组织蛋白酶S和组织蛋白酶K在人曲张大隐静脉平滑肌细胞中韵表达

目的：探讨组织蛋白酶S（Cats）、组织蛋白酶K（CatK）在曲张大隐静脉平滑肌细胞（SMC）中的表达变化。方法：术中收集曲张及正常大隐静脉标本，采用免疫组织化学显色方法、免疫印迹法检测CatS、CatK在曲张和正常大隐静脉管壁的表达。结果：CatS和CatK在曲张大隐静脉中免疫反应阳性，阳性细胞主要位于SMC胞质，曲张组SMC中CatS和CatK阳性细胞平均光密度值较正常组明显增高，其蛋白表达明显高于正常组，免疫荧光显色可见CatS、CatK与曲张大隐静脉SMC共定位。结论：CatS和CatK在曲张大隐静脉SMC中表达明显增高，表明两者可能参与了静脉曲张的发生，并可能作为该疾病的生物学标志。     

组织蛋白酶L及其抑制剂Cystatin C在曲张大隐静脉平滑肌细胞中的表达

目的:组织蛋白酶L(Cat L)及其抑制剂Cystatin C在曲张大隐静脉平滑肌细胞(SMC)中的表达。方法:术中收集曲张及正常大隐静脉标本,采用免疫组织化学、荧光免疫组织化学染色方法及计算机图像分析技术观察检测。结果:免疫组化染色可见Cat L、Cystatin C分别在曲张、正常大隐静脉中免疫反应阳性,阳性细胞主要位于SMC胞质;曲张组SMC中Cat L阳性细胞平均光密度值明显增高,Cystatin C明显降低,与正常组间差异显著;荧光免疫组织化学染色可见Cat L与曲张大隐静脉SMC共定位,Cystatin C与正常大隐静脉SMC共定位。结论:在曲张大隐静脉的SMC中Cat L表达增强,Cystatin C表达下降。这一改变可能作为大隐静脉曲张的生物学标志。     

粥样硬化动脉壁胶原含量和组织蛋白酶B的表达

目的:检测动脉粥样硬化(AS)血管壁胶原含量和组织蛋白酶B的表达水平,探讨其对AS发生发展的作用。方法:9例AS患者(AS组)和5例正常成人(对照组),采用Van Gieson法染色检测其动脉血管胶原含量,应用免疫组织化学染色(SP法)和Western-blotting检测组织蛋白酶B的表达。结果:与对照组相比,AS组胶原含量明显升高(P<0.05),组织蛋白酶B的表达也明显升高(P<0.05)。结论:组织蛋白酶B可能通过破坏细胞外基质促进AS进展。     

基质金属蛋白酶-9及其组织抑制物-1在妊娠输卵管黏膜中的表达

目的:检测基质金属蛋白酶-9(MMP-9)及其组织抑制物-1(TIMP-1)在输卵管黏膜中的表达,探讨与输卵管妊娠的关系。方法:采用免疫组织化学显色技术和图像半定量分析法,检测MMP-9和TIMP-1在人妊娠输卵管黏膜、人正常输卵管黏膜及正常宫内早孕子宫蜕膜组织中的表达。结果:MMP-9和TIMP-1在正常宫内早孕组中表达最强,在输卵管妊娠组中的表达较强,在正常输卵管组中表达较弱。两两比较差异均有显著性。结论:MMP-9/TIMP-1参与了输卵管妊娠中胚胎着床过程,且与输卵管妊娠缺乏蜕膜化反应有关。     

组织蛋白酶B在大鼠肝脏的分布

目的研究组织蛋白酶B(cathepsins B,CB)在正常SD大鼠肝脏组织中的表达。方法采用免疫荧光组织化学的方法,观察CB在大鼠肝脏组织中的分布。结果 CB免疫反应阳性细胞主要分布在肝细胞、枯否细胞、肝窦内皮细胞,枯否细胞、肝窦内皮细胞的免疫活性比较强,肝细胞的免疫活性比较弱,而肝星状细胞没有CB免疫活性。阳性物质分布于细胞质,细胞核阴性。结论 CB在大鼠肝脏组织的表达提示其可能在肝脏起着非常重要的作用。     

泛素蛋白酶系统在四氧嘧啶诱导的糖尿病大鼠视网膜中的表达

目的:建立泛素蛋白酶系统(UPS)在正常和8周糖尿病大鼠视网膜的表达图谱,探讨其在糖尿病视网膜病变(DR)发生发展中可能的分子机制。方法:在限制片段差异显示PCR(RFDD-PCR)技术建立正常和8周糖尿病大鼠视网膜基因表达谱的基础上,对差异片段进行生物信息学分析,筛选UPS DR相关基因,并以免疫组织化学、半定量RT-PCR技术进行验证。结果:RFDD-PCR结果显示,泛素蛋白酶系统DR相关基因UBS3A、PSMD8和PSMD11在糖尿病组表达上调。RT-PCR结果显示,糖尿病组UBS3A表达比正常组明显增高,PSMD8和PSMD11则仅在糖尿病组中表达。免疫组织化学结果显示,正常组UBE3A阳性免疫反应物见于内丛状层,外丛状层和锥、杆体细胞层,PSMD8和PSMD11未见阳性细胞;糖尿病组UBE3A、PSMD8和PSMD11阳性细胞明显增多,见于节细胞层、内核层和外核层。结论:UPS与DR发生发展有关。     

人组织蛋白酶K基因研究进展

组织蛋白酶K(cathepsin K,CTSK)是一种溶酶体半胱氨酸蛋白酶,属于木瓜蛋白酶超家族成员,是近年来发现的存在于溶酶体的细胞内蛋白酶中重要的一个细胞外基质降解酶类.组织蛋白酶K不仅参与机体正常生理调节,如在破骨细胞中表达、与骨吸收和骨重塑相关、对软骨胞外基质Ⅱ型胶原有降解作用,而且在多种实体瘤中都存在组织蛋白酶K异常表达.该文综述了CTSK基因的功能及其与多种疾病和肿瘤的相关性.     

金属蛋白酶-9在糖尿病肾病发病机制中的作用

目的:探讨大鼠糖尿病肾病发展过程中肾局部金属蛋白酶-9(MMP-9)与糖尿病肾病发生的相关性.方法:健康SD大鼠,随机分成糖尿病组和正常对照组,糖尿病组大鼠采用一次性腹腔内注射链脲佐菌素诱导制造糖尿病大鼠模型.分别于造模后第4、 8周通过免疫组织化学方法观察肾局部MMP-9的表达,测其肾小球阳性细胞表达率;应用免疫印迹法对MMP-9在肾组织不同时间点的含量进行定量检测.结果:MMP-9在正常肾小球呈强阳性表达,造模后4、 8周末,糖尿病组大鼠MMP-9在肾小球表达均下降.正常对照组MMP-9含量最高,糖尿病4、8周组大鼠肾脏MMP-9含量明显下降.结论:糖尿病肾病时肾组织中MMP-9表达水平降低,可能是糖尿病早期病理改变的重要原因之一.     

组织蛋白酶参与皮肤生理的研究进展

组织蛋白酶(cathepsin,Cat)是半胱氨酸蛋白酶家族的主要成员,与皮肤肿瘤、慢性炎症等多种皮肤病密切相关.近年来多项研究表明Cat在人类皮肤生理方面的重要意义,主要集中于CatD/CatL/CatV维持表皮正常分化和完整屏障,CatB/CatD/CatG/CatK维持真皮细胞外基质(extracellular matrix,ECM)的稳态,CatV保证毛囊正常发育和生长周期.这些研究已深入细胞信号、基因调控,增进对正常皮肤生理调控的理解,未来也可能应用到疾病的基因治疗领域.     

人脑桥静脉平滑肌细胞α-肌动蛋白的表达

方法:人脑标本12例,共计62支桥静脉.应用H-E染色、丽春红-维多利亚蓝组合染色观察桥静脉SMC的组织学特点,运用免疫组织化学显色、免疫荧光化学显色和免疫印迹法检测平滑肌特异标记物α-平滑肌肌动蛋白(α-SMA)的表达.结果:桥静脉管壁中膜可见散在、孤立的SMC,大多呈纵切面,α-SMA在桥静脉中免疫反应阳性;桥静脉流出端硬脑膜侧α-SMA表达高于蛛网膜侧.结论:桥静脉管壁中有SMC的存在,且分布不对称.     

Immunohistochemical expression of cathepsin D in meningiomas

We retrospectively studied the expression of cathepsin D by immunohistochemical analysis in 86 meningiomas (World Health Organization [WHO] grade I, n = 44; WHO grade II, n = 21; WHO grade III, n = 21) and correlated the results with tumor grade and outcome. Staining was scored semiquantitatively based on distribution among neoplastic cells as follows: 0, no staining; 1+, 5% or less of the cells; 2+, 6% to 20%; 3+, 21% to 50%; and 4+, more than 50% of the cells. Cathepsin D expression was observed as follows: 0, 10 cases (12%); 1+, 25 cases (29%); 2+, 15 cases (17%); 3+, 12 cases (14%); and 4+, 24 cases (28%). A higher degree of cathepsin D immunostaining was associated with low tumor grade (P = .0014), low mitotic count (P < .0001), low apoptotic count (P < .0001), and the development of recurrence (P = .035). There was no correlation with outcome or MIB-1 proliferation index. Cathepsin D expression by immunohistochemical analysis was identified in the majority (88% [76/86]) of meningiomas studied. A greater degree of immunoreactivity was observed in the WHO grade I group.     

Activation of neutrophil collagenase by cathepsin G

Collagenase is secreted from neutrophils as a latent or proenzyme. In an effort to understand the mechanism of collagenase activation in inflammation, human peripheral neutrophils (PMNs) were isolated and incubated with the tumor promotor, phorbol myristate acetate (PMA), which induces the neutrophils to degranulate and secrete proteinases. Neutrophil media were then treated with various activators or inhibitors of collagenase and other proteinases, and the collagenase activity was measured. A serine proteinase secreted from neutrophils, cathepsin G, was found to activate latent collagenase, but it was also found to require activation itself. Both hypochlorous acid (HOCl) and oxidized glutathione (GSSG) were tested for their coilagenase-activating ability and were found to be successful only in the presence of active cathepsin G. A specific cathepsin G inhibitor (0.5 mM Z-Gly-Leu-Phe-CH₂Cl) prevented the activation of latent collagenase by HOCl. To confirm these results, purified neutrophil cathepsin G was incubated with a neutrophil proteinase mixture which contained latent collagenase. The collagenase was shown to be activated upon incubation with purified cathepsin G. These results indicate that cathepsin G is a key mediator in neutrophil collagenase activation.     

Novel cathepsin B and cathepsin B-like cysteine protease of Naegleria fowleri excretory–secretory proteins and their biochemical properties

Naegleria fowleri causes a lethal primary amoebic meningoencephalitis (PAM) in humans and experimental animals, which leads to death within 7–14 days. Cysteine proteases of parasites play key roles in nutrient uptake, excystment/encystment, host tissue invasion, and immune evasion. In this study, we cloned N. fowleri cathepsin B (nfcpb) and cathepsin B-like (nfcpb-L) genes from our cDNA library of N. fowleri. The full-length sequences of genes were 1,038 and 939 bp (encoded 345 and 313 amino acids), and molecular weights were 38.4 and 34 kDa, respectively. Also, nfcpb and nfcpb-L showed a 56 and 46 % identity to Naegleria gruberi cathepsin B and cathepsin B-like enzyme, respectively. Recombinant NfCPB (rNfCPB) and NfCPB-L (rNfCPB-L) proteins were expressed by the pEX5-NT/TOPO vector that was transformed into Escherichia coli BL21, and they showed 38.4 and 34 kDa bands on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis using their respective antibodies. Proteolytic activity of refolded rNfCPB and rNfCPB-L was maximum at a pH of 4.5, and the most effective substrate was Z-LR-MCA. rNfCPB and rNfCPB-L showed proteolytic activity for several proteins such as IgA, IgG, IgM, collagen, fibronectin, hemoglobin, and albumin. These results suggested that NfCPB and NfCPB-L cysteine protease are important components of the N. fowleri ESP, and they may play important roles in host tissue invasion and immune evasion as pathogens that cause N. fowleri PAM.     

Antigonococcal activity of human neutrophil cathepsin G.

We have shown that lysosomal cathepsin G, prepared from acid extracts of granules derived from human polymorphonuclear granulocytes, exhibits potent in vitro antimicrobial activity against Neisseria gonorrhoeae. An isolated isozyme of cathepsin G was found to exhibit antigonococcal activity by a nonenzymatic mechanism in a time-dependent manner. Moreover, we observed that the antigonococcal activity of cathepsin G was relatively independent of pH and evident over a pH range resembling that invoked for maturing phagolysosomes. Using a number of isogenic strains, we determined that certain mutations known to alter cell envelope structure rendered gonococci more susceptible to cathepsin G. This suggests that the susceptibility of gonococci to cathepsin G, and possibly other antimicrobial proteins derived from PMN granules, is genetically determined and possibly related to the structure of the gonococcal cell envelope.