Matrix metalloproteinases stimulate epithelial-mesenchymal transition during tumor development

Matrix metalloproteinases (MMPs) are a family of more than 28 enzymes that were initially identified on the basis of their ability to cleave most elements of the extracellular matrix (ECM) but have subsequently been found to be upregulated in nearly every tumor type. As digestion of the ECM is essential for tumor invasion and metastasis, MMPs have been studied for their role in these later stages of tumor development. More recently, exposure to these enzymes has been found to impact cellular signaling pathways that stimulate cell growth at early stages of tumor progression. MMPs have also been found to cleave intracellular targets and so inducing mitotic abnormalities and genomic instability. Emerging evidence indicates that tumor-associated MMPs can also stimulate processes associated with epithelial-mesenchymal transition (EMT), a developmental process that is activated in tumor cells during cell invasion and metastasis. Investigations of potential therapeutic MMP inhibitors aimed at blocking the protumorigenic tissue alterations induced by MMPs have been complicated by the side effects associated with nonspecific inhibition of normal physiological processes; recent investigations have shown how delineation of the extracellular targets and intracellular signaling pathways by which MMP action on cancer cells can induce EMT provides insight into novel therapeutic targets. Here, we provide an overview of recent findings of MMP action in tumors and the mechanisms by which MMPs induce both phenotypic and genotypic alterations that facilitate tumor progression.     

晚期糖基化终末产物通过氧化应激诱导大鼠软骨细胞损伤

目的:探讨晚期糖基化终末产物(advanced glycation end products,AGEs)能否通过氧化应激引起大鼠软骨细胞损伤。方法:原代培养SD大鼠软骨细胞,对细胞表型进行鉴定;应用CCK-8法检测软骨细胞生存率;DCFH-DA染色荧光显微镜下检测胞内活性氧簇(reactive oxygen species,ROS)的水平;Hoechst 33342核染色法及Annexin V-FITC/PI流式细胞法测定软骨细胞的凋亡率;RT-PCR法检测软骨细胞中Bax、Bcl-2、caspase-3、MMP3、MMP13和COL2的mRNA水平;Western blotting法检测软骨细胞中cleaved caspase-3、MMP3、MMP13和COL2的蛋白水平。结果:与对照组相比,AGEs可显著上调胞内ROS水平(P0.05),但经抗氧化剂N-乙酰半胱氨酸(NAC)抑制后ROS的生成明显减少(P0.05);另外,NAC可抑制AGEs引起的软骨细胞凋亡相关分子Bax/Bcl-2和caspase-3水平的上调,并减少MMP3和MMP13表达及COL2的丢失(P0.05)。结论:AGEs可通过氧化应激诱导大鼠软骨细胞损伤。     

Antagonism of sulfadoxine and pyrimethamine antimalarial activity in vitro by p-aminobenzoic acid, p-aminobenzoylglutamic acid and folic acid

The activity of pyrimethamine and sulfadoxine against two strains of Plasmodium falciparum has been studied in vitro by a radioisotopic technique. Low level antagonism of pyrimethamine resulted from the inclusion of p-aminobenzoic acid, p-aminobenzoylglutamic acid or folic acid in the test medium. Sulfadoxine activity was antagonised slightly by p-aminobenzoic but not by p-aminobenzoylglutamic acid, and antagonised markedly by folic acid at concentrations above 4 X 10(-8) M. At 10(-7) M folic acid, a concentration lower than that of normal RPMI medium 1640, sulfadoxine activity was reduced 7000 to 9000-fold in comparison with controls. These results are of importance in terms of the utilisation of folates by P. falciparum, the susceptibility of the parasite to antifolate drugs and the in vitro determination of parasite susceptibility.     

Matrix metalloproteinases activation in Toxocara canis induced pulmonary pathogenesis

BackgroundToxocara canis, a source of visceral larva migrans, causes toxocariasis and induces respiratory symptoms. The reasons by which the pulmonary pathological alteration in the lungs infected with T. canis remain unclear.MethodsThe involvement of the pulmonary pathological alteration by histology, enzyme activity, and Western blot analysis in the lungs of BALB/c mice after the infection of 2000 embryonated eggs.ResultsThe pathological effects gradually increased after the infection culminated in severe leukocyte infiltration and hemorrhage from days 4–14 post-inoculation. Gelatin zymography using substrate showed that the relative activity of matrix metalloproteinase (MMP) −9 and MMP-2 significantly increased in T. canis-infected mice. Western blot analysis indicated that the MMPs protein level of fibronectin monomer significantly increased in T. canis-infected mice compared with that in uninfected control. T. canis larvae mainly initiated leukocyte infiltration and hemorrhage in the lungs.ConclusionThese phenomena subsequently induced the activities of MMPs in parallel with the pathological changes in early stage pulmonary inflammation. In conclusion, T. canis larval migration activated the MMPs and caused pulmonary pathogenesis.     

血清基质金属蛋白酶活性与动脉粥样硬化斑块稳定性的实验观察

目的观察血清MMP-2、MMP-9在不同时期的兔动脉粥样硬化动物模型中的变化,研究其影响因素及与斑块稳定性的关系。方法将30只长耳白兔随机分为3组:普通饮食组(n=10),高脂饮食组(n=10),球囊损伤 高脂饮食组(n=10),3个月后以中国斑点蝰蛇毒和组胺触发使斑块破裂。其间监测血脂情况,并行血清MMP-2、MMP-9检测。动物处死后查找动脉硬化斑块行病理检测。结果普通饮食组实验中各检测项目均无明显差异。高脂饮食及球囊损伤 高脂饮食组一个月后血脂明显升高。药物触发后高脂饮食组血清MMP-9活性明显增加,球囊损伤 高脂饮食组血清MMP-2、MMP-9活性均明显增加。结论在兔动脉粥样硬化的动物模型中,斑块处于不稳定状态时血清MMP-9活性明显增加,而MMP-2的活性升高可能与内皮机械损伤有关。     

miR-30c抑制宫颈癌细胞恶性表型的分子机制研究

目的:探讨微小RNA(miR)-30c过表达抑制宫颈癌细胞恶性表型的分子机制。方法:运用Lipofectamine 2000转染法将pGenesil-1-miR-30 c质粒转染宫颈癌细胞系C33A、HeLa、SiHa和CaSki,以转染p-Geresil-1的细胞为阴性对照;运用TaqMan real-time PCR检测各组细胞中miR-30c的表达水平;采用MTT法、集落形成实验、Transwell法、Annexin V-FITC及流式细胞术分别测定细胞活力抑制率、集落形成能力、迁移率及凋亡率;Western blot检测Bax、Bcl-2、基质金属蛋白酶(MMP)-9、MMP-13和金属蛋白酶组织抑制物1(TIMP-1)的蛋白表达水平。结果:转染p Genesil-1-miR-30c质粒的宫颈癌细胞系中miR-30c表达水平均显著高于阴性对照组(P0.01);过表达miR-30 c的宫颈癌细胞活力抑制率显著高于阴性对照组(P0.05),细胞集落形成率和迁移率显著低于阴性对照组(P0.05);流式细胞术检测结果显示,miR-30c过表达的宫颈癌细胞凋亡率显著高于对照组(P0.05);Western blot结果显示miR-30 c过表达促进Bax和TIMP-1蛋白的表达,而抑制Bcl-2和MMP-13蛋白的表达(P0.05或P0.01)。结论 :miR-30 c过表达抑制宫颈癌细胞活力和迁移,诱导宫颈癌细胞凋亡,其机制与激活细胞凋亡通路及抑制MMP-13表达有关。     

胸腺上皮肿瘤组织中基质金属蛋白酶-2活性与Ⅰ型膜型基质金属蛋白酶 mRNA表达的相关性

目的探讨胸腺瘤和胸腺癌中基质金属蛋白酶（MMP）-2、Ⅰ型膜型（MT1）-MMP、金属蛋白酶组织抑制剂（TIMP）-2mRNA的表达和MMP-2蛋白活性的关系。方法分别用real-time逆转录．聚合酶链反应（RT-PCR,Taqman法）、明胶酶谱法和Filmin situ gelatin-Zymography（FIZ）对正常的胸腺组织（2例）、胸腺瘤（12例）和胸腺癌（2例）患者的新鲜肿瘤组织中MMP-2、MT1-MMP、TIMP-2mRNA的表达,pro-MMP-2的活性率及活性蛋白的定位进行测定。结果MMP-2、MT1-MMP及TIMP-2mRNA在Ⅰ期与Ⅱ期和Ⅲ期与Ⅳ期中的表达差异均无统计学意义（P〉0．05）,在Ⅰ～Ⅱ期与Ⅲ～Ⅳ期和胸腺癌3组中差异均有统计学意义（P〈0．01）。在AB、B1型（混合型和淋巴细胞为主型）与B2、B3型（皮质型和多角细胞为主型）以及胸腺癌3组中差异均有统计学意义（P〈0．05）。MMP-2的蛋白活性率（MMP-2／pro—MMP-2＋MMP-2）在Ⅰ～Ⅱ期、Ⅲ～Ⅳ期和胸腺癌各组中差异有统计学意义（P〈0．05）,在AB、B1型与B2、B3型以及胸腺癌各组中的差异均具有统计学意义（P〈0．05）。胸腺瘤各期及各型中MT1-MMP、TIMP-2mRNA与MMP-2蛋白活性表达均呈正相关,且相关程度相似（r=0．7235、r=0．7647、P〈0．005）。MMP-9的蛋白表达在各组间差异均无统计学意义。结论MMP-2、MT1-MMP及TIMP-2的mRNA表达与胸腺瘤临床分期、病理分型相关,MMP-2的活性与MT1-MMP和TIMP-2的表达升高正相关。推测MT1-MMP通过TIMP-2对MMP-2的激活起促进作用。     

Dithiocarbamates inhibit IL-1-induced cartilage degradation in bovine articular cartilage explants

Interleukin-1-stimulated cartilage degradation in bovine articular cartilage explants is effectively inhibited by several different dithiocarbamates with IC₅₀'s in the micromolar range.accepted by W. B. van den BergSupported by OsteoArthritis Sciences, Inc.     

抗人基质金属蛋白酶-2的单抗制备及功能研究

目的在体外原核表达MMP-2纤维连接蛋白样片段(MFD),制备抗MMP-2特异性的单克隆抗体(McAb),以期得到能够有效抑制MMP-2活性的单抗。方法运用原核表达的MFD蛋白纯化浓缩后免疫BALB/C小鼠,尾静脉加强后取脾细胞融合,ELISA法筛选阳性克隆,制备腹水。运用Westernblotting对抗体进行鉴定,并观察抗体与天然MMP-2的反应情况,进行亚型鉴定,组织特异性鉴定同时进行功能实验增殖实验、侵袭实验以及体外血管新生实验。结果小鼠脾细胞与杂交瘤细胞融合后筛选获得一株持续分泌单克隆抗体的细胞株,命名为SZ-117。ELISA法测定上清和腹水效价分别为2×10-3和2×10-5。抗体为IgG1亚型,可以与天然的MMP-2相结合,与血细胞表面没有交叉反应。免疫组化提示胃、胆囊、脾脏、卵巢、前列腺、输卵管以及淋巴结的间质中都有阳性表达,而在甲状腺、小肠、肝脏等组织器官未见阳性表达。该抗体可以有效地抑制血管内皮细胞株Eahy926细胞以及胰腺癌细胞株1990细胞的侵袭行为,同时还可以抑制Eahy926细胞在体外的血管生成。结论抗MMP-2特异性抗体的获得为MMP-2测定提供了一个有效工具,并且该抗体还可以有效地抑制内皮细胞在体外的血管生成和肿瘤细胞的侵袭行为,有望成为一种新的抗肿瘤物质。     

一氧化氮与基质金属蛋白酶介导的血管重构在动脉瘤中的作用

Nitric oxide (NO) and matrix metaUoproteinases (MMPs) are important in vascular remold-ing, especially in abdominal aortic aneurysm. NO may be associated with aneurysms by modulating MMPs expression and activity.     

妊娠期基质金属蛋白酶的调控及其与胎盘发育的关系

 基质金属蛋白酶（matrix metalloproteinases，MMP） 是一类蛋白水解酶，可以降解多糖以外的细胞外基质（extracellular matrix，ECM）的多种成分，在组织塑型过程中影响着 ECM 的代谢、新生血管的形成等过程。在胎盘发育过程中滋养细胞的入侵包括细胞基质的溶解和重塑，其生物学行为与有着高度侵蚀性的肿瘤细胞相似，又被称为假恶性细胞。MMP 是降解蜕膜基质的重要酶类，可以降解几乎所有基质成分，在胚胎植入、胎盘形成及子宫螺旋动脉重构中发挥重要作用。MMP 在滋养层不同类型细胞中的分布存在特异性，在不同时期胎盘中的表达和活性也有所不同，妊娠期 MMP 的调控直接影响胎盘的发育，对维持正常妊娠至关重要。本文主要就妊娠期 MMP 的调控及其与胎盘发育关系的研究进展做一综述。 1 MMP 分类及特点 MMP 是一族 Zn2+ 依赖性内肽酶，至今已经发现至少26 种^[1]。根据蛋白结构的相似性以及降解底物不同，MMP 通常被分为 5 个亚类：①间质胶原酶（MMP-1、MMP-8和MMP-13），来源于纤维细胞、癌细胞及滋养细胞，降解底物为 Ⅰ~ Ⅲ 型胶原等。②明胶酶（MMP-2、MMP-9），部分来源于滋养细胞、纤维细胞及巨噬细胞等，酶解底物为明胶变性胶原和 Ⅳ 型胶原等。③基质溶解素（MMP-3、MMP-7、MMP-10、MMP-11、MMP-12 和 MMP-26），来源于成纤维细胞和癌细胞等，降解底物为 Ⅲ ~ Ⅴ 型胶原、明胶、蛋白聚糖及糖蛋白等。④膜型金属蛋白酶（MT-MMP），包括 MT1-MMP、MT2-MMP、MT3-MMP、MT4-MMP、MT5-MMP 及 MT6-MMP，存在于肿瘤细胞表面及滋养细胞表面，降解底物为 Ⅳ 型胶原、明胶等。有证据表明，MT1-MMP、MT2-MMP、MT3-MMP 和 MT5-MMP 参与激活 MMP-2^[2]。⑤其他基质金属蛋白酶，包括 MMP-19、MMP-20、MMP-23 等，可以降解抗胰蛋白酶及釉质。     

MMP-1、MMP-2在盆底器官脱垂疾病患者阴道组织中的表达及意义

 目的 研究基质金属蛋白酶MMP-1、MMP-2在盆底器官脱垂患者阴道组织中的表达情况,从而探讨基质金属蛋白酶与女性盆底障碍性疾病( PFD) 的关系。方法 选择我院妇产科收治的盆底器官脱垂患者20例为研究组,20例非盆底器官功能障碍患者作为对照。RT-PCR方法检测不同组MMP-1mRNA 的表达情况,明胶底物酶方法测定阴道前壁组织中MMP-1、MMP-2的表达。结果 脱垂患者阴道组织中MMP-1的表达明显增加,但是两组患者阴道壁组织中MMP-2的表达没有明显差别,而MMP-2前体在脱垂患者阴道壁组织中表达明显增加。结论 MMP-1使盆底组织胶原代谢分解增加,增加的MMP-1表达与盆底器官脱垂有关。     

Protein kinase C activation by phorbol ester increases in vitro invasion through regulation of matrix metalloproteinases/tissue inhibitors of metalloproteinases system in D54 human glioblastoma cells

To elucidate possible mechanisms of phorbol 12-myristate 13-acetate (PMA) induced in vitro invasiveness of glioblastoma cells, we examined expression levels of membrane-type 1 matrix metalloproteinase (MT1-MMP), MMP-2, MMP-9 and tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 using Western blotting and gelatin zymography assay, and found that PMA induced the secretion of MMP-9, activated MMP-2 proenzyme to fully active form of 59 kDa, down-regulated the TIMP-1 and TIMP-2 secretion, and increased MT1-MMP on the cell surface. However, PKC inhibitor Go 6983 reversed all of these effects brought about by PMA. We, therefore, conclude the activation of PKC by PMA in these cells plays a critical role in the regulation of MMPs/TIMPs system, which has a major role in tumor invasion and metastasis.     

Breast cancer progression: insights into multifaceted matrix metalloproteinases

The restricted view of matrix metalloproteinases (MMPs) as simple destroyers of extracellular matrix components has largely ignored their substantial contribution in many aspects of cancer development and metastatic dissemination. Over the last few years, the relevance of MMPs in the processing of a large array of extracellular and cell surface-associated proteins has grown considerably. Our knowledge about the complex functions of MMPs and how their contribution may differ throughout cancer progression is rapidly expanding. These new findings provide several explanations for the lack of success of MMP inhibition in clinical trials. A complete understanding of MMP biology is needed before considering them, their substrates or their products as therapeutic targets. In this review, we explore the different faces of MMP implication in breast cancer progression by considering both clinical and fundamental aspects.     

太子参对心肌梗死后慢性心衰大鼠心功能与基质金属蛋白酶表达的影响

目的：研究太子参对心肌梗死诱导的慢性心衰大鼠心功能与基质金属蛋白酶（MMPs）表达和活性的作用。方法:采用结扎大鼠冠状动脉复制急性心肌梗死诱导慢性心衰动物模型，以血流动力学指标分析心功能，RT-PCR分析MMP-2与MMP-9的mRNA表达，酶谱法分析MMP-2与MMP-9的活力。结果:大鼠冠脉结扎6周后，心功能显著紊乱；左心室组织MMP-2与MMP-9 mRNA水平显著提高、酶活力增加。太子参水煎液连续灌胃给药5周，可显著改善心衰大鼠的血流动力学，抑制左心室组织MMP-2与MMP-9的活力和mRNA的水平增加（与模型组比较，差异显著，P<0.01，P<0.05）。结论:大鼠冠脉结扎6周形成慢性心衰模型，太子参水煎液可改善大鼠冠脉结扎所诱导的心衰，其机制可能与MMPs的表达和活性有关。     

基质金属蛋白酶-9与炎症反应研究进展

MMP - 9 is one of the family of matrix metalloproteinases, which degrades extracellular matrix. MMP- 9 is induced by many inflammatory factors, including IL - 1β, IL - 8 and TNF - α. Mean-while, MMP - 9 potentiates inflammatory factors by aminoterminal processing. The signal transduction by which inflammatory factors induce MMP - 9 is also an important part in recent research. This review will give a sunmamy in all these fields.     

前列腺间质细胞中雌二醇对MMP-2，MMP-9及其组织抑制因子表达的影响

目的： 研究雌二醇(E2)对前列腺间质细胞中基质金属蛋白酶(MMP-2、MMP-9）及其组织抑制因子1(TIMP-1)、TIMP-2和雌激素受体α、β（ERα、ERβ）的影响。方法： 实时定量PCR法检测E2在前列腺间质细胞中对MMP-2、MMP-9、TIMP-1、TIMP-2 mRNA水平的影响；半定量RT-PCR法检测E2对ERα、ERβ mRNA水平的影响。酶谱电泳法检测MMP-2、MMP-9的活性。Western blotting检测E2在前列腺间质细胞中对ERα蛋白水平的影响。结果： 前列腺间质细胞中有MMP-2和ERα mRNA的表达，未检测到MMP-9 mRNA；培养液中检测到MMP-2前体（pro-MMP-2），未检测到其活性形式，也未检测到MMP-9前体及其活性形式。用E2处理间质细胞后MMP-2 mRNA水平降低，pro-MMP-2蛋白量减少，雌激素受体抑制剂ICI 182.780可抑制此作用。E2对TIMP-1,2 mRNA表达无显著影响。E2能够增加前列腺间质细胞中ERα mRNA及其蛋白表达的水平。结论： E2能够通过ERα下调前列腺间质细胞中MMP-2的表达。     

基质金属蛋白酶及其组织型抑制剂活性及表达失衡与高血压性左心室重塑的关系

目的： 探讨慢性压力负荷增高时，大鼠左心室（LV）基质金属蛋白酶（MMPs）/组织型基质金属蛋白酶抑制剂（TIMPs）失衡与LV重塑的关系。方法：40只6周龄雄性卒中易感性自发性高血压大鼠（SHR-SPs）作为研究对象，10只同周龄雄性Wistar-Kyoto（WKY）大鼠作为对照。6个月后，以Millar压力容积导管评价2组大鼠的在体LV血流动力学，并对2组大鼠的心脏进行组织病理学、明胶酶谱和免疫印迹法分析。结果：反映LV收缩与舒张功能的血流动力学参数在2组间有显著差异（P＜0.05）；SHR-SPs心脏胶原容积分数、血管周胶原面积/管腔面积、心肌横断面积、心室壁动脉中膜面积/管腔面积均增高（P＜0.05）；心肌MMP-2活性、蛋白含量及TIMP-1蛋白含量在SHR-SPs中明显增高（P＜0.05）。结论：慢性压力超负荷能够导致心脏细胞外基质代谢失调及MMPs/TIMPs系统失衡，继而产生心室腔扩张、LV收缩与舒张功能障碍。     

MMP-9/TIMP-1表达在糖尿病鼠皮肤伤口愈合过程中的变化及意义初探

目的：观察基质金属蛋白酶-9（MMP-9）、组织金属蛋白酶抑制剂-1（TIMP-1）的表达和MMP-9/TIMP-1比值在糖尿病组和正常组大鼠皮肤伤口愈合过程中不同时点表达的动态变化，探讨其可能的作用机制。 方法:糖尿病大鼠形成6周后行皮肤伤口造模，采用HE染色、Masson染色和免疫组织化学方法评估伤口形成后3、7、14 d伤口组织的再上皮化、炎症细胞浸润、肉芽组织厚度、新生血管形成和胶原纤维密度的情况；通过逆转录-聚合酶链反应(RT-PCR)和Western印迹检测术后不同时点MMP-9、TIMP-1在伤口组织中的表达情况。结果:糖尿病大鼠伤口愈合明显迟缓。术后第3 d两组间胶原纤维、肉芽组织、新生血管和再上皮化没有差异，术后第7 d糖尿病组以上指标得分均低于正常组，第14 d这种趋势更加明显；而第3 d至14 d，糖尿病组的单核巨噬细胞浸润得分均低于正常组。术后第3 d两组MMP-9表达均达高峰，第7、14 d呈逐渐下降趋势，糖尿病组MMP-9表达水平在各时点均高于正常组；术后第3 d两组TIMP-1表达均达高峰，第7、14 d呈逐渐下降趋势，糖尿病组TIMP-1表达水平在各时点均低于正常组；正常组MMP-9/TIMP-1蛋白水平比值始终维持在一个动态平衡的稳定水平，而糖尿病组却长期处于高水平状态。结论:异常的胶原产生、新生血管重建、炎症反应、再上皮化、肉芽形成可能是糖尿病鼠创面愈合减慢的组织病理学基础；皮肤组织MMP-9/TIMP-1的平衡性改变可能是这种组织病理学异常的重要原因之一。     

Matrix metalloproteinases-2 and -3 are reduced in cerebrospinal fluid with low beta-amyloid1–42 levels

Matrix metalloproteinases (MMPs), a family of extracellular soluble or membrane bound endopeptidases, are implicated in many physiological and pathophysiological functions—based on their capability to cleave all protein components of the extracellular matrix. Recent studies have implicated several forms of MMPs in chronic neurodegenerative diseases like Alzheimer's disease (AD), vascular dementia (VD), and Parkinson's disease (PD). The aim of the present study was to analyse eight MMPs (MMP-1, -2, -3, -7, -8, -9, -10, -13) in the human cerebrospinal fluid (CSF) and to correlate with the well established biomarkers beta-amyloid_1–42 (Aβ), total-tau and phospho-tau-181. Our data show a significant decrease of MMP-2 and MMP-3 levels in the CSF in samples with significantly reduced Aβ levels. It is concluded that MMP-2 and MMP-3 are directly linked to Aβ in the brain and a dysfunction may influence the processing of Aβ.