蛋白酶激活受体1与胃癌转移关系的研究进展

蛋白酶激活受体1(protease activated receptor 1,PAR1)是介导血栓形成的一种G蛋白偶联受体,属于蛋白酶激活受体(protease activated receptors,PARs)家族成员,其主要特征是作为凝血酶的受体参与血栓形成及凝血活动。PAR1表达于人多种正常细胞,在血栓形成、炎症反应、血管形成及免疫功能等过程中发挥重要作用。据最近研究发现,PAR1在多种恶性肿瘤中高表达,提示高表达的PAR1可能介导某些肿瘤的恶性生物学行为,并且其表达水平的改变与胃癌的侵袭转移有关,有望成为胃癌新的诊断标志和干预靶点。     

脑血康对脑出血蛋白酶激活受体-1表达的影响

目的:探讨脑出血后蛋白酶激活受体-1(PAR-1)的动态表达及脑血康的干预作用.方法:72只大鼠随机分为正常组、脑出血模型6 h、24 h、3 d、7 d组和脑血康治疗6 h、24 h、3 d、7 d组.用Ⅶ-S型胶原酶诱导大鼠脑出血模型.免疫组化方法测定各时间点大鼠脑出血后血肿周围水肿组织PAR-1蛋白表达;逆转录聚合酶链反应(RT-PCR)检测PAR-1 mRNA表达.结果:正常组大鼠大脑PAR-1蛋白和PAR-1 mRNA表达轻度阳性;模型组6 h时PAR-1蛋白和PAR-1 mRNA表达强度开始增强,24 h表达进一步增加,于3 d达到高峰,然后开始下降,7 d时明显下降.在6 h、24 h、3 d、7 d各时间点,模型组和脑血康组PAR-1阳性细胞数、PAR-1 mRNA吸光度(A)比值升高,与正常组比较差异均有显著性(P<0.05或P<0.01);脑血康组PAR1阳性细胞数、PAR-1 mRNA A比值降低,与模型组各时间点比较差异均有显著性(P<0.05或P<0.01).结论:脑出血后PAR-1受到凝血酶的持续活化,脑出血后凝血酶的作用可能通过PAR-1介导;脑血康可抑制PAR-1活化,这可能是脑血康治疗脑出血的主要机制之一.     

海水对肺腺癌细胞株A549细胞蛋白酶激活受体2表达的作用研究

目的 观察海水对肺腺癌细胞株A549细胞的炎性损伤及对蛋白酶激活受体2(PAR-2)表达的影响,探讨海水导致急性肺损伤的作用机制.方法 将培养的A549细胞接种于6孔板中,按随机数字表法把细胞分为对照组及海水处理2、4、8 h组,相应时间点用实时荧光定量逆转录-聚合酶链反应(RT-PCR)和蛋白质免疫印迹法(Western blotting)检测PAR-2的mRNA和蛋白表达;收集细胞培养上清液,用酶联免疫吸附法(ELISA)检测肿瘤坏死因子-α(TNF-α)和白细胞介素-8(IL-8)含量.结果 用海水处理后,A549细胞PAR-2 mRNA和蛋白表达均于2 h显著增加(P均<0.05),4 h达高峰,分别为对照组的1.8倍和2.2倍,随后下降,但仍显著高于对照组(P均<0.01);TNF-α和IL-8含量则于2 h即显著升高并达最高峰[分别为(214.35±20.85)ng/L,(55.86±5.65)ng/L],随后下降,但仍显著高于对照组[分别为(25.86±3.85)ng/L,(6.97±1.77)ng/L,P均<0.01].结论 海水可致A549细胞炎症反应明显,并可诱导A549细胞PAR-2表达增高.     

蛋白酶激活受体在过敏反应中的作用及检测方法

蛋白酶激活受体（protease activated receptors，或proteinase activated receptors，PARs）是1991年后陆续发现的G蛋白偶联受体（GPCRs）家族中的4个新成员。PARs的表达几乎遍及所有涉及过敏反应的细胞，现将PARs在过敏反应中的作用特点等综述如下。     

蛋白酶激活受体-1在结肠癌组织中的表达及其临床意义

目的：观察并探讨蛋白酶激活受体-1（proteinase activated receptor-1,PAR-1）在结肠癌组织中表达的临床意义。方法：用免疫组织化学方法检测65例结肠癌及其癌旁肠组织中PAR-1的表达情况。结果：PAR-1的表达在结肠癌组织中高于癌旁肠组织,随肌壁癌组织浸润度和淋巴结转移程度的提升而增高,均有显著差异（P〈0.01）;PAR-1的表达虽随着癌分化程度降低而增高,但无显著差异（P〉0.05）。结论：PAR-1的表达与结肠癌发展及临床病理分期有关,可作为结肠癌临床预后指标之一。     

反复自然流产患者外周血中性粒细胞蛋白酶激活受体2和组织因子的表达

摘要:目的：检测蛋白酶激活受体2（proteaseactivated receptor 2, PAR2）和组织因子（tissue factor,TF）在反复自然流产（recurrent spontaneous abortion,RSA）患者外周血中性粒细胞的表达。 方法：用real-time PCR、western blot及TF活性分析方法,在mRNA、蛋白质及活性水平检测35例RSA患者和35例健康非孕或健康孕妇外周血中性粒细胞PAR2和TF的表达。 结果：与健康非孕或健康孕妇比较,RSA患者外周血中性粒细胞PAR2和TF在mRNA、蛋白质及活性水平均显著升高,且PAR2和TF mRNA水平呈明显正相关（r=0.864,P＜0.05）,PAR2蛋白质水平与TF活性水平间有正相关关系（r=0.942,P＜0.05）。 结论：RSA患者外周血中性粒细胞高表达PAR2和TF,为进一步研究PAR2和TF在RSA中的作用提供了依据。     

蛋白酶激活受体在ALI/ARDS炎症反应中的作用研究进展

急性肺损伤(ALI)/急性呼吸窘迫综合征(ARDS)目前仍是困扰医学界的复杂难题,其发病机制尚未完全阐明.蛋白酶激活受体(PARs)是新近发现的一种G蛋白偶联受体亚类,以其独特的激活方式通过细胞内复杂的信号转导,对ALI/ARDS的炎症过程发挥了重要的调节作用.本文简要叙述了PARs的结构、激活和灭活方式、细胞内信号转导及其效应,着重介绍了在ALI/ARDS中PARs四种亚型介导的炎症反应,以及炎性介质对PARs的调控,从而阐明PARs激活在推动肺部炎症病理过程发展中的作用.     

蛋白酶激活受体-1在动脉粥样硬化中的作用及其机制研究

目的 分析蛋白酶激活受体-1(PAR-1)在动脉粥样硬化中的作用及其机制研究。方法 选取40只ApoE⁽(-/-))小鼠,将40只小鼠随机分为对照组、模型组、PAR-1模拟物组、PAR-1抑制剂组,每组10只。比较各组的PAR-1相对表达量、小鼠斑块泡沫细胞占比、斑块脂质核心面积百分比、丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、胆固醇(TC)、甘油三酯(TG)水平、MAPKs、NF-κB信号通路蛋白。结果 PAR-1抑制剂组的PAR-1相对表达量低于PAR-1模拟物组、模型组,高于对照组,差异均有统计学意义(t分别=10.63、8.49、-8.49,P均<0.05);PAR-1抑制剂组的斑块泡沫细胞占比、斑块脂质核心面积百分比低于PAR-1模拟物组、模型组,差异均有统计学意义(t分别=56.21、42.76;20.65、21.72,P均<0.05);PAR-1抑制剂组的ALT、AST、TC、TG低于PAR-1模拟物组、模型组,高于对照组,差异均有统计学意义(t分别=16.84、5.18、-0.99;11.66、5.24、-1.73;31.30、...     

蛋白酶激活受体（PARs）在心血管系统中的作用

蛋白酶激活受体（PARs）属于G蛋白耦联受体超家族，在感染、创伤愈合、血管舒张、血管再生、动脉粥样硬化、胃肠道疾病、神经系统疾病、肺部炎症等病理生理机制中发挥了重要的作用^[1]。研究表明PARs在心血管系统广泛分布，它们促使血管舒张、影响心脏的电活动和机械活动、参与血管和心肌层的结构重构等，[第一段]     

人类GM-CSF受体胞外结构域及功能的研究进展

人类粒-巨噬细胞集落刺激因子(granulocyte-macrophage colony-stimulating factor,GM-CSF)通过细胞膜表面特异性受体(GM-CSF receptor,GMR)介导维持细胞存活,刺激细胞增殖与分化,调节成熟细胞功能。GMR为跨膜受体由α链和β链共同组成,结构上分为胞外域、跨膜部分和胞内域。α链和β链胞外域均含有一种造血生长因子受体超家族的共同结构模体(cytokine receptor modules,CRM)并依靠保守的半胱氨酸残基形成的二硫键维持胞外空间结构。GM-CSF和α链通过电荷识别结合并诱导β链加入,形成完整的配体受体复合物。在二硫键的作用下,GMR多聚化使两条以上β链胞内部分相互接近活化JAK2,完成GM-CSF对细胞的激活过程。     

阿霉素诱导Egr-1启动子调控的造血因子GM-CSF基因表达对荷瘤小鼠造血功能恢复的影响

本研究探讨阿霉素（ADM）诱导早期生长因子（Egr-1）启动子调控的造血因子基因表达对荷瘤小鼠化疗后造血功能恢复的作用。将构建的携带有Egr-1启动子的粒-巨噬细胞集落刺激因子（GM-CSF）cDNA和增强型绿色荧光蛋白（EGFP）cDNA双顺反子真核表达载体导入基质细胞HFCL后,输入重症联合免疫缺陷（SCID）小鼠体内。实验小鼠随机分4组：①ADM诱导的HFCL／EG组（HFCL／EG＋ADM组）,②ADM诱导的HFCL组（HFCL＋ADM组）,③单纯输注HFCL／EG组（HFCL／EG组）和④单纯输注HFCL组（HFCL组）,每组6只动物。观察外周血象动态改变,用流式细胞术检测eGFP^＋人基质细胞,用RT-PCR和Westernblot分别检测GM-CSFmRNA及其蛋白的表达。结果表明：化疗组与未化疗组相比外周血白细胞降低,而且下降幅度较低,恢复加快;各组间CFU-GM数无显著性差异;肿瘤抑制率与化疗相关,而与外源基因表达不相关;ADM处理后实验组小鼠骨髓可见绿色荧光阳性的基质细胞;RT,PCR和Westernblot显示GM-CSFmRNA和GM-CSF蛋白表达增强。结论：ADM诱导的Egr-1启动子造血因子基因疗法具有化疗后促进造血恢复作用。     

抗原致敏、GM-CSF基因修饰的树突状细胞诱导免疫杀伤多发性骨髓瘤细胞U266的研究

本研究探讨负载有多发性骨髓瘤细胞U266可溶性抗原并携带外源基因粒-巨噬细胞集落刺激因子（GM—CSF）的树突状细胞（DC）,在体外激活自身T淋巴细胞形成细胞毒T淋巴细胞（CTLs）对U266细胞的杀伤作用。用U266可溶性抗原致敏DC,再用含有外源基因GM—CSF的腺病毒感染DC,将所得的DC与T细胞混合培养以形成对U266具有特异杀伤作用的CTL,最后通过测定乳酸脱氢酶（LDH）以计算CTL对U266的杀伤率。结果表明：负载抗原及外源基因组、负载抗原组和对照组之间的杀伤率存在显著差异（n=3,F=10．939,P〈0．05）;两两比较表明,负载抗原及外源基因组的杀伤率高于其它两组（P〈0．001）,且负载抗原组高于对照组（P〈0．001）。结论：以U266可溶性抗原致敏的DC可诱导出对U266具有特异杀伤作用的CTL,当所致敏的DC通过腺病毒感染而带有外源基因GM—CSF时,所诱导的细胞毒杀伤反应则进一步增强。     

rhTpo／GM-CSF融合蛋白的构建、表达及活性测定

本研究的目的是找出一个治疗由于放化疗等原因造成的造血组织损伤导致的贫血、感染和出血的有效方法。通过RT-PCR的方法从人胎肝中克隆了重组人血小板生成素(rhTpo)的基因,利用基因工程的手段将其与重组人粒-巨噬细胞集落刺激因子(rhGM-CSF)的基因相融合,并在原核细胞中表达。研究结果表明,大肠杆菌JM101表达的融合蛋白rhTpo/GM-CSF在体外小鼠骨髓造血祖细胞的培养中保留了Tpo对巨核细胞系和红细胞系的刺激作用,并增加了GM-CSF对粒细胞系的刺激活性。结论提示,原核细胞表达的融合蛋白rhTpo/GM-CSF具有刺激骨髓红细胞系、粒细胞系及巨核细胞系造血的活性。     

Modulation of neutrophil complement receptor 3 expression by pneumococci

Complement receptor 3 (CR3; CD18/CD11b) plays an important role in the recognition and clearance of Streptococcus pneumoniae (pneumococci) by neutrophils. The purpose of the present study was to characterize the modulation of CR3 surface expression on neutrophils exposed to pneumococci and to assess its functional significance. CR3 was detected with fluorescent phytoerythrin-labelled anti-CR3 (CD11b) antibodies, quantified with a fluorescence cell counter (FACS) and localized by confocal fluorescence microscopy. Uptake of fluorescent FITC-labelled pneumococci was quantified by FACS. Whole blood from healthy volunteers was exposed at 37 degrees C to killed whole type III Streptococcus pneumoniae (KSP; 10(8)/ml) or to a positive control ( Escherichia coli lipopolysaccharide) that enhanced CR3 surface expression on neutrophils to a comparable extent. Varying the concentration of KSP between 10(5) and 10(8) organisms/ml progressively augmented CR3 surface expression measured at 1 h, whereas the response declined at 10(9)/ml. The diminished response to 10(9) KSP/ml proved to be time-dependent, with surface CR3 up-regulated maximally within 5 min, and down-regulated thereafter. Labelling of CR3 during exposure demonstrated accelerated receptor sequestration, and confocal fluorescence microscopy demonstrated internalized CR3. Cooling to 16 degrees C, to inhibit the up-regulation of CR3 surface expression, also inhibited the uptake of FITC-labelled KSP and morphological changes. Accelerated down-regulation of surface CR3 expression by exposure to 10(9)/ml unlabelled KSP diminished the uptake of labelled KSP added subsequently. In contrast, lipopolysaccharide-induced up-regulation of CR3 expression increased the uptake of labelled KSP. Together, these experiments reveal dynamic modulation of CR3 expression on the surface of neutrophils exposed to pneumococci and a functional correlate of this modulation. Thus neutrophil expression of CR3 changes dynamically in response to exposure of neutrophils to progressively higher concentrations of pneumococci, conditions that mimic early neutrophil recruitment to densely infected lung tissue in acute pneumococcal pneumonia.     

Clinical pharmacokinetic studies of a human haemopoietic growth factor, GM-CSF

The pharmacokinetics of glycosylated recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) was studied following intravenous (i.v.) and subcutaneous (s.c.) bolus injection of rhGM-CSF, 8 micrograms kg-1 employing a sensitive radioimmunoassay. After a single i.v. bolus injection, an initial high serum level of rhGM-CSF was observed, followed by a rapid decrease that occurred in two phases with a half-life (t1/2) alpha of 20.0 +/- 5 min and a t1/2 beta of 68.3 +/- 8 min. Following s.c. bolus injection the absorption was more prolonged. Peak serum concentrations did not occur until about 15-20 h, and were followed by a more protracted elimination than by the i.v. route. In all patients the single rhGM-CSF injection led to an increase in peripheral white blood cells (WBC), after a temporary drop of 2-5 h duration. The increase in WBC was of longer duration after s.c. than after i.v. bolus treatment. Since the subcutaneous administration leads to prolonged serum concentration of rhGM-CSF and prolonged increase in peripheral WBC, it seems preferable to i.v. bolus injection, and as effective as continuous i.v. infusion.     

脐血清GM-CSF、IL-3和IL-6水平检测及意义

目的：探讨脐血的造血支持作用及其作用机理,为脐血的临床应用提供理论依据。方法：采用固相ＥＬＩＳＡ法检测３４例脐血清和２０例成人血清的ＧＭ－ＣＳＦ、ＩＬ－３和ＩＬ－６含量。结果：脐血清ＧＭ－ＣＳＦ、ＩＬ－３和ＩＬ－６含量分别为１９．１±１４．８、１９．２±１２．７和５７．５±２９．１ｐｇ／ｍｌ,成人血清ＧＭ－ＣＳＦ和ＩＬ－６未测出,ＩＬ－３含量为２．２±２．９ｐｇ／ｍｌ,差异均有极显著意义（Ｐ均＜０．００１）。结论：脐血含有丰富的造血生长因子,可用于再生障碍性贫血和化疗等原因引起的粒细胞减少的治疗。     

Proteolysis of the endothelial cell protein C receptor by neutrophil proteinase 3

BACKGROUND: The endothelial cell protein C receptor (EPCR) presents protein C to the thrombin:thrombomodulin complex on the endothelium of large vessels, and enhances the generation of activated protein C (APC) and activation of protease-activated receptor-1. A previous report has demonstrated binding of soluble (s) EPCR to activated neutrophils via surface proteinase 3 (PR3). METHODS: We now report further characterization of this interaction. Activated neutrophils and purified PR3 both decrease endothelial cell (EC) surface EPCR, suggestive of its proteolysis. RESULTS: When added to purified recombinant sEPCR, PR3 produced multiple cleavages, with early products including 20 kDa N-terminal and C-terminal (after Lys(176)) fragments. The binding of active site blocked PR3 to sEPCR was studied by surface plasmon resonance. Estimates of the K(D) of 18.5-102 nM were obtained with heterogeneous binding, suggestive of more than a single interaction site. CONCLUSIONS: This work demonstrates PR3 binding to and proteolysis of EPCR and suggests a mechanism by which anticoagulant and cell protective pathways can be down-regulated during inflammation.     

特尔立治疗白血病化疗后白细胞减少的临床观察及药物经济学

目的：比较国产与进口粒细胞巨噬细胞集落刺激因子(GM—CSF)在白血病化疗后骨髓抑制期的疗效及经济学价值。方法：选自1997—2001年住院急性白血病按随机对照方法分为特尔立组15例，生白能组13例。结果：两组在缩短骨髓抑制的天数和疗效无明显差异，但药物经济学有明显差别。结论：特尔立较进口同种制剂生白能疗效无明显差别，费用明显降低。     

Inhibition of IgE-mediated mast cell activation by the paired Ig-like receptor PIR-B

The potential of the paired Ig-like receptors of activating (PIR-A) and inhibitory (PIR-B) types for modifying an IgE antibody-mediated allergic response was evaluated in mouse bone marrow-derived mast cells. Although mast cells produced both PIR-A and PIR-B, PIR-B was found to be preferentially expressed on the cell surface, where it was constitutively tyrosine phosphorylated and associated with intracellular SHP-1 protein tyrosine phosphatase. PIR-B coligation with the IgE receptor (FcepsilonRI) inhibited IgE-mediated mast cell activation and release of serotonin. Surprisingly, the inhibitory activity of PIR-B was unimpaired in SHP-1-deficient mast cells. A third functional tyrosine-based inhibitory motif, one that fails to bind the SHP-1, SHP-2, and SHIP phosphatases, was identified in parallel studies of FcepsilonRI-bearing rat basophilic leukemia (RBL) cells transfected with constructs having mutations in the PIR-B cytoplasmic region. These results define the preferential expression of the PIR-B molecules on mast cells and an inhibitory potential that can be mediated via a SHP-1-independent pathway.