人正常胚胎和葡萄胎绒毛中纤蛋白溶酶原激活与抑制因子及其尿激酶受体的表达

本研究应用原位杂交技术检测人正常胚胎及葡萄胎绒毛中纤蛋白溶酶原激活与抑制因子（ｔＰＡ,ｕＰＡ,ＰＡＩ－１）及其尿激酶受体（ｕＰＡ－Ｒ）的表达。结果表明,人正常胚胎绒毛的合体滋养层和细胞滋养层细胞中均含有ｔＰＡ、ｕＰＡ、ＰＡＩ－１及ｕＰＡ－ＲｍＲＮＡ,葡萄胎绒毛中ｔＰＡ、ＰＡＩ－１及ｕＰＡ－ＲｍＲＮＡ表达明显增强。提示：（１）ＰＡ－ＰＡＩ的协同作用在维持妊娠期间正常的纤蛋白溶解可能是重要的;（２）纤溶激活与抑制在葡萄胎的发展过程中可能起作用     

大黄酸对转化生长因子诱导内皮细胞纤溶酶原激活物抑制物1表达的影响

目的 探讨大黄酸对转化生长因子β1（TGF-β1）诱导人脐静脉内皮细胞纤溶酶原激活物抑制物1（PAI-1）mRNA表达和蛋白合成的影响。以阐明大黄酸对内皮细胞的保护作用。方法 以人脐静脉细胞系ECV-304为研究对象，Northern blot检测PAI-1mRNA的表达。流式细胞仪法检测PAI-1蛋白的合成。免疫沉淀法和Western blot检测p44/p42丝裂原活化蛋白激酶（MAPK）活性。结果 大黄酸可以快速抑制TGF-β1诱导的内皮细胞PAI-1mRNA的表达，且呈明显的剂量依赖效应。如果作用时间延长，则对蛋白的合成同样具有明显抑制作用。进一步的研究还发现，大黄酸对TGF-β1激活的p44/p42MAPK活性具有明显下调作用。结论 大黄酸对TGF-β1诱导的PAI-1高表达的抑制作用可能是其保护内皮细胞功能从而防治糖尿病及其血管并发症的作用机制之一。     

血清t-PA和PAI-1水平在腹腔镜手术中的临床意义

目的:探讨腹腔镜与开腹手术中血浆纤维蛋白溶酶的变化。方法:在腹腔镜与开腹手术前、术后8h分别测量血浆中组织型纤溶酶原激活物(Tissue-type p lasm inogen activator,t-PA),纤溶酶原激活物抑制物-1(P lasm inogen activator inh ib itortype-1,PAI-1)浓度。结果:血浆t-PA浓度在腹腔镜手术和传统腹部手术两组病例术后均下降,但在传统腹部手术组下降更显著(P<0.05),而血浆PAI-1浓度在腹腔镜手术和传统腹部手术两组病例术后均升高,但在传统腹部手术组升高更显著(P<0.05)。结论:本次临床研究结果表明:腹腔镜手术组术前、术后血浆中t-PA、PAI-1浓度的变化小于传统腹部手术组,腹腔镜手术组引起腹腔粘连严重程度也低于传统腹部手术组。     

转化生长因子β1对系膜细胞纤溶酶原激活物抑制物1表达的影响

目的观察转化生长因子(TGF)β1对肾小球系膜细胞(GMC)纤溶酶原激活物抑制物(PAI)-1表达的影响,并探讨反应性氧基(ROS)在TGF-β1诱导的PAI-1表达中的作用。方法体外培养大鼠GMC,分别用TGF-β1(2ng/ml)和葡萄糖氧化酶(GO)(10mU/ml)刺激,并用BSO和抗氧化剂N-乙酰半胱氨酸(NAC)进行干预处理。采用Western印迹检测PAI-1蛋白表达;RT-PCR和Northern杂交检测PAI-1mRNA表达;合成的荧光素纤溶酶底物测定纤溶酶活性。结果外源性TGF-β1和GO可显著上调大鼠系膜细胞PAI-1蛋白和mRNA的表达并降低纤溶酶活性。BSO可显著增强TGF-β1和GO诱导的系膜细胞PAI-1mRNA的表达;而NAC可显著地逆转由TGF-β1和GO诱导的PAI-1mRNA表达的上调作用。结论TGF-β1可显著上调系膜细胞PAI-1的表达并抑制纤溶酶活性。ROS在TGF-β1诱导的系膜细胞PAI-1表达上调的信号传递途径中可能起了信号传递分子的作用。     

纤维蛋白对肾小球内皮细胞表达纤溶酶原激活物及纤溶酶原激活物抑制物的作用

目的：观察体外培养人肾小球内皮细胞（GEC）表面原位形成的纤维蛋白对GEC表达纤溶酶原激活物及纤溶酶原激活物抑制物（PA／PAI）的影响。方法：应用逆转录聚合酶链反应（RT－PCR）,酶谱分析法与反向酶谱法分别在基因转录水平与蛋白质活性水平上检测纤维蛋白对GEC表达tPA,uPA gn PAI-1r 作用,纤维蛋白平板法检测纤维蛋白对GEC PA／PAI系统的综合效应,结果：纤维蛋白能够明显促进tPA,uPA与PAI－1的mRNA表达上调,无血清RPMI 1640培养下的GEC几乎检测不到PAI知性,但可检测到PAI－1的活性。纤维蛋白能够浓度依赖性刺激GEC tPA与uPA活性增加以及PAI01的活性增加,呈浓度依赖性与时间依赖性,相同剂量的纤维蛋白原与纤维蛋白的作用相似,放线菌酮与放线菌素D均可抑制纤维蛋白上调GEC表达tPA,uPA与PAI的作用,纤维蛋白平板法显示,纤维蛋白对GEC PA／PAI系统的综合效应是以升高PA活性为主,其活性能够被抑肽酶完全阻断。结论：肾脏局部毛细血[管内沉积的纤维蛋白可能通过对GEC PA／PAI系统的调节发挥其病理作用。     

Postoperative changes in plasma tissue-type plasminogen activator and type I plasminogen activator inhibitor

To clarify the changes which occur postoperatively in intravascular fibrinolysis, plasma levels of tissue-type plasminogen activator (t-PA) antigen, the total plasminogen activator inhibitor type-1 (PAI-1) antigen, and the t-PA-PAI-1 complexes were assayed in this study. Blood samples were taken the morning before surgery, then at 0, 12, 24, 36, 60, 108, and 156h postoperatively in ten patients who underwent radical surgery for thoracic esophageal cancer. The plasma levels of the t-PA and total PAI-1 antigens, and the t-PA-PAI-1 complexes were then measured by enzyme immunoassay. The plasma t-PA and total PAI-1 levels increased significantly in the immediate postoperative period, the percent increase of the latter being much greater than that of the former. Moreover, the calculated free t-PA antigen level was decreased throughout the postoperative period, suggesting postoperative hypofibrinolysis. The platelet count and neutrophil elastase level were significantly correlated with the free t-PA antigen level atr = 0.630,P < 0.001, andr = -0.447,P < 0.01, respectively. The results of this study indicated that postoperative hypofibrinolysis caused by the increased synthesis of PAI-1 may enhance postoperative hypercoagulability, and this may lead to the development of organ damage. Thus, the concentration of the PAI-1 antigen may be a potentially important index for the prediction of postoperative illness.     

纤溶酶原激活及抑制因子在人蜕膜腺体与间质细胞中的生成及调节

为了探讨人早孕蜕膜腺体与间质细胞离体培养下释放活性纤蛋白溶酶原激活及抑制因子（ＰＡ，ＰＡＩ）的平衡关系与激素调节。应用琼脂糖纤蛋白铺盖及反向铺盖技术，测得腺体及间质细胞均仅释放尿激酶型ＰＡ（ＵＰＡ），而不释放组织型ＰＡ（ｔＰＡ），并同时生成Ⅰ型ＰＡ抑制因子（ＰＡＩ－１）。雌二醇（Ｅ２）及ＲＵ４８６刺激ｕＰＡ、抑制ＰＡＩ－１活性，孕酮（Ｐ）的作用则相反，提示Ｐ在人早孕蜕膜中的作用之一是使ＰＡ－ＰＡ１的动态平衡趋于纤溶活性受抑的安静状态，而ＲＵ４８６激活蜕膜纤溶过程，从而增强细胞外基质蛋白水解，可能是其抗早孕作用机理之一。     

尿激酶对糖尿病肾脏病患者血纤溶酶原激活物抑制物1的影响

目的观察尿激酶对糖尿病肾脏病（DKD）患者血纤溶酶原激活物抑制物1（PAI-1）的影响及其临床疗效。方法选择在我院住院的DKD患者88例,其中Ⅲ期43例、Ⅳ期45例。将DKDⅢ、Ⅳ期患者分别分为对照组（DKDⅢ-C组、DKDⅣ—C组）和观察组（DKDⅢ-O组、DKDⅣ—O组）。对照组给予常规降糖、保护肾脏及血管紧张素转化酶抑制剂（ACEI）等药物治疗。观察组在常规治疗的基础上给予尿激酶50000U加入100ml生理盐水中静脉滴注,每天1次,共14d。比较各组24h尿白蛋白量、空腹血糖、血肌酐、D-二聚体和血PAI-1水平。结果DKDⅢ—C组和DKDⅣ—C组治疗前、后24h尿白蛋白量、空腹血糖、血肌酐、D-二聚体和血PAI-1均无统计学差异（P〉0．05）。DKDⅢ-O组和DKDⅣ—O组治疗后24h尿白蛋白量和血PAI-1均降低（P〈0．05）,而空腹血糖、血肌酐、D二聚体治疗前、后均无统计学差异（P〉0．05）。治疗后,DKDⅢ-O组血PAI-1及24h尿白蛋白下降程度较DKDⅣ—O组明显（P〈0．01）。结论尿激酶可通过降低血PAI-1水平来减少DKD患者尿白蛋白量,对保护肾功能、延缓DKD进展有积极意义,且小剂量应用未增加出血倾向,对DKD是一种安全有效的治疗方法。     

Ⅰ型纤溶酶原激活物抑制物在IgA肾病肾小管间质损伤中的作用研究

目的 探讨Ⅰ型纤溶酶原激活物抑制物（ＰＡＩ－１）在ＩｇＡ肾病肾小管间质损害中的作用。方法 采用原位杂交和免疫组织化学技术,分别在基因和蛋白质水平检测３８例ＩｇＡ肾病患者肾小管间质中的ＰＡＩ－１表达,同时观察α－平滑肌肌动蛋白和增殖细胞核抗原（ＰＣＮＡ）的表达变化。     

纤溶酶原激活物抑制物1与肝细胞癌

目的研究纤溶酶原激活物抑制物１（ＰＡＩ１）在肝细胞癌（ＨＣＣ）蛋白和ｍＲＮＡ水平的表达及其与ＨＣＣ生物学特性的关系。方法取ＨＣＣ石蜡标本４８例,肝良性肿瘤石蜡标本１２例（对照组）做免疫组化染色;液氮冻存ＨＣＣ标本２０例,肝血管瘤５例（对照组）做免疫印迹杂交。结果肝癌细胞与癌周细胞及对照组肝细胞相比,ＰＡＩ１抗原蛋白和ｍＲＮＡ表达显著升高,差异有显著意义,Ｐ值分别＜００１和＜０．０５。术后２年内死亡病例与生存病例相比,ＰＡＩ１阳性率有显著意义的升高,Ｐ＜００５。ＰＡＩ１和纤溶酶原激活物（ｕＰＡ）及其受体（ｕＰＡＲ）同时阳性患者与同时阴性患者相比,前者侵袭性病例较后者升高有显著性意义（Ｐ＜００５）。结论ＨＣＣ中ＰＡＩ１蛋白和ｍＲＮＡ表达明显增高。ＰＡＩ１与ＨＣＣ浸润转移和预后密切相关。     

尿激酶型纤溶酶原激活物及其受体高表达增强血管收缩性重塑和内膜增生的研究

目的　探讨血管壁中尿激酶型纤溶酶原激活物 (uPA)和其受体 (uPAR)表达与血管收缩性重塑和内膜增生的相关性。方法　建立兔髂动脉粥样硬化再狭窄模型 ,比较内膜和中膜层的uPA、uPAR表达及其与内膜面积和血管重塑指数的相关性。结果　内膜层uPA、uPAR的抗原和mRNA表达明显强于中膜层 (P <0 .0 1) ,血管壁内膜uPA、uPAR的抗原和mRNA表达水平与血管重塑指数呈负相关 (r =-0 .870 0 7,P =0 .0 10 9;r =-0 .860 3 8,P =0 .0 13 0和r =-0 .845 5 5 ,P =0 .0 165 ,r =-0 .862 40 ,P =0 .0 12 5 ) ,与内膜面积呈正相关 (r=0 .92 0 0 0 ,P =0 .0 3 3 0 ;r=0 .90 772 ,P =0 .0 0 47和r=0 .92 14 9,P =0 .0 0 3 2 ;r =0 .90 614 ,P =0 .0 0 49) ,血管重塑指数和血管内膜面积与中膜uPA、uPAR表达无明显相关性。结论　内膜层uPA、uPAR高表达增强了血管重建后血管壁收缩性重塑和内膜增生。     

ERK1/2信号通路在甲状旁腺激素致人近曲小管上皮细胞分泌纤溶酶原激活物抑制剂1中的作用

目的 从体外观察ERK信号通路在甲状旁腺激素（PTH）致人近曲小管上皮细胞（HK-2）合成纤溶酶原激活物抑制物1（PAI-1）中的作用。 方法 以HK-2细胞株为研究对象,用不同浓度PTH（10-8、10-9、10-10、10-11、10-12 mol/L）作用细胞48 h,以及10-10 mol/L PTH作用细胞不同时间（12、24、36、48、72 h）,分别用RT-PCR法检测PAI-1 mRNA表达,Western印迹法检测PAI-1蛋白表达。以10-10 mol/L PTH作用细胞48 h,分别观察细胞ERK1/2抑制剂预处理前后磷酸化（p）ERK1/2、PAI-1mRNA及蛋白的变化情况。 结果 10-12 mol/L PTH可促进细胞在基因及蛋白水平合成PAI-1,随着PTH浓度逐渐上升,PAI-1mRNA及蛋白浓度均相应增加,以10-10 mol/L PTH组刺激作用最显著,分别为对照组的4.01倍和3.81倍（均P < 0.01）。但随着PTH浓度进一步增加,PAI-1mRNA及蛋白浓度却随之下降。10-10 mol/L PTH作用细胞,12 h时有少量PAI-1表达,72 h时达峰值,并呈时间依赖性,分别为0 h组的4.06倍和4.03倍（均P < 0.01）。10-10 mol/L PTH作用细胞48 h,有大量的p-ERK1/2合成（P < 0.01）,经ERK1/2抑制剂预处理后,PAI-1及ERK均显著下降（均P < 0.01）,但仍高于对照组（均P < 0.05）。 结论 ERK信号通路部分参与PTH致HK-2细胞合成PAI-1的作用。     

白介素1β对肾小球系膜细胞表达纤维蛋白溶解酶原激活物抑制物-1和纤维连接蛋白的影响

目的探讨白介素１β（ＩＬ１β）对人肾小球系膜细胞表达纤维蛋白溶解酶原（纤溶酶原）激活物抑制物１（ＰＡＩ１）和纤维连接蛋白（ＦＮ）的影响。方法应用Ｎｏｒｔｈｅｒｎ杂交检测体外培养的肾小球系膜细胞在ＩＬ１β刺激后ＰＡＩ１和ＦＮ的ｍＲＮＡ表达,采用纤维蛋白平板法检测系膜细胞培养上清中ＰＡＩ１的活性,利用ＥＬＩＳＡ法检测ＦＮ的水平。结果ＩＬ１β刺激组系膜细胞ＰＡＩ１和ＦＮｍＲＮＡ的表达显著增高（与对照组比,Ｐ＜００５）,细胞培养上清中ＰＡＩ１的活性和ＦＮ的水平亦明显增加（与对照组比,Ｐ＜００１）。结论ＩＬ１β可以上调系膜细胞ＰＡＩ１和ＦＮ蛋白质及ｍＲＮＡ的表达,提示ＩＬ１β可能通过抑制细胞外基质降解和增加细胞外基质合成而导致细胞外基质的积聚。     

Thrombospondin-1 and transforming growth factor beta-1 upregulate plasminogen activator inhibitor type 1 in pancreatic cancer

Controlled degradation of the extracellular matrix by proteases is crucial in tumor cell invasion. We have shown that thrombospondin-1 (TSP-1), through activation of transforming growth factor beta-1 (TGF-β1), regulates the plasminogen/plasmin protease system in breast cancer. To determine whether this occurred in other epithelial neoplasms, we studied the role of TSP-1 and TGF-β1 in the regulation of the plasminogen/plasmin system in pancreatic cancer. ASPC-1 and COLO-3S7 pancreatic cancer cells were treated with TSP-1 or TGF-β1 at varying concentrations. The TSP-1 and TGF-β1-treated cells were also treated with either anti-TSP-1, anti-TSP-1 receptor, or anti-TGF-β1 antibodies. Urokinase plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) expression was determined by enzyme-linked immunosorbent assay. TSP-1 and TGF-β1 promoted a dose-dependent upregulation of ASPC-1 and COLO-3S7 PAI-1 expression. The TSP-1 effect could be blocked with anti-TSP-1 or anti-TGF-β1 antibodies. The TGF-β1 effect could be blocked only with anti-TGF-β1 antibody. Anti-TSP-1 receptor antibody blocked the TSP-1 effect on PAI-1 expression but had no effect on TGF-β1-mediated PAI-1 expression. Neither TSP-1 nor TGF-β1 had an effect on uPA production. We conclude that TSP-1, in a receptor-mediated process that involves the activation of TGF-β1, upregulates PAI-1 expression in pancreatic cancer without an effect on uPA production. Supported in part by National Institutes of Health grants CA65675 and CA69722 (Dr. Tuszysnki). Dr. Berger is the recipient of an American Cancer Society Clinical Career Development Award 96-09. Presented at the Thirty-Eighth Annual Meeting of The Society for Surgery of the Alimentary Tract, Washington, D.C., May 11–14,1997.     

Elevated plasminogen activator inhibitor levels in cyclosporin-treated renal allograft recipients

Atherosclerosis and thrombosis, two major causes of morbidityand mortality in renal transplant recipients, share the sameclinical risk factors including decreased fibrinolysis and lipiddisturbances. In a crosssectional study we have determined parametersof fibrinolysis in control subjects (n=23) and stable renalallograft recipients without cyclosporin CsA (n=10) and withCsA (n=87) in their immunosuppressive treatment. In CsA-treatedpatients, tissue-type plasminogen activator was moderately increasedcompared to patients without CsA (8.4±3.3 vs 5.5±2.8ng/ml). The plasminogen activator inhibitor (PAI) activity inplasma was clearly increased in CsA-treated patients: 14.5±8.8vs 7.2±3.2 in normal controls and 8.5±2.4 AU/mlin patients without CsA. Total cholesterol and LDL cholesterollevels were higher in CsA-treated patients (256±62 and169±60 mg/dl) than in patients without CsA (209±45and 136±44 mg/dl). The two groups did not differ in HDLcholesterol, triglycerides, and lipoprotein(a). Hypercholesterolaemia,obesity, and steroid-induced diabetes could be identified asrisk factors for elevated plasma PAI activity in CsA-treatedpatients. Hypofibrinolysis induced by elevated PAI levels andincreased LDL cholesterol may contribute to the increased thrombogenicityand accelerated atherosclerosis observed in cyclosporin-treatedpatients.     

结缔组织生长因子反义寡核苷酸对肾小管上皮细胞纤溶酶原激活物抑制物1表达的影响

目的探讨结缔组织生长因子(CTGF)反义寡核苷酸(ODN)对转化生长因子β1(TGF-β1)诱导的肾小管上皮细胞纤溶酶原激活物抑制物1(PAI-1)mRNA表达和蛋白产生的影响,以明确CTGF在肾脏细胞外基质(ECM)降解中的作用。方法体外培养人近曲小管上皮细胞(HKC),以脂质体介导方法将CTGF反义ODN转染细胞。以TGF-β1(5μg/L)刺激HKC不同时间后,用逆转录-聚合酶链式反应(RT-PCR)方法检测PAI-1mRNA表达;流式细胞仪法检测胞内PAI-1蛋白合成;Western印迹方法检测HKC分泌到上清中的PAI-1蛋白含量。结果TGF-β1可诱导HKC高表达CTGF和PAI-1。转染后6h,CTGF反义ODN可显著抑制TGF-β1诱导的HKCPAI-1mRNA表达。转染后24h,CTGF反义ODN可明显抑制胞内PAI-1蛋白合成,并减少了HKC分泌到胞外的PAI-1。结论CTGF在肾小管间质纤维化时ECM降解中起了关键调控作用,其反义ODN的导入可能是延缓肾小管间质纤维化的有效手段之一。     

Expression of cytokines and growth factors in human glomerulonephritides

Numerous experimental studies point to the potential role of cytokines and growth factors in the pathogenesis of renal disease. However, from the various autocrine and paracrine mediators identified in vitro and in animal models, so far only a few have been demonstrated in selected human glomerulopathies. We examined two types of glomerulonephritis (GN): extracapillary GN with anti-neutrophil cytoplasmic autoantibodies (ANCA), an example of an acute form of GN, and mesangial IgA GN, usually a chronic form of GN, with immunocytochemistry, in situ hybridization and the polymerase chain reaction. Normal renal tissue from tumour nephrectomies served as a control. In ANCA-positive GN with active renal lesions (crescents, glomerular and vascular necrosis), infiltrating mononuclear cells in glomeruli and in the interstitium expressed interleukin (IL)-1, tumour necrosis factor (TNF)-, IL-2, interferon (IFN)-, platelet-derived growth factor (PDGF) and transforming growth factor (TGF)-. Cytokine expression was also observed in activated resident cells, including endothelial cells, capsular epithelial cells, smooth muscle cells of vessel walls, fibroblasts and some tubular epithelial cells. In addition, we noted an increase in the cytokine and growth factor receptors TNF-R, IL-1R type II, IL-2R, IFN-R and PDGF-R. In contrast, in mesangial IgA-GN, IL-1, TNF-, IFN- and IL-2 were usually absent in glomeruli. Mesangial expansion in this disorder was accompanied by an increased expression of PDGF, PDGF-R, TGF- and IL-6 in mesangial areas. In both conditions a good correlation was observed between cytokine expression at the mRNA (in situ hybridization) and protein level (immunocytochemistry). These results demonstrate that different cytokine and growth factor patterns are expressed in the various forms of GN, and suggest that the local production of these peptides plays an important role in the pathogenesis and progression of human glomerulonephritides.