尿激酶型纤溶酶原激活物对糖尿病大鼠肾脏系膜基质的影响

目的 探讨尿激酶型纤溶酶原激活物（uPA）对糖尿病大鼠肾脏系膜基质表达的影响及机制.方法 8周龄健康雄性SD大鼠（体质量150～200 g）20只尾静脉注射链脲佐菌素,成功建立糖尿病大鼠模型后采用数字表法随机分为糖尿病组（n=10）和uPA组（n=10;尾静脉注射2500 U·kg^-1·d^-1 uPA,共4周）.另以10只正常大鼠为对照组.29d后处死大鼠,心脏取血检测血糖、血肌酐水平.过碘酸六胺银染色测定肾小球平均面积、肾小球平均容积和肾小球系膜区面积.免疫组织化学法检测肾脏尿激酶型纤溶酶原激活物受体（uPAR）、纤溶酶原激活物抑制剂-1（PAI-1）和Ⅳ型胶原表达水平.采用方差分析和q检验进行数据统计.结果 与正常对照组比较,糖尿病组大鼠明显出现尿蛋白[（25.4±4.3）mg/24 h vs（5.5±2.1）mg/24 h,q=4.27,P＜0.01],肾小球体积及系膜基质显著增加,肾小球系膜uPAR、PAI-1、Ⅳ型胶原表达显著增加（q值分别为3.63、3.97、4.21,均P＜0.05）.糖尿病大鼠注射uPA后,尿蛋白明显减少[（12.6±5.4）mg/24 h,q=3.45,P＜0.05],肾小球体积、系膜基质异常有所改善（q值分别为4.34、4.27,均P＜0.01）,肾小球系膜PAI-1、Ⅳ型胶原表达明显减少（q值分别为3.98、4.17,均P＜0.05）.结论注射uPA可明显降低糖尿病大鼠肾小球PAI-1蛋白表达,对uPAR表达的影响不大,提示uPA可能通过与uPAR结合、摄取PAI-1并加速其降解,从而调节肾小球系膜细胞及其基质表达,改善糖尿病系膜基质病变.     

下调PAI对HpG2细胞组织型纤溶酶原激活物和尿激酶纤溶酶原激活物的影响

目的探讨纤溶酶原激活物抑制剂(PAI)的发卡样小干扰RNA(siRNA)真核表达载体对人肝癌HpG2细胞组织型纤溶酶原激活物(tPA)和尿激酶PA(uPA)表达的影响。方法设计短发夹状PAI siRNA的寡核苷酸链,将其插入到真核细胞表达载体pSilencerTM 2.1-U6 neo载体中构建重组载体后,采用脂质体转染方法将构建的重组载体导入人肝癌细胞HpG2中,采用逆转录聚合酶链反应(RT-PCR)、Western印迹法分别检测转染细胞PAI mRNA和蛋白对tPA和uPA表达的影响。结果转染后细胞中PAI含量降低,tPA和uPA的表达减少(均P<0.01)。结论测序结果表明发卡样的PAI siRNA真核表达载体转染HpG2细胞后获得稳定表达,并可特异性封闭PAI的表达,PAI siRNA能有效阻断PAI的蛋白合成,同时抑制细胞中tPA和uPA的表达。     

血管紧张素转换酶抑制剂对糖尿病大鼠血浆PAI-1和tPA活性的影响

目的 探讨血管紧张素转换酶抑制剂(ACEI)对糖尿病(DM)大鼠血浆纤溶酶原激活物抑制剂1(PAI-1)和组织型纤溶酶原激活物(tPA)活性的影响及其机制.方法 将链脲佐菌素诱导的DM大鼠分为正常对照组、培哚普利治疗组、培哚普利联合一氧化氮合酶(NOS)抑制剂治疗组.治疗4周后比较各组血浆PAI-1和tPA活性及PAI-1/tPA比值.结果 与正常对照组相比,DM大鼠血浆PAI-1活性和PAI-1/tPA比值显著升高,tPA活性降低.培哚普利治疗使PAI-1活性下降,tPA活性增加.联合NOS抑制剂在一定程度上抵消了培哚普利的这种作用.结论 DM状态下存在纤溶异常,ACEI能够通过内源性的NO改善纤溶平衡.     

糖基化终末产物对大鼠肾皮质PAI-1表达和活性的影响

目的观察糖基化终末产物（AGEs）对大鼠肾皮质纤溶酶原激活物抑制物-1（PAI-1）表达和活性的影响。方法大鼠尾静脉注射AGE修饰大鼠血清蛋白（AGEs）6周，其中部分大鼠同时腹腔注射AGE形成抑制剂氨基胍（AG），以注射天然大鼠血清蛋白（RSP）和正常大鼠（Con）作为对照。免疫组织化学、Western blot、RT—PCR检测PAI-1表达水平，酶谱法分析纤溶酶原激活物（PA）活性，PAS染色评估细胞外基质（ECM）含量。结果与Con组及RSP组比较，AGEs组大鼠肾皮质ECM含量、PAI-1蛋白及mRNA表达水平均明显增高，PA活性明显降低（PAI-1 mRNAP〈0．01，其余P〈20．05），而同时注射AG的大鼠上述变化明显减轻。结论AGEs上调大鼠肾皮质PAI-1的表达，抑制PA活性，可能是糖尿病肾病ECM积聚的原因之一。     

化瘀通络中药对5/6肾切除大鼠纤溶系统的影响

目的 观察化瘀通络中药肾络通对5/6肾切除大鼠纤溶系统的影响.方法 46只健康雄性SD大鼠随机分为假手术组、模型组、苯那普利组、肾络通组.复制5/6肾切除肾衰大鼠模型,采用免疫组织化学及原位杂交方法检测各组大鼠肾组织纤溶酶原激活物组织型(t-PA)、纤溶酶原激活物尿激酶型(u-PA)、纤溶酶原激活物的抑制物-1(PAI-1)及PAI-1 mRNA的表达,并进行病理学检测.结果 模型组t-PA、u-PA的相对含量及ILD值均显著低于假手术组(P＜0.01),而PAI-1及PAI-1 mRNA则显著高于假手术组(P＜0.01);肾络通组、苯那普利组与模型组相比,t-PA、u-PA的相对含量及ILD值均明显升高(P＜0.01或P＜0.05),而PAI-1及PAI-1 mRNA则明显降低(P＜0.01或P＜0.05).结论 化瘀通络中药肾络通能纠正5/6肾切除大鼠纤溶系统的紊乱,延缓肾脏疾病的进程.     

TGF-β1和PAI-1在老年IgA肾病患者中的表达及意义

目的探讨血液及尿液中转化生长因子(TGF)-β1和纤溶酶原激活物抑制物(PAI)-1在老年IgA肾病患者中的水平及意义。方法免疫组织化学和ELISA方法检测研究对象肾脏组织、血液及尿液中TGF-β1和PAI-1水平。结果与老年人和青少年非IgA肾病患者相比,老年IgA肾病患者肾脏组织、血液和尿液中TGF-β1和PAI-1表达水平显著升高(P0.05)。随着病情的加重,老年IgA肾病患者肾脏组织、血液及尿液中TGF-β1和PAI-1表达水平显著升高(P0.05)。结论老年IgA肾病患者肾脏组织、血液和尿液中TGF-β1和PAI-1表达水平与疾病严重程度相关,可作为老年IgA肾病辅助诊断及判断疾病进展的手段之一。     

非诺贝特对2型糖尿病大鼠肾皮质纤溶酶原激活物抑制物1表达的影响

小剂量链脲佐菌素加高脂饮食诱导的糖尿病大鼠给予非诺贝特灌胃8周，检测显示糖尿病大鼠肾皮质纤溶酶原激活物抑制物1（PAI-1）mRNA表达明显下降，同时肾小球肥大部分改善，提示非诺贝特对糖尿病肾脏的保护作用可能与其下调肾皮质PAI-1基因表达有关。     

纤溶酶原激活物抑制剂-1与肾间质纤维化

肾间质纤维化(RIF)是各种慢性肾脏疾病发展到终末期肾病的共同归路.纤溶酶原激活物抑制剂(PAI)的表达及活性异常在RIF病变中发挥着至关重要的作用.该文就PAIs系统在RIF发生发展过程中的作用做一综述.     

马来酸罗格列酮对大鼠血浆tPA和PAI-1活性的影响

STZ诱导的糖尿病大鼠研究显示，罗格列酮治疗（每日4mg／kg，3周）能增加组织型纤溶酶原激活物（tPA）的血浆水平，降低PAI—1的活性，降低PAI-1／tPA的比值。因此，罗格列酮可能有心血管保护作用。     

同型半胱氨酸和叶酸对内皮细胞纤溶系统影响

同型半胱氨酸（Hcy）水平的升高与动脉粥样硬化及心脑血管疾病密切相关，叶酸补充治疗可以减少Hcy水平，然而高胱氨酸血症（HHE）的致病机制及叶酸的保护机制还不完全清楚。为了探讨Hcy及叶酸对纤溶系统的影响，我们首次研究了生理及超生理浓度的Hcy与叶酸共同作用对人脐静脉血管内皮细胞（HUVEC）的组织型纤溶酶原激活物（tPA）和纤溶酶原激活物抑制剂-1（PAI-1）的抗原合成及mRNA表达的影响。     

组织型纤溶酶原激活剂及其抑制剂与肺血栓栓塞症

肺血栓栓塞症（PTE）的发病与机体的纤溶和凝血系统功能密切相关。组织型纤溶酶原激活物（t-PA）及其抑制物（PAI-1）因调节机体的纤溶系统而在静脉血栓形成及栓塞性疾病的发病机制中发挥重要作用,因此,本文对t—PA和PAI-1与PTE的关系作如下综述。     

Competition between plasminogen and tissue plasminogen activator for cellular binding sites

Cellular receptors for plasminogen and tissue plasminogen activator (t- PA) regulate plasminogen activation and cell-associated proteolytic activity. The characteristics of the interactions of both ligands with monocytes and monocytoid cell lines bear certain similarities, including affinity (kd approximately 1 mumol/L) capacity and susceptibility to carboxypeptidase treatment. Therefore, we have undertaken the present study to determine directly whether t-PA and plasminogen share common binding sites on cells. We found that recombinant human single-chain t-PA (rt-PA) could inhibit the binding of 125I-plasminogen to the cells and, conversely, plasminogen could inhibit 125I-rt-PA binding. This relationship was observed with 9 cell types, including both adherent cells and cells in suspension. In addition, under several conditions of cell treatment, plasminogen and t- PA receptor expression was modulated in parallel. Furthermore, molecules that have been implicated as candidate plasminogen receptors, gangliosides, and an alpha-enolase--related molecule, also interacted with t-PA. These results suggest that at least a component of the binding sites for plasminogen is shared with t-PA. Occupancy of these sites by either or both ligand(s) should result in arming the cells with the proteolytic activity of plasmin.     

Thyroid follicular cells secrete plasminogen activators and can form angiostatin from plasminogen

Angiostatin, a 38 kDa fragment of plasminogen, potently inhibits the growth of blood vessels. Angiostatin is generated from plasminogen by urokinase-type (uPA) and tissue-type (tPA) plasminogen activators in the presence of free sulphydryl donors. Angiogenesis inhibitors may be important in regulating angiogenesis in developing goitre. We have examined angiostatin formation in human primary thyrocyte cultures and a rat thyrocyte cell line (FRTL-5). We found that human thyroid cells in culture secrete plasminogen activators (both tPA and uPA) as well as matrix metalloproteinase 2 into the medium. When human thyrocyte conditioned medium was incubated with plasminogen (10 microg/ml) and N-acetylcysteine (100 microM) for 24 h, a 38 kDa fragment of plasminogen, which is consistent with angiostatin, was generated. The appearance of the 38 kDa fragment was increased by agents that increase cAMP (forskolin and 8 BrcAMP). FRTL-5 cells, which do not secrete uPA or tPA, did not generate angiostatin. Thyroid cells produce several angiogenic growth factors, and human thyrocyte conditioned medium stimulated growth of endothelial cells. When the conditioned medium was incubated with plasminogen and N-acetylcysteine, this stimulatory effect was lost, consistent with the production of a growth inhibitory factor. We conclude that thyroid cells can produce angiostatin from plasminogen in vitro, and this may play a role in vivo in limiting goitre size.     

Activity of tissue plasminogen activator and plasminogen activator inhibitor in noninsulin-dependent diabetes mellitus

The activity of free tissue plasminogen activator (f-tPA) and plasminogen activator inhibitor (PAI) in the plasma of 82 noninsulin-dependent diabetics (NIDDM) was measured by bioimmunoassay of the euglobulin fraction obtained from the plasma, and the levels were compared with those of age- and gender-matched normal subjects. Comparison of these levels in both groups revealed that the f-tPA activity tended to be lower in NIDDM than in the controls, although the differences were not significant. Normal activity of PAI was seen, but f-tPA in NIDDM, when accompanied by macroangiopathy such as ischemic heart disease, was significantly depressed. When glycosylated hemoglobin levels were in excess of 10%, the f-tPA activity was significantly decreased, but no reduction was found in PAI activity as compared with controls. When NIDDM is associated with either macroangiopathy or high glycosylated hemoglobin levels, a decreased f-tPA activity, rather than an increased PAI activity, may contribute to the development of a defective fibrinolytic state.     

Tissue plasminogen activator release and plasminogen activator inhibitor levels in coronary artery disease

Unstable angina and Q wave myocardial infarction are associated with intraluminal coronary thrombosis, a process to which impaired fibrinolysis may contribute. The authors examined the extrinsic fibrinolytic system, including tissue plasminogen activator antigen, plasminogen activator inhibitor activity and antigen, and euglobulin clot lysis time before and after venous occlusion in 56 patients undergoing coronary angiography for chest pain syndromes and in 16 healthy controls. Fibrinolysis variables were similar (with greater than 95% confidence) in the patients with thrombus-associated coronary syndromes as compared with those with chest pain syndromes not due to coronary thrombosis. These fibrinolytic variables were also similar to those in patients without coronary artery disease and in healthy controls. Their data suggest that defective fibrinolysis is not involved, at least systemically, in the pathogenesis of thrombus-associated coronary artery syndromes.     

Kinetic studies of plasminogen activation by epithelial tissue plasminogen activator

Initial-rate kinetic studies of the activation of plasminogen by epithelial activator were performed in the absence and in the presence of fibrinogen and CNBr-digested fibrinogen, under assay conditions similar to those described by Hoylaerts et al. (J Biol Chem, 257, 2912, 1982). In the purified system, and in the absence of any stimulator, Lys-plasminogen is more readily activated than Glu-plasminogen, with catalytic rate constants of 0.01 s-1 and 0.0034 s-1 and Michaelis constants of 1.2 microM respectively. With Glu-plasminogen, double reciprocal plots deviated from linearity at low concentrations of plasminogen, in agreement with the findings reported for melanoma activator. In the presence of fibrinogen, activation rates for both Lys and Glu-plasminogen were increased. (kcat = 0.017 and 0.041 s-1 and km = 1.4 and 41 microM, respectively). In the presence of CNBr-fragments of fibrinogen, the Michaelis constant is lowered for both forms of plasminogen, (km = 0.3 microM) thus indicating high affinity and efficient activation of plasminogen on fibrin clot. Comparison of the kinetic data with those reported for melanoma activator suggest that even though the values of the kinetic constants are different, epithelial activator has a similar mechanism of action for the activation of plasminogen as the melanoma enzyme.