Study on the Mechanism of the Annexin I -Mediated Co-Assembly of t-PA and Plasminogen

In order to further investigate the effect of annexin Ⅱ (Ann- Ⅱ ) on tissue plasminogen activator (t-PA)-dependent plasminogen (PLG) activation and its interactive mechanism, recombinant native Ann- Ⅱ bound t-PA, PLG and plasmin with high affinity was examined. The flow cytometric assay showed that the ann- Ⅱ expression rate was higher in the human umbilical vein endothelial cell (HUVEC) (87. 65 %) than in the HL-60 cells as controls (35. 79 %). Two irrelevant proteins,bovine serum albumin (BSA) and equine IgG (EIG) had no effect on the production of plasmin.Ann- Ⅱ -mediated enhancement of t-PA-dependent PLG activation was inhibited by ε-aminocaproic acid or by pretreatment of Ann- Ⅱ with carboxypeptidase B with the inhibitive rate being 77.8 % and 77. 0 %, respectively. It was revealed that the effect of Ann- Ⅱ on PLG activation was specific for tPA. Urokinase didn‘t bind to Ann- Ⅱ , demonstrating the role of receptor-related lysine residues on activation of PLG, showing that the Ann- Ⅱ -PLG interaction was dependent upon carboxyl-terminal lysine residues. These findings suggest that annexin Ⅱ -mediated co-assembly of t-PA and PLG may promote plasmin generation and play a key role in modulating fibrinolysis on the endothelial surface.     

高脂血症患者血小板体积及有关参数的研究

本文报道242例高脂血症患者血小板(PLT)、平均血小板数(MPLT)及平均血小板体积(MPV)等参数的测定结果,其中高胆固醇并高甘油三酯患者,PLT、MPLT及巨大血小板比例(Macro-PLT)高于正常,MPV正常;而单纯高胆固醇或高甘油三酯患者,上述指标与正常人无显著性差异。提示高胆固醇并高甘油三酯患者,存在血小板生成动力学异常,其血小板的破坏增加,生成率加速、但处于一种高水平的动态平衡之中。这可能是高脂血症出现高凝状态的原因之一。同时表明此类患者使用抗血小板疗法具有一定的合理性和必要性。     

血浆D二聚体和GMP-140测定在DIC诊断中的意义

目的：探讨分子标志物在弥散性血管内凝血（DIC）诊断中的价值。方法：分别检测DIC患者20例（DIC组）、DIC疑诊组11例（DIC疑诊者）和正常人10例（正常对照组）血浆D二聚体和血小板颗粒膜蛋白（GMP－140）水平。结果：以上三组血浆D二聚体水平分别为4．43±3．43，2．03±0．17，0．36±0．17（mg／L）。血浆GMP－140水平分别为53．78±27．55，33．32±16．50，17．29±4．84（ng／ml）。DIC组血浆D二聚体和GMP－140显著高于DIC疑诊组和正常对照组，而DIC疑诊组与正常对照组差异尤显著性。动态观察发现，血浆D二聚体升高的6例DIC疑诊者有3例随后转变成DIC。结论：血浆D二聚体和GMP－140是DIC时敏感指标，监测二指标有助于DIC的早期诊断。     

INCREASED APOPTOSIS IN LYMPHOMA BONE MARROW ERYTHROID CELLS： POSSIBLE ANEMIC PATHOPHYSIOLOGIC SIGNIFICANCE

Objective:To investigate whether excessive erythroid cell apoptosis and/or suppression of proliferation would be implicated in the pathogenesis of anemia associated with lymphoma. Methods: Bone marrow (BM) erythroid cell (CD71^ , GPA^ ) apoptosis(annexin V) and proliferation (Ki-67) were measured by flow cytometry in lymphoma patients and normal donors. Results: BM CD71^ and GPA cells showed a significantly greater proportion of apoptotic cells in lymphoma than in normal samples. Proliferation rate was also significantly increased in lymphoma BM CD71^ and GPA^ cells than in normal samples. Conclusion:Increased apoptosis in lymphoma BM erythroid cells may contribute to the anemia characteristic of this disorder. Proliferation might be up-regulated on BM erythroid cells via negative feedback loop resulting from a state of apoptosis.     

骨髓或外周血中CD4+CD25+Foxp3+调节性T淋巴细胞与aGVHD的关系

目的 研究患者异基因造血干细胞移植(allo-HSCT)前后骨髓或外周血单个核细胞中CD4+CD25+Foxp3+调节性T淋巴细胞(以下简称CD4+CD25+Foxp3+T细胞)的比率、Foxp3 mRNA和Foxp3蛋白的表达与急性移植物抗宿主病(aGVHD)的关系.方法 选择37例行HIA相合异基因造血干细胞移植(allo-HSCT))的患者作为研究对象.采用流式细胞术检测患者allo-HSCT前7天(-7 d)、移植当天(0 d)、移植后30天(+30d)、+60 d和+90 d时骨髓或外周血单个核细胞中CD4+CD25+Foxp3+T细胞所占的比率;定量聚合酶链反应(PCR)检测Foxp3 mRNA相对表达量;蛋白印迹(Western blot)法检测Foxp3的蛋白表达水平.结果 37例患者存移植后100 d内发生aGVHD 13例(aGVHD组),未发生aGVHD 24例(无aGVHD组).CD4+CD25+Foxp3+T细胞的比率在0 d时最低,与-7、+30、+60、+90d时比较.差异均有统计学意义(P<0.01);与aGVHD组比较,无aGVHD组患者的CD4+CD25+Foxp3+T细胞的比率明显升高,差异有统计学意义(P<0.01).aGVHD组患者移植后Foxp3的表达水平逐渐升高,但明显低于无aGVHD组,尤其在+60和+90 d时的差异有统计学意义(P<0.01).Western blot法检测结果 表明,aGVHD组受者的Foxp3蛋白表达也明显低于无aGVHD组.结论 CD4+CD25+Foxp3+T细胞对防止发生aGVHD具有重要作用,监测患者骨髓或外周血中CD4+CD25+Foxp3+T细胞的比率可以预测aGVHD的发生.     

Effect of Xiaoyu Zhixue Tablet (消瘀止血片) on Expression of Platelet Membrane Glycoproteins in Patients with Hemorrhagic 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 thrornbopathy 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) Ⅰ b/Ⅸ, GP Ⅱ b/Ⅲ a complexes, GP Ⅰ b, GP Ⅱ b, GPⅢ a 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, GP Ⅱ b/Ⅲ a complexes, GP Ⅰ b, GP Ⅲ a 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 Ⅰ b/Ⅸ, GP Ⅱ b/Ⅲ a complexes, GP Ⅰ b, GPⅢ a and P-selectin at the level of receptor protein.     

川芎嗪对凝血－纤溶系统酶活性的影响

实验以川芎嗪对凝血－纤维蛋白溶解系统酶活性的影响来探讨其活血化瘀机理．结果表明：川芎嗪对凝血过程中的凝血活酶和凝血酶的生成及活性有明显抑制作用，且抑制作用随浓度增加而加强；川芎嗪对纤溶酶原无直接激活作用，但能加速或增加尿激酶对纤溶酶原的激活作用或者增强纤溶酶对纤维蛋白的降解作用．因而川芎嗪对凝血－纤溶系统酶活性的影响是其活血化瘀机制的重要因素．     

PLK1基因缄默在K562细胞凋亡中的作用

目的观察缄默PLK1基因的短发夹状RNA(short hairpin RNA,shRNA)对K562细胞内PLK1表达和细胞凋亡的影响,探讨PLK1在白血病发病中的作用,寻找更为有效的白血病治疗途径。方法设计合成针对PLK1基因1416~1436位点的shRNA片段,并将其克隆于绿色荧光蛋白的表达载体pEGFP-H1中,命名为pEGFP-H1/PLK1。通过电穿孔转染法将其导入K562细胞中,分为对照组、pEGFP-H1空载体转染组和pEGFP-H1/PLK1shRNA重组质粒转染组,转染后24,48h,分别通过RT-PCR和Western blot检测各组细胞PLK1基因和蛋白水平的变化,以MTT方法测各组细胞的增殖活性,比色法检测各组细胞的半胱天冬酶3(caspase-3)蛋白酶活性,并通过流式细胞仪检测各组细胞的凋亡和G2/M期转变情况。结果PLK1mRNA相对水平(与内参的灰度比值)对照组为1.25±0.07,空载体转染24,48h组分别为1.21±0.08和1.23±0.09,shRNA重组质粒转染24,48h组分别为0.52±0.04和0.25±0.02,shRNA重组质粒转染组较其他两组明显降低(P<0.01)。各组细胞PLK1的蛋白水平变化趋势与PLK1mRNA表达相似。相应地,对照组、空载体转染48h组、shRNA重组质粒转染24,48h组的凋亡率分别为(8.3±0.6)%、(8.7±0.7)%、(49.7±3.8)%和(82.3±6.9)%,后两者与前两组之间差异有统计学意义(P<0.05)。此外,与对照组和空载体转染组相比,shRNA重组质粒转染组的细胞增殖能力明显下降(P<0.05),且位于G2/M期的细胞较对照组和空载体转染组显著增加。以上结果均于转染后48h时较明显(P<0.05)。结论构建的shRNA能明显抑制转染细胞PLK1的表达和增殖,并可促进转染细胞的凋亡,并使停留于G2/M期的细胞显著增加。     

发色底物法测定血浆凝血酶活性的研究

近年来的研究表明,血栓栓塞性疾病已成为危害人类健康和生命的主要疾病之一.国内外学者为谋求预测血栓形成的有效实验指标已做了大量的研究.其中最受关注的就是对人体内凝血酶活性的测定。正常人体内仅存在极微量有活性的凝血酶。如果机体受到某种疾病的损害,凝血和抗凝平衡失调,体内凝血酶活性即有所改变。国外多采用单克降抗体测定人体内凝血酶—抗凝血酶复合物。国内采用测定凝血酶的作用产物来间接了解其在体内的活性改变。本文介绍发色底物法直接测定血浆内凝血酶活性的方法。实验原理     

恶性肿瘤患者血栓弹力图检测的临床意义

目的：探讨恶性肿瘤患者体内凝血状态。方法：对65例恶性肿瘤患者于治疗前进行血栓弹力图（TEG）及常现凝血象的检测。结果：恶性肿瘤患者TEG大多数表现为高凝图象：γ值、K值、M值明显缩短，均值分别为2．80±0．81（分）、1．10±0．30（分）、26．40±5．98（分）。Ma值明显增宽，均值为66．42±8．11（mm）。少部分晚期肿瘤患者TEG表现为低凝图象：γ值、K值、M值明显延长，均值分别为8．88±1．51（分）、5．63±1．33（分）、38．89±6．95（分）。Ma值明显变窄，均值为46．08±4．85（mm）。结论：恶性肿瘤患者体内存在凝血功能异常，临床可根据TEG的改变来判断恶性肿瘤患者临床所处凝血状态，正确指导治疗。