平。结果:治疗组的腹痛缓解时间、尿淀粉酶恢复时间和平均住院时间均短于治疗对照组(P<0.05或0.01),临床疗效也优于治疗对照组(H c=5.7902,P<0.05)。治疗前治疗组和治疗对照组的P-sel、ICAM-1比较差异无显著性(P>0.05),治疗后治疗组的P-sel下降较明显,与治疗对照组比较差异有非常显著性(P<0.01),两组治疗后ICAM-1水平差异未见显著性(P>0.05)。结论:复方丹参注射液结合西药治疗对急性胰腺炎有显著疗效,可能与复方丹参注射液能调节免疫细胞粘附分子表达,抑制炎症细胞聚集,进而抑制炎症反应有关。     

Effects of ambroxol combined with low-dose heparin on production of ICAM-1 and E,P-selectin in acute lung injury

AIM: To study the production of intercellular adhesion molecule-1(ICAM-1), E-selectin and P-selectin in serum, lung tissues and bronchoalveolar lavage fluid(BALF)of acute lung injury(ALI) model and to observe the effects of ambroxol combined with low-dose heparin on the changes of the 3 factors above.METHODS: Twenty-four healthy rabbits were randomly divided into 3 groups: normal saline control group (NC), oleic acid injury group (OA), ambroxol+ heparin treatment group (AH). The rabbit ALI model was induced by oleic acid injection through auricular vein. Partial pressure of O₂ in artery(PaO₂) was analyzed.The concentrations of ICAM-1 and E-selectin were detected by ELISA.The apoptosis index(AI) was measured by TUNEL method.The expression of P-selectin was determined by immunohistochemical method.The ultrastructural changes of the lung tissues were observed under electron microscope, and the lung wet/dry ratio(W/D) was calculated.RESULTS: PaO₂ in AH group and OA group was significantly lower (P<0.01) than that in NC group, and PaO₂ in AH group was significantly higher than that in OA group (P<0.01). The concentrations of ICAM-1 and E-selectin in serum, lung tissues and BALF, and AI and W/D in lung tissues in AH group were higher (P<0.05 or P<0.01) than those in NC group, and was lower than those in OA group (P<0.05 or P<0.01). In NC group, no significant change of the above parameters at all time points was observed (P>0.05). In OA group, PaO₂ was significantly decreased (P<0.01) with the pathological process developed, and the concentrations of ICAM-1 and E-selectin were significantly increased. In AH group, PaO₂ was decreased (P<0.05),and the concentrations of ICAM-1 and E-selectin were increased with the process of ALI developed. The P-selectin expression in lung tissues of OA group was distributed mainly in inflammatory cells, capillary endothelial cells and plasma. From low to high levels, the order was NC group < AH group < OA group in the expression of P-selectin. The most obvious apoptosis was observed in OA group. No apoptosis or occasional positive cells were found in NC group. The apoptotic rate in AH group was significantly reduced compared with that in OA group.CONCLUSION: In ALI induced by OA, ICAM-1, E-selectin and P-selectin are significantly increased and are involved in the occurrence and development of ALI. Ambroxol combined with low-dose heparin reduces the levels of ICAM-1, E-selectin and P-selectin, the pulmonary edema and the lung injury, improves pulmonary functions, and plays an important role in the prevention and treatment of acute lung injury.     

Flow cytometric detection of activated platelets in the dog

Background: Platelet activation appears to play a role in a variety of canine thrombotic disorders. At present, tests for the detection of activated platelets are not used routinely in veterinary clinical laboratories. Objective: The purpose of this study was to develop a clinically applicable method to detect activated canine platelets. Methods: A flow cytometric assay was developed to detect activated platelets, platelet aggregates, and platelet microparticles in the dog. Blood was collected from healthy dogs using EDTA or sodium citrate as the anticoagulant, and platelet‐rich plasma was prepared. Platelets were activated by adding phorbol myristate acetate. In some experiments, platelets were fixed by incubation with 0.5% paraformaldehyde. In other experiments, platelets were stored for 4 or 24 hours at 4°C before analysis. Activated platelets were detected by measuring surface expression of P‐selectin and by determining the percentages of platelet aggregates and microparticles using forward‐angle vs side‐angle light scatter plots. Results were analyzed by using 2‐way ANOVA and the SchefféF‐test. Results: Platelets collected in EDTA had minimal expression of P‐selectin, whereas platelets collected in sodium citrate had greater median fluorescence intensity. Fixation with 0.5% paraformaldehyde before labeling platelets with anti‐P‐selectin did not affect antibody binding or the percentages of platelet aggregates and microparticles. Storage of platelet‐rich plasma at 4°C for 4 hours did not affect antibody binding or the percentages of platelet aggregates or microparticles. Activation of platelets ex vivo by addition of 10 ng/mL phorbol myristate acetate resulted in a large increase in expression of P‐selectin but only slight increases in platelet aggregates and microparticles. Conclusion: Determination of platelet P‐selectin expression and percentages of platelet aggregates and platelet microparticles may provide a clinically applicable means for detection of activated platelets in dogs. The capacity to use EDTA‐anticoagulated blood samples and to fix platelets for evaluation at a later time makes the test attractive as a routine diagnostic tool.     

Progress of P-selectin glycoprotein ligand-1 in kidney diseases

P-selectin glycoprotein ligand-1 (PSGL-1) is an adhesion molecule mainly expressed on the surface of leukocytes and platelets, which plays a vital part in immune recognition, inflammatory response and thrombosis. The prevalence of chronic kidney disease (CKD) is increased gradually and it has been one of major diseases that threaten the world's public health. In addition, its etiology is complicated, and most of the pathogenesis is incompletely understood. Researches show that PSGL-1 participates in the development of kidney diseases in a variety of ways, and its mechanism may involve signaling pathways such as TGF-β1/Smad and PI3K/AKT/GSK-3β, as well as key renal regulators such as connective tissue growth factor and chemokine (C-C motif) ligand 2. This review summarizes the research progress of PSGL-1 in renal fibrosis, lupus nephritis, obesity-related kidney disease and acute kidney injury.     

Alterations in normal canine platelets during storage in EDTA anticoagulated blood

Abstract: Flow cytometric detection of platelet surface-associated IgG (PSAIgG) can be used to determine whether immunologic factors are contributing to thrombocytopenia in dogs. In vitro alterations in platelet activation and morphology, however, could impact the results of this test. The purpose of this study was to determine whether the PSAIgG test for immune-mediated thrombocytopenia was valid on whole blood in EDTA anticoagulant after 24–72 hours of storage, and to characterize other alterations in canine platelets that could impact immunologic testing. Platelets were harvested and analyzed immediately after blood collection and after 24, 48, and 72 hours of storage at 4°C. Spontaneous and thrombin-induced changes in the following platelet parameters were evaluated using flow cytometric techniques: PSAIgG, platelet microparticle formation, membrane expression of P-selectin and glycoprotein CD61, exogenous IgG binding, surface-exposed phosphatidylserine, and fibrinogen binding. The amount of PSAIgG increased 6-to 9-fold in stored samples compared with fresh samples. Platelet microparticle formation was spontaneous in stored samples and increased significantly over time. Membrane phosphatidylserine, P-selectin, and fibrinogen binding were not altered by storage, indicating that platelet activation was minimal in stored samples. Although storage decreased the percentage of platelets positive for CD61 by 8-to 10-fold compared with fresh samples, activation by high-dose thrombin partially restored the percentage of CD61-positive platelets in 24-hour-old samples. In conclusion, even though platelets stored in EDTA for up to 72 hours remain in a resting state, aged platelets have an increased tendency to form microparticles and have increased surface IgG and decreased surface CD61, which may contribute to false-positive results for tests of immune-mediated thrombocytopenia.     

Effect of N,N-dimethylsphingosine on the expression of adhesion molecules in human umbilical vein endothelial cell line EA.hy926

AIM: To investigate whether and how N, N-dimethylsphingosine (DMS) plays a role in modulating the adhesion of monocytes to vascular endothelial cells, and identify whether human umbilical vein endothelial cell line EA.hy926 take place of the vascular endothelial cells.METHODS: Adhesion ratio was measured by flow cytometry, and immunohistochemistry was used to detect the expression of ICAM-1 and P-selectin in HUVEC: EA.hy926 cells after the effect of DMS. RESULTS: DMS inhibited the adhesion of monocytes to HUVEC: EA.hy926 cells in a time-dependent and concentration-dependent manner by reducing the expression of ICAM-1 and P-selectin. CONCLUSIONS: DMS reduced adhesion molecule expression in vascular endothelial cells. DMS may be an important contributor to reduce adhesion ratio, suggesting that DMS plays a negative role in proinflammatory and immune functions of the modified vascular endothelial cells during atherosclerosis and restenosis.