膜型基质金属蛋白酶

生命科学研究LIFE SCIENCE RESEARCH1999年 第3卷 第3期 Vol.3 No.3 1999基质金属蛋白酶吴二喜　王凤飞　Norman McKIE摘　要：基质金属蛋白酶是一类分解细胞外基质组分的锌蛋白酶。

它们在有机体生长发育中的细胞外基质逆转与重塑以及疾病中的病理损害起着极为重要的作用。

基质金属蛋白酶的表达和活性在不同细胞水平受到严密调控，如细胞因子、生长因子以及激素的调节。

基质金属蛋白酶以酶原形式分泌，随后被其它蛋白酶如胞浆素或非蛋白酶类化学物质如有机汞所激活。

所有基质金属蛋白酶都受到天然抑制剂金属蛋白酶组织抑制剂所抑制。

两者的不平衡导致许多疾病的发生，如肿瘤侵入及转移。

合成基质金属蛋白酶组织抑制剂所抑制，如Marimastat能控制肿瘤转移的发生及进一步扩散。

本文将对基质金属蛋白酶的特征、分子区域结构、底物特性、激活机制、调控方式等方面进行最新概述。

关键词：基质金属蛋白酶；金属蛋白酶组织抑制剂；胶原酶；明胶酶；基质酶；膜型基质金属蛋白酶中图分类号：Q51；Q55 文献标识码：AMatrix MetalloproteinasesWU Er-xi1*，WANG Feng-fei1*，Norman McKIE2（1．Department of Human Metabolism and Clinical Biochemistry，University of Sheffield Medical School，Beech Hill Road，Sheffield S10 2RX，UK； 2．Department of Rheumatology，University of Newcastle-upon-Tyne Medical School Framlington Place，Newcastle-upon-Tyne NE2 4HH，UK）Abstract：Matrix metalloproteinases (MMPs) are zinc proteinases that degrade compounds of the extracellular matrix (ECM). These enzymes play a pivotal role in turnover and remodelling of the ECM during organism growth and development and the pathological destruction of tissues in diseases. The activities of metalloproteinases are tightly controlled at several different cellular levels such as modulation by cytokines, growth factors and hormones. MMPs are secreted as zymogens which can be activated by other proteinases such as plasmin or non-proteolytic agents such as organomercurials. All MMPs are inhibited by their natural inhibitors,tissue inhibitors of matrix metalloproteinases(TIMPs). Imbalance between MMPs and TIMPs has been implicated in many diseases such as tumour invasion and metastasis. The synthetic MMP inhibitors such as Marimastat can prevent the growthand further spread of established metastases.Key words：MMPs；TIMPs；collagenases；gelatinases；stromelysins；membrane type MMPsT0　Introduction Matrix metalloproteinases (MMPs) are also called matrixins. Since the first MMP was discovered by Gross and Lapiere in 1962, numerous other MMPs have been described and characterized. To date at least 14 MMPs have been found (Table 1). According to their structural properties and substrate specificities, MMPs can be divided into sub-Table　1　The　matrix　metalloproteinase　family*Enzymes MMP No．**EC No．Mr（kDa）latent/activeExtracellular matrixsubstratesCollagenases Interstitial collagenase MMP1EC 3．4．24．757/48Collagen Ⅰ，Ⅱ，Ⅲ，Ⅶ，Ⅹ；gelatin，entactin，tenascin，aggrecan，progelatinaseA，progelatinase BNeutrophil collagenase MMP8EC3．4．24．3485/65Collagen Ⅰ，Ⅱ，Ⅲ；aggrecanCollagenase 3MMP13　60/48Collagen Ⅰ，ⅡGelatinases Gelatinase A MMP2EC 3．4．24．2472/66Collagen Ⅰ，Ⅳ，Ⅴ，Ⅶ，Ⅹ；gelatin，fibronectin，laminin，aggrecan elastin，progelatinase BGelatinase B MMP9EC 3．4．24．3592/84Collagen Ⅳ，Ⅴ；gelatin，elastin，entactin，aggrecan，vitronectinStromelysins Stromelysin 1MMP3EC 3．4．24．1760/50Collagen Ⅱ，Ⅳ，Ⅸ，Ⅹ，Ⅺ；gelatin，laminin，fibronectin，elastin，tenascin，aggrecan，procollagenase，progelatinase B，neutrophil procollagenaseStromelysin 2MMP10EC 3．4．24．2253/47Collagen Ⅳ，laminin，fibronectin，elastin，aggrecan，procollagenaseStromelysin 3MMP11　65/45Serpins，α1-PI，α2-antiplasmin，insulin-like growth factor-bindingprotein-1Others Matrilysin MMP7EC 3．4．24．2328/21Collagen Ⅳ，gelatins，laminin，fibronectin，entactin，elastin，aggrecan，progelatinaseA，progelatinase B，procollagenaseMetalloelastase MMP12EC．3．4．24．6552/43ElastinMembrane type MMPs MT1-MMP MMP14　63/54Progelatinase A，procollagenase 3，collagen，proteoglycan，fibronectin，tenascinMT2-MMP MMP15　72/61Progelatinase A，procollagenase 3，collagen，proteoglycan，fibronectin，tenascinMT3-MMP MMP16　64/55Progelatinase A MT4-MMP MMP17　70/54Unknown*Compiled from sources including Sang and Douglas[2]；Shingleton et al．[3]；Nagase [4]；Pei et al．[19]；Mari et al．[44]；Murphy et al．[96]；Takino et al．[28，46]andCockett et al．[116]**Some MMPs have been omitted．MMP5 and MMP2 are the same enzyme[5]，MMP4 and MMP6 have been described in only one laboratory and there are no sequence data to date [5]．There are some novel MMPs which have been found recently，see text for details．groups: collagenases, gelatinases, stromelysins, membrane type MMPs (MT-MMP) andothers[1～4]. This review summarises the characteristics, domain structure, substratespecificity, activation mechanisms, regulation, functions of the MMPs.1　Main characteristics of the MMPs All MMPs have a number of common characteristics which are also helpful to identify new members. These properties[2, 5～7]are: 1）they share a common domain structure comprising signal, propeptide, catalytic, and C- terminal hemopexin-like domains (except MMP7) (Fig．1).Fig．1　The domain structure of the matrix metalloproteinases2）They are secreted as zymogens. 3）Their activation can be achieved by other proteinases or organomercurials. 4）Their proteinase activity is blocked by 1,10-phenanthroline and chelating agents. 5）Activation is accompanied by a loss of molecular weight. 6）The active site contains metal ion zinc. 7）Their activity is inhibited by tissue inhibitor of metalloproteinases (TIMPs). 8）The active enzymes cleave one or more components of the extracellular matrix. 9）The enzymes act in neutral pH and need calcium ions for stability.2　Domain structure of MMPs After comparing the primary amino acid sequences of the MMP members, it can be seen that these proteins are divided into several distinct domains that are conserved among family members[1]. The largest MMP member (MMP9) has 7 domains in order from N-terminal: signal, propeptide, catalytic, fibronectin-like,α2V collagen-like, hinge, and C-terminal hemopexin-like. The simplest member MMP7 has only signal, propeptide and catalytic domains. The newly discovered MT-MMPs have a transmembrane domain[8]. All MMPs produce a signal as a leader sequence which cells cleave prior to secretion. The propeptide is lost on activation. For example, the human fibroblast collagenase is synthesized as a preproenzyme of Mr 54 kDa (57 kDa in Table 1) with the signal peptide of19 amino acids[9]. The primary secretion products of human fibroblast collagenase consist ofa minor glycosylated form of Mr 57 kDa and a major unglycosylated polypeptide of Mr 52 kDa[9]. 81 amino acids are removed after proteolytic activation of human fibroblast collagenase[9]. The catalytic domain contains a conserved zinc binding site comprising the sequence HEXGHXXGXXH[2]. The zinc acts as an active site and is ligated by the three-histidine residues of the zinc binding consensus sequence. The glutamic acid residue in the conserved zinc binding site acts as the catalytic base and proton shuttle during proteolysis and is involved in the fixation of a zinc-bound water molecule[10]. The structural integrity of the zinc-binding active site is maintained by a conserved methionine that is called “Met-turn”[10]. The catalytic domain also contains a calcium-binding region where the calcium ion is believed to stabilize the enzyme[11]. With the exception of matrilysin (MMP7), the MMPs contain an additional feature, that is their hemopexin-like or vitronectin-like C-terminal domain[1]. This domain is thought to help determine substrate specificity[1,5]. Thiscould be true, since recently Gohlke et al.[12]have found that the topology and the side chain arrangements of gelatinase A (MMP2) and fibroblast collagenases are very similar, but there are significant differences in surface charge and contouring. They thought these differences may be a factor in allowing the MMP2 C-terminal domain to bind to TIMP2. Very recently, Brooks et al.[13,14]have found that MMP2 can bind αvβ3 through its hemopexin-like domain. Besides the prototype domain structure, the gelatinases contain three tandem domains with sequences similar to the collagen-binding domain of fibronectin. The fibronectin-like domain is thought to be involved in the binding of two gelatinases to their substrate[5]. The MT-MMPs contain a transmembrane domain and a furin recognition site which is also found in stromelysin 3[8,15](Fig．1).3　Substrate specificity of MMPs The collagenases mainly cleave interstitial collagens (type I, II and III), unlike other MMPs, their substrate specificity is well defined. Collagenase action on the α2-macroglobulin results in the cleavage of Gly-Leu Peptide bond[5]．They cleave the Gly-Ile peptide bond of α1(I) chain of collagen and Gly-Leu peptide bond of α2(I) chain of collagen[5]. The substrate cleavage pattern for collagenases is that P1 "residue is invariably hydrophobic (Leu, Ile, Val) and that P1 is usually Gly or a hydrophobic residue[5,16]. The gelatinases cleave denatured collagens and type IV collagen. As shown in table 1, while gelatinase A (MMP2) can also cleave the fibronectin and laminin, major components of the basement membrane, gelatinase B (MMP9) can only cleave the basement membrane component entactin[2]. Stromelysin 1 and 2 have a broad pH optimum and more general activity and are able to degrade many ECM proteins including proteoglycans, gelatins, fibronectin, laminin, elastin, type IV collagen and type IX collagen[17]． Niedzwiecki et al.[18]tested the substrate specificity of the human stromelysin 1 and found the preferences at P3, P2, P1, P1 ", and P2 "are for the hydrophobic amino acids Pro, Leu, Ala, Nva, and Trp, respectively. The mature stromelysin 3 does not degrade any major ECM components. Up to now, the only known substrates for stromelysin 3 are α1-proteinase inhibitor, serine proteinase inhibitors, α2-antiplasmin, and insulin-like growth factor-binding protein-1 [19,20]. However, the substrate specificity overlaps among the MMP members. The gelatinase A (MMP2) can also cleave triple helical type I collagen generating the 3/4 and 1/4 length collagen fragments characteristic of interstitial collagenases[21]. The cleavage site is the same Gly-Ile/Leu bond as the interstitial collagenases[21]. In fact all of the MMPs cleave gelatin and fibronectin at some rate[1]. Almost every MMP degrades an octapeptide containing collagen sequence GPQGIAGQ[5].4　Activation mechanisms of MMPs The MMPs are secreted in a latent form that is subsequently converted into the mature enzymes. The inactive zymogen can be processed into active forms by numerous reagentsincluding proteinases such as trypsin and plasmin; conformational perturbants such as sodium dodecyl sulfate (SDS); heavy metals such as Au (I) compounds and organomercurials; oxidants such as NaSCN; disulfide reagents such as oxidized glutathione; and sulfhydryl alkylating agents such as N-ethylmaleimide[22]. The latent proform of MMPs contains the highly conserved PRCGVNPD sequence with an unpaired cysteine residue. The cysteine residue links to the active site zinc, which blocks the active site. A ‘cysteine switch’　activation mechanism has been proposed[22,23](Fig． 2). In other words, in the latent form of MMPs, the cysteine residue with the thiol group coordinates the catalytically essential zinc ion in the active site of enzyme. Some reagents such as SDS can dissociate the cysteine residue from the zinc ion[22]. From this hypothesis, each MMP should have an unpaired cysteine residue and a zinc-binding site. In fact, all MMPs contain the PRCGVNPD sequence with an unpaired cysteine residue in the propeptide domain and the HEXXHXXGXXH sequence in the catalytic domain. Very recently, it has been shown that he structure of human pro-MMP2 supports this hypothesis[24]. The loops within the propeptide domain act as bait for activating proteinases. The prodomain structure breaks down and its shielding of the catalytic cleft is withdrawn upon cleavage, which allow water to enter and hydrolyze the coordination of the cysteine residue to the zinc ion[24]. All MT-MMPs found to date and stromelysin 3 contain a consensus sequence RXR/KR, which has already been found to be essential in the activation of stromelysin 3 and MT1-MMP by furin [15,25,26]. MMPs also have the ability to activate one another[27]. MT1-MMP[8]and MT3-MMP[28]have been shown to activate MMP2. MMP7 can activate MMP1, MMP2, MMP3 and MMP9[29]. MMP3 activates MMP1[30], MMP8[31], MMP9[32]and MMP13[33]. MMP2 and MT1-MMP activate MMP13[34], while MMP10 activates MMP8[35]. It has also been well documented that MMPs and serine proteinases can act on the proforms of one another [36,37]. Plasmin can cleave the prodomains of MMPs such as collagenase and stromelysin and activate these enzymes[36,37]. MMP7 can catalyze the formation of low molecular weight pro-urokinase and urokinase[38].Fig．2　Cysteine switch mechanism for activation of MMPsProteinases such as trypsin and plasmin cleave the propeptide，ahead of the cysteine to generate intermediate forms．Alternatively，nonproteolytic agents such as aminiphenyl-mercuric acetate （APMA） and sodium dodecy1 sulfate （SDS） will modify the cysteine．In a second step，these intermediate forms can be autoproteolytically cleaved to remove the propeptide and confer permanent activity．5　Regulation of MMPs The activities of metalloproteinases are tightly controlled at several different cellular levels. There are five points: (i) modulation of gene expression by cytokines, growth factors and hormones; (ii) synthesis and secretion of proMMPs; (iii) selective expression of MMP genes in specific tissue/cell types; (iv) activation of proenzymes; and (v) inhibition of the active enzymes[7,36,39,40]. It is known that the MMPs are not constitutively expressed in most tissue types but their mRNAs can be induced by treatment with many kinds of agents such as cytokines, growth factors, hormones, tumour promoter and oncogene products. In some cases the induction of more than one MMP is coordinately regulated, for example, MMP1 and MMP3 are often coordinately expressed[40]. The synthesis of MMP1 and MMP3 can be upregulated by 12-O-tetradecanoyl-phorbol-13-acetate (TPA), interleukin 1 (IL1), tumor necrosis factor α(TNFα), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and platelet derived growth factor (PDGF) and be downregulated by transforming growth factor β　(TGF-β), interferon γ (IFN-γ) retinoic acid and dexamethasone[40], and MMP1 is down regulated by IL4[41]. The selective expression of MMP genes in specific tissue/cell types is one method of regulation of MMPs. Interstitial collagenase (MMP1) arises from connective tissue fibroblasts and macrophages. Neutrophil collagenase (MMP8) is only produced by cells of the neutrophil lineage. Stromelysin 1 (MMP3) is not widely expressed normally, but can be readily induced by growth factors, cytokines, tumour promoters and oncogene products in cultured mesenchymal cells such as chondrocytes and connective tissue fibroblasts. Stromelysin 2 (MMP10) is usually expressed in macrophages, keratinocytes and tumour cells[40,42]. Stromelysin 3 (MMP11) is often expressed in tumour stromal cells[37,40,43,44]. Gelatinase A (MMP2) is the most widespead of all MMPs and is frequently elevated in malignancies as well as occurring in connective tissue cells[36,40]. Gelatinase B (MMP9) is expressed in transformed and tumour-derived cells, neutrophil, corneal epithelial cells, cytotrophoblasts and keratinocytes[36]. Matrilysin is expressed in immature monocytes[36]. Metalloelastase (MMP12) is found in macrophages. MT1-MMP is expressed in tumour stroma cells[45]. MT2-MMP is produced by a human oral malignant melanoma and a human placenta[46]. MT3-MMP is expressed in normal tissues such as lung and kidney and cultured cells such as the squamous cell carcinoma cell line OSC-19 and human embryonal lung fibroblasts[28]. MT4-MMP is expressed in primary breast carcinomas and breast cancer cell lines examined[47]. All MMPs are inhibited by natural inhibitors called tissue inhibitors of metalloproteinases (TIMPs) of which four have been described (Table 2). TIMPs 1, 2 and 4 are secreted extracellularly in soluble form whereas TIMP3 binds to the ECM. All TIMPs share 35%～40% sequence identity and considerable higher structural similarity. The TIMPs bind with high affinity and 1∶1 molar ratio to active MMPs resulting in the loss of proteinase activity. All TIMPs contain 12 cysteine residues that have been shown to form disulfide bonds generating 6 loops. The mechanism of inhibition of MMPs by TIMPs is widely felt to be unclear. Murphy[39]. pointed out that the TIMP C-terminal domain has several different MMP binding sites that act to increase the rate of inhibition. Further studies such as site-directed mutagenesis and the crystal structure research of the complex of MMP and TIMP will facilitate understanding of the mechanism. TIMPs play a pivotal role in the regulation of ECM degradation/remodelling. Disruption of the balance between MMPs and TIMPs has been implicated in many diseases such as tumour invasion and metastasis. Besides their inhibitory activities, the TIMPs play a role in growth-promotion[48]. Some effective synthetic MMP inhibitors also can inhibit MMPs[49,50]. For example, the hydroxamate-based inhibitors such as BB94 (batimastat) and BB2516 (marimastat) can inhibit metastatic spread of tumoursand block the process of tumour neovasculariation[51～54]. The hydroxamate group in thesemolecules can combine with the zinc atom in the active site of MMPs[49,50].Table 2　Comparison of TIMP family members*　TIMP1TIMP2TIMP3TIMP4 Molecular mass28kDa21kDa24kDa22kDa Glycosylation yes no no/yes no Binding properties proMMP9proMMP2ECM proMMP2Transcripts0.9kb 1.1 and 3.5kb 4.5,2.8,2.4and 1.2kb1.4，4.1，2.1，1.2，and 0.97kbModulation of gene expression by TGF-β1updown or noobvious effectup not determinedMajor sites ofexpressionOvary，bone Placenta Kidney，brain Heart Chromosome locationof human genexq1117q2522q12.1～13.22p25 *Compiled from sources including Stetler-Stevenson[117]；Greene et al．[118]；Wick et al．[119]；Chamber and Matrisian[120]and Olson et al．[121]6　MMPs in physiological processes and pathological destruction Normal expression of MMPs is associated with turnover and remodelling of the ECM during growth and development. Cawston[7]summarised the involvement of MMPs in the normal turnover of connective tissue matrix. The MMPs are involved in the normal physiological processes such as ovulation and embryo implantation, embryological development, angiogenesis, bone turnover, uterine resorption and cervical ripening[7]. The MMPs are associated with the pathological destruction of tissues in diseases[1,7]. They are implicated in the processes such as wound healing, corneal ulceration, tumour growth and metastasis, periodontal disease, rheumatoid arthritis, arteriosclerosis and aortic aneurism[1,7]. Considerable research has been directed toward understanding both the steps involved in tumour cell invasion and metastasis and the molecular mechanism of the process. MMPs are thought to be one of the main contributors to tumour invasion and metastasis since they can degrade all of the components of basement membranes which the tumour cellsmust traverse[55～58]. Now the gene-targeting experiments have facilitated the examinationsof the effects that their absence, mutation or overexpression, in various physiological and pathological processes[59]. In the following sections more detailed information is presented on several MMP family members.7　Collagenases I, II and III (MMP1, MMP8 and MMP13) The group of enzymes termed collagenases includes 3 members: MMP1 (interstitialcollagenase or fibroblast collagenase), MMP8 (neutrophil collagenase) and MMP13 (collagenase 3). Their sequences share around 50%[33]. They cleave all three α chains of native types I, II, III collagens at a single site resulting in fragments corresponding to three-quarters and one-quarter of their initial length by hydrolyzing the peptide bond Gly-[Ile orLeu][60～62]. They do not degrade collagen IV and V, which are cleaved by otherproteinases[63～66]. The collagenases can also cleave type X collagen[67]. The threecollagenases have their own preference in cleaving the collagens. The preferred substrate of collagenase 3 (MMP13) is collagen type II[33], fibroblast collagenase (MMP1) preferentially cleaves collagen type III[68]and neutrophil collagenase (MMP8) prefers to cleave type I collagen[69]. Also collagenase 3 has a much stronger gelatinolytic activity than its homologous counterparts MMP1 and MMP8[33]. To date MMP1 expression has not been found in rats or mice[33]. All collagenases are located in the human chromosome 11q22.3 cluster[70,71]and share a highly conserved gene structure[72,73]. The expression of MMP13 in cartilage and its preference to degrade type II collagen suggests that it plays a critical role in the arthritides[74]. X ray crystallographic analyses of MMP1 and MMP8 are available now. The structures show that their catalytic domains harbour two zinc ions and one or twocalcium ions[75～77]. The structure consists of a five-stranded β sheet and three α helices[75].8　Gelatinases A and B (MMP2 and MMP9) Gelatinases, also called type IV collagenases, contain two members: MMP2 and MMP9. They degrade denatured collagens, type IV, V, VII, X, and XII collagens,vitronectin, aggrecan, elastin, galectin 3 and laminin[78～83]. Recently, it has beendemonstrated that MMP2 degrades native interstitial collagens[21]. MMP2 is the most widely distributed MMP[1]. Overexpression of gelatinases has been demonstrated in many tumorsystems and has been linked to tumour invasion[84～87]. While the proMMP2 is often foundcomplexed with the TIMP2 and is activated by MT1-MMP[88,89], the proMMP9 is associated with the TIMP1[78,90]. The regulation of MMP2 and MMP9 gene expression is different[91]. The promoter of MMP9 gene has a TATA-like sequence and a TPA response element, while MMP2 gene has no TATA motif-like sequence in the vicinity of the start site for transcription and TPA response element in its promoter[91]. There is a structural difference between MMP2 and MMP9 that comprises an extended 54 amino acid hinge region sequence which shares some homology with the α2 chain of type V collagen[78]. Brooks et al.[13,14]have found that MMP2 can directly bind integrin αvβ3. They further demonstrated that MMP2 binds αvβ3 through its hemopexin-like domain. It is likely to be distinct from other αvβ3-directed ligands as MMP2 has no RGD sequence[13,14]. Brooks et al.[14]have also found that PEX, a fragment of MMP2, which contains the C-terminal hemopexin-like domain, prevents MMP2 binding to αvβ3 and blocks cells surfacecollagenolytic activity. PEX is likely to be a natural breakdown product of MMP2 since the active form (62 kDa) of MMP2 can be further processed to a smaller species (43 kDa) resulting from autocatalytic removal of the 29　kDa hemopexin-like domain without the presence of TIMP2[14,92]. Both MMP2 and MMP9 are located on human chromosome 16[93].9　Stromelysins 1, 2 and 3 (MMP3, MMP10 and MMP11) MMP3 (stromelysin 1), MMP10 (stromelysin 2) and MMP11 (stromelysin 3) belong to this group. Stromelysins have wide substrate specificity. Stromelysin 1 degrades aggrecan, fibronectin， gelatin， laminin， type II， IV， IX， X， XI collagens and elastin[2,94]. It also participates in activation of other proMMPs such as proMMP1, proMMP8 and proMMP9[31,32,80]. MMP3 is not readily expressed in tissue but can be induced by cytokines such as IL1 and TNFα, growth factors and tumour promoters such as PMA[1,5,95]. Stromelysin 2 cleaves aggrecan, laminin, fibronectin, elastin and type IV[2]. It is transcriptionally active in normal human cells such as keratinocytes and it encodes the secreted Stromelysin 2[42]. Stromelysin 3 is a newly characterized MMP. It can degrade serpin, α1-proteinase inhibitor (α1-PI), α2-antiplasmin, and insulin-like growth factor-binding protein-1[19,20]; the truncated stromelysin 3 can also degrade fibronectin, laminin, aggrecan and type IV[96]. Stromelysin 3 is often found expressed in stromal cells surrounding primary and metastatic carcinomas[43]and may be involved in promoting local tumor development. Recently it has been suggested that the tumor-specific processing of stromelysin 3 to the 35 kDa protein is likely to be an important regulatory mechanism, since the generation of 35 kDa stromelysin in tumour/stroma coculture requires basic fibroblast growth factor (bFGF) and an MMP-like enzyme[44]. Unlike other MMPs, prodomain of stromelysin 3 contains a furin cleavage site, and therefore stromelysin 3 can be processed directly to its 45 kDa active form by furin within the constitutive secretory pathway[15]. MMP3 and MMP10 are located in the human chromosome cluster 11q22.3[70,72], while MMP11 is mapped to the Q11.2 region of chromosome 22[97]．10　Membrane type-matrix metalloproteinases (MMP15, MMP16, MMP17 and MMP18) Membrane type matrix metalloproteinases (MT-MMPs), comprising MT1-MMP, MT2-MMP2, MT3-MMP and MT4-MMP (MMP14, MMP15, MMP16 and MMP17, respectively) are a novel subgroup of MMPs which contain a transmembrane domain and a short cytoplasmic domain in addition to the signal, pro-, catalytic, hemopexin-like and C-terminal domains which are common to other MMPs. To date, four MT-MMPs have been described [8,28,47,98]. From the alignment of amino acid sequences for MT-MMPs[47], it can be seen that they have at least 30% sequence homology to each other. Like other MMPs, they contain cysteine switch sequence, zinc binding site and Met-turn. However, they have an insertion between the propeptide and the catalytic domain in addition to the above mentionedmembrane-binding domain and short cytoplasmic domain. This insertion as in stromelysin 3 contains a potential cleavage site of furin or furin like convertase[8,15]. It has already been demonstrated that furin can cleave this sequence in pro-stromelysin 3 and proMT1-MMP [15,25,26]. However, the proMT1-MMP is proteolytically activated by human plasmin[99]. Okumura et al.[99]proposed that this is an extracellular proteinase activator of proMT1-MMP whereas the intracellular mechanism of activation is mediated by furin or a furin-like enzyme. The second insertion is found in proteinase domain; the function of these 8 amino acids is not known. The third insertion which contains the hydrophobic transmembrane domain is located in the C-terminal. The transmembrane domain of MT-MMPs plays an essential role in the progelatinase A (proMMP2) activation function of MT-MMPs, although some portions of the truncated form remain on the cell surface after the removal of this domain[100]. In MT1-MMP, the activation complex is a trimer comprising MT1-MMP, progelatinase A and TIMP2[88,89]. A recent study has suggested that MT1-MMP is a TIMP2 receptor[101]. This is further confirmed by another recent study[73]that TIMP2 and MT1-MMP form a complex for activation of progelatinase A. TIMP2 and TIMP3 can efficiently inhibit MT1-MMP whereas TIMP1 is a poor inhibitor for MT1-MMP[102]. Will et al.[102] also demonstrated that TIMP2 and TIMP3 can bind more rapidly to the catalytic domain of MT1-MMP than to the catalytic domain of MMP2. At the same time, two groups showed that MMP14, MMP15 and MMP16 locate in chromosomes 14, 16 and 8, respectively [103,104]. The MT-MMP gene loci are dispersed whereas most MMPs are clustered on chromosome 11q22.3[70,71], suggesting that the MT-MMPs may be a genetically distinct subgroup of MMPs. It has been suggested that the MT-MMPs play a critical role in tumor cell invasion and in ECM degradation[8,57,102]. MT-MMPs also can degrade ECM components[25,105,106].11　Others This group includes MMPs such as matrilysin (MMP7) and metalloelastase (MMP12). Matrilysin, also called putative metalloproteinase 1 (pump 1), is the smallest member of the MMP family. Unlike other MMPs, it lacks a C-terminal hemopexin-like domain. MMP7 can degrade various ECM components. It also can activate other proMMPs such as proMMP1, 3, 2 and 9[29]. Metalloelastase (MMP12) is also called macrophage elastase (ME). The expression of human macrophage elastase (HME) is mainly restricted to tissue macrophages [107]. Besides the substrate elastin listed in Table 1, the purified recombinant HME can degrade fibronectin, laminin, entactin, type IV collagen, chondroinan sulfate, and heparan sulfate[108]. Chandler et al.[109]also demonstrated that HME can degrade myelin basic protein and processes a TNF α fusion protein. Therefore Gronski et al.[108]suggested that HME may be essential for macrophages to penetrate basement membranes and remodel injured tissue during inflammation. MMP7 and MMP12 are allocated in the human chromosome cluster 11q22.3[70,72,107].。

MMP-9，也被称为基质金属蛋白酶9，或者是92 kDa IV型胶原酶、92 kDa 明胶酶、明胶酶B，属于锌-金属蛋白酶家族。

MMP-9基因位于染色体20q11.1~13.1，具有13个外显子和9个内含子。

它的主要功能是降解和重塑细胞外基质的动态平衡，并参与许多正常生理过程，如胚胎发育、繁殖、血管形成、骨骼发育、伤口愈合、细胞迁移、学习和记忆等。

此外，MMP-9也在一些病理过程中发挥重要作用，如中性粒细胞迁移通过基底膜的调节因子，血管和新生血管形成等。

在人类呼吸道上皮愈合过程中，MMP-9的水平会显著上调，同时它也与许多其他病理过程相关。

基质金属蛋白酶（ＭＭＰｓ）与冠心病【关键词】基质金属蛋白酶冠心病基质金属蛋白酶(MMPs)是一组分解细胞外基质的内源性蛋白水解酶的总称,是目前倍受注目的性症因子之一。

越来越多的证据表明MMPs参与了肿瘤增殖、侵袭和转移，心脏和血管结构和功能的改变、哮喘和慢阻肺的发生、发展、伤口愈合，胎儿发育等多个层面的病理过程。

MMPs降解外基质（ECM）和基底膜的作用是主要病理生理机制所在。

目前认为ECM与冠心病的发生、发展密切相关，故目前MMPs与冠心病的关系逐渐成为心血管病研究领域的热点。

1 MMPs生物学特性所有的MMPs家族成员都具备以下基本特征：①降解ECM；②以酶原方式分泌，被纤溶酶和细胞因子激活和修饰后发挥生物活性；③活性依赖与锌离子；④均需要钙离子保持稳定性；⑤在中性PH值下发挥作用；⑥在体内存在着天然的激活剂和抑制剂。

MMPs根据底物特异性和结构差异分为四大类［1］。

第一类为胶原酶，它们的主要水解底物是纤维类胶原，即Ⅰ、Ⅱ、Ⅲ型胶原；第二类为明胶酶，主要水解变性胶原及基膜的主要成分Ⅳ型胶原；第三类为基质水解酶类，MMPs3和MMPs10的水解底物比较广泛，如Ⅲ、Ⅳ型胶原，蛋白聚糖，明胶及糖蛋白等；第四类为模型MMPs，能降解几种ECM外，还能激活其他MMPs。

MMPs的调节：MMPs活性有三种调控方式。

①转录水平调节；大多数MMPs 在正常组织中表达一般保持在较低水平，在出现一定病理生理变化时可能升高。

炎性因子、激素、生长因子都有一定影响。

②酶原的激活：MMP以无活性的酶原形式分泌到细胞外，须经过蛋白酶的水解才能激活。

目前已经发现的激活机制有：逐步激活、通过MT-MMP激活、细胞内激活。

MMPS的最初激活往往涉及纤溶酶、胰蛋白酶、弹性蛋白酶、激胎释放酶等，其中纤溶酶被活认为是体内最强的生理激活剂。

③内源性金属蛋白酶组织抑制剂（TIMPS）的调节作用：TIMPS 是体内MMPs活性的重要内源性抑制剂，是一种低分子量蛋白。

扶正化淤方对四氯化碳肝纤维化大鼠基膜型基质金属蛋白酶活性的影响（作者:___________单位: ___________邮编: ___________）作者：崔红燕，刘成海，孙保木，刘成【摘要】目的探讨扶正化淤方通过影响基膜型基质金属蛋白酶活性而抗肝纤维化的作用机制。

方法四氯化碳(CCl4)皮下注射与高脂低蛋白饲料复合建立大鼠肝纤维化模型。

随机分为正常组、模型组与扶正化淤方药物干预组。

药物组自造模之日起以扶正化淤方稀释液灌胃，共用药6周。

HE染色观察肝组织炎性病理变化，天狼猩红染色观察肝组织胶原沉积，盐酸水解法测定肝组织羟脯氨酸含量，Western 印迹法分析肝组织α-SMA、Ⅳ型胶原、MMP-2、MMP-9、TIMP-2和MT-MMP1蛋白表达水平；明胶酶图法检测肝组织MMP-2/9活性。

结果与正常大鼠比较，模型大鼠血清肝功能异常、肝组织炎症明显、肝组织IV型胶原表达与沉积增加，α-SMA、Ⅳ型胶原、MMP-2、MMP-9、TIMP-2和MT-MMP1蛋白表达升高，而MMP-2/9活性水平明显上升。

扶正化瘀方显著改善模型大鼠血清肝功能、减轻肝脏炎症和胶原沉积；减少肝星状细胞活化；抑制Ⅳ型胶原蛋白表达和沉积；降低MMP-2、TIMP-2和MT1-MMP蛋白表达；抑制MMP-2/9活性。

结论扶正化瘀方可通过抑制MMP-2的激活与活性水平，并减少Ⅳ型胶原沉积，而起到减轻肝组织的破坏与重构而发挥抗肝纤维化的作用。

【关键词】扶正化淤方基质金属蛋白酶基膜型肝纤维化Abstract：ObjectiveTo investigate the effects of Fuzheng Huayu recipe (FZHY recipe) on the activity of membrane-type matrix-metalloproteinases and its mechanism in the treatment of liver fibrosis. MethodsLiver fibrosis was induced in rats by injection of CCl4 subcutaneously and fed with high lipid and lower protein diet．Rats were randomly divided into 3 groups：normal，model control and Fuzheng Huayu decoction treated group. For the treated group，rats were administered with Fuzheng Huayu decoction by gavage for 6 weeks. Inflammation in liver was determined by HE staining,Collagen deposition in liver was studied by Sirius red staining,the content of hepatic hydroxyproline(Hyp) were measured with Jamall’s methods; protein expressions of α-SMA, collagen IV，MMP-2, MMP-9 and TIMP-2 were tested by Western blot, and MMP-2/9 activities were analyzed by gelatin zymography.ResultsCompared with normal group，abnormal liver function, obvious hepatic inflammation, excess collagen deposition, high protein expressions of collagen IV，MMP-2/9，TIMP-2 andMT1-MMP, and increased activities of MMP-2/9 were observed in model group. Compared with model group, FZHY recipe significantly improved the rat liver function, alleviated liver inflammation and the excess collagen deposition.In addition, it also significantly down-regulated the protein expression of collagen IV , MMP-2, TIMP-2 and MT1-MMP, and inhibited the activity of MMP-2. ConclusionFuzheng Huayu decoction can down-regulate the activities of MMP-2 /9 and inhibit the activation of MMP-2，reduce the excess collagen IV deposition in liver tissue, and to alleviate the damage and regeneration of liver.Key words：FZHY Decoction；MMP；Membrane-type；Liver Fibrosis肝纤维化是诸多慢性肝病的共同病理特点，以肝脏细胞外基质(extracellular matrix，ECM)代谢失衡，生成超过降解，并大量沉积于肝组织为特征。

金属基质蛋白酶9与疾病的研究进展摘要：金属基质蛋白酶（MMPs）是一组可以选择性降解细胞外基质（ECM）的内肽酶，金属基质蛋白酶9（MMP-9）又名明胶酶B，它是MMPs家族中一个重要成员，能水解弹性蛋白、胶原蛋白、明胶、纤维蛋白等多种细胞外基质成分，在生理情况下参与人体的正常发育，是ECM降解的生理性调节因子。

所有MMPs都受金属基质蛋白酶抑制剂所抑制，两者的不平衡导致许多疾病的发生。

金属基质蛋白酶抑制剂1（tissue inhibitor of metalloproteinase-1，TIMP-1）为MMP-9特异性抑制剂，目前认为MMP-9与TIMP-1是调节ECM降解、合成的主要酶类，MMP-9/TIMP-1比值失衡是多种疾病发生、发展的重要机制之一。

近年研究发现MMP-9还与全身多系统疾病的发生、发展有关，现就MMP-9与多种疾病的相关研究进展进行综述。

关键词：金属基质蛋白酶9；疾病；研究进展金属基质蛋白酶（matrix metalloteinases，MMPs）是一组锌离子依赖的肽链内肽酶，因含金属离子（锌、钙）而得名，现至少已发现26种MMPs，称为MMPs家族，是构成细胞外基质降解最重要的蛋白水解系统。

其抑制剂（tissue inhibitor of metalloproteinases，TIMPs）是调节MMPs的重要因素。

MMPs与TIMPs在维持ECM的动态平衡中起重要作用。

细胞外基质（extracellullar matrix，ECM）是一类由胶原、蛋白聚糖及糖蛋白等大分子物质组成的动态网状结构，ECM不仅起细胞与细胞之间机械支持和连接作用，也是细胞与细胞之间信号传递的桥梁，现已识别丝氨酸蛋白酶、半胱氨酸蛋白酶、天冬氨酸蛋白酶、基质金属蛋白酶均能降解ECM[1]。

金属基质蛋白酶9（matrix metalloteinases-9，MMP-9）是MMPs家族分子量最大的明胶酶。

MMP-2、MMP-14、TIMP-2在胃癌组织中的表达及意义张金玲;费雁;陈伟;冯刚【摘要】Objective To examine the expression of MMP-2, MMP-14 and TIMP-2 in gastric cancer and its clinical significance. Methods The expression of MMP-2,MMP-14 and TIMP-2 was immunohistochemically detected in 80 samples of gastric cancer tissues and 30 adjacent normal gastric tissues. Results The positive expression rate of MMP-2 and MMP-14 was 75.00% and 82. 50% respectively,in gastric cancertissues,significantly higher than that in normal gastric tissues(36. 67% and 33. 33% ,respectively) (P<0. 01). The positive expression rate of TIMP-2 was 45. 00% and 40. 00% in gastric cancer tissues and normal gastric tissues with no significant difference being found. The expression of MMP-2 and MMP-14 in gastric cancer tissues was closely correlated to the cell differentiation,invasion,lymph node metastasis and TNM stages(P<0. 01). But the expression of TIMP-2 in gastric cancer tissues was only related to the lymph node metastasis (P<0. 05). The expression levels of MMP-2 and MMP-14 were significantly higher in survival time <2 years group than in survival time ≥2 years group(P<0. 01). Conclusion MMP-2 and MMP-14are over-expressed in gastric cancer. Combined detection for MMP-2/MMP-14 or MMP-2/MMP-14/TIMP-2 may help evaluate the malignancy of gastric cancer and predict the prognosis of patients with gastric cancer.%目的探讨基质金属蛋白酶(matrix metalloproteinase,MMP)-2、14及基质金属蛋白酶组织抑制因子-2(tissue inhibitor of metalloproteinase,TIMP-2)在胃癌中的表达及其意义.方法用免疫组织化学方法 (SP法)检测80例胃癌手术患者胃癌组织及30例同期癌旁≥5 cm正常组织中MMP-2、MMP-14及TIMP-2的表达.结果①在胃癌、正常胃组织中,MMP-2阳性表达率分别为75.00%和36.67%,MMP-14阳性表达率分别为82.50%和33.33%,差异有统计学意义(均P＜0.01);TIMP-2阳性表达率分别为45.00%和40.00%,差异无统计学意义.②癌组织中,MMP-2、MMP-14的表达与肿瘤分化程度、浸润深度、淋巴结有无转移和TNM临床分期密切相关(均P＜0.01).TIMP-2的表达仅与淋巴结有无转移相关(P＜0.05).③生存期＜2年患者的MMP-2、MMP-14表达率明显高于生存期≥2年患者(均P＜0.01).结论胃癌组织中,MMP-2、MMP-14高表达,联合检测MMP-2、MMP-14或MMP-2、MMP-14、TIMP-2可作为胃癌恶性程度及预后判断的指标.【期刊名称】《华中科技大学学报（医学版）》【年(卷),期】2013(042)002【总页数】4页(P227-230)【关键词】胃癌;基质金属蛋白酶-2;基质金属蛋白酶-14;基质金属蛋白酶组织抑制因子-2【作者】张金玲;费雁;陈伟;冯刚【作者单位】华中科技大学同济医学院附属普爱医院肿瘤科,武汉,430033【正文语种】中文【中图分类】R735.2胃癌是我国最常见的恶性肿瘤之一，年死亡率高，其确切病因尚未完全明了。

基质金属蛋白酶,分类基质金属蛋白酶，听起来就像是科学界里的一群超级英雄，专门对付那些让人头疼的细胞外基质。

想象一下，咱们的身体里面，就像是一个错综复杂的城市，血管、神经、细胞，啥都有，它们得靠一种叫做“细胞外基质”的东西来保持秩序，就像是城市里的道路、桥梁和建筑。

而基质金属蛋白酶呢，就是那些能够“拆迁”和“改造”这些基础设施的特种部队。

这些家伙可不是吃素的，它们有着强大的分解能力，能把那些老旧的、不再需要的细胞外基质给拆掉，让新的、更有活力的结构能够长出来。

就像是城市里的拆迁队，虽然看起来是在搞破坏，但实际上是在为城市的发展腾出空间，让新的建筑能够拔地而起。

基质金属蛋白酶这个大家族里，成员可不少，每个成员都有自己的特长和喜好。

有的擅长对付胶原蛋白，那可是构成皮肤、骨骼和血管的重要成分；有的则喜欢分解弹性蛋白，让咱们的皮肤能够保持弹性，不容易长皱纹；还有的专门对付那些让细胞黏在一起的物质，就像是城市里的“胶水”，让细胞们能够紧紧地团结在一起。

这些基质金属蛋白酶，平时都乖乖地待在自己的岗位上，不会随便乱动。

但是，一旦身体里面出现了什么异常，比如炎症、癌症啥的，它们就会像是被激活了一样，开始疯狂地工作起来。

就像是城市里的消防员，平时默默无闻，但一旦有火情发生，就会立刻冲上前去，扑灭大火。

不过，话说回来，这些基质金属蛋白酶要是太过头了，也会带来麻烦。

就像是拆迁队拆得太猛了，把好好的房子也给拆了，那可就不妙了。

如果它们过度活跃，就会破坏掉那些原本应该保留的细胞外基质，导致身体出现问题。

比如，皮肤上的伤口久久不能愈合，骨骼变得脆弱容易骨折，甚至还会出现肿瘤。

所以，咱们的身体里，还得有一套机制来管理这些基质金属蛋白酶，让它们既能够发挥自己的作用，又不会过度破坏。

就像是城市里的规划局，得时刻关注着拆迁队的动向，确保他们的工作既合法又合规。

科学家们对基质金属蛋白酶的研究，也是越来越深入了。

他们就像是侦探一样，不断地寻找着这些特种部队的踪迹，了解它们的习性，以便更好地利用它们，为人类的健康服务。

基质金属酶介绍基质金属酶（Matrix Metalloproteinases，MMPs）是一类能够降解基质蛋白分子的酶，在生物体中广泛存在。

这一类酶在许多生理和病理过程中发挥重要作用，包括胚胎发育、组织修复和炎症反应等。

本文将全面探讨基质金属酶的结构、功能以及其在不同疾病中的作用。

一、基质金属酶的结构基质金属酶是一类含有金属离子的蛋白酶，主要包括九个家族，分别是MMP-1至MMP-9。

这些家族中的每个成员都具有相似的结构，包括一个信号肽、一个前体区、一个附着的结构域以及一个C末端的催化结构域。

催化结构域中含有活性位点，可结合底物并参与酶解反应。

二、基质金属酶的功能基质金属酶主要通过降解基质蛋白分子来调节细胞外基质的合成和降解。

基质蛋白分子在维持细胞外基质结构和功能中起重要作用，但过度积累会导致疾病的发生。

基质金属酶的主要功能包括： 1. 降解胶原蛋白：基质金属酶可以降解胶原蛋白，维持胶原蛋白的动态平衡。

2. 降解纤维连接蛋白：基质金属酶可以降解纤维连接蛋白，参与组织修复和创伤愈合过程。

3. 调节细胞迁移：基质金属酶通过调节细胞外基质的降解和重建，影响细胞迁移的速度和方向性。

4. 参与血管生成：基质金属酶在血管生成过程中发挥重要作用，调控内皮细胞的迁移和管腔形成。

三、基质金属酶在疾病中的作用基质金属酶的异常表达和活性会导致多种疾病的发生和发展。

以下是基质金属酶在不同疾病中的作用： ### 1. 炎症性疾病基质金属酶参与调节炎症反应过程，当炎症失控时，基质金属酶的过度活化会导致组织损伤和病变。

2. 癌症转移基质金属酶在肿瘤转移过程中起关键作用。

它能够降解基质蛋白分子，破坏细胞外基质屏障，促进肿瘤细胞的侵袭和迁移。

3. 心血管疾病基质金属酶参与动脉粥样硬化病变的形成，它能够降解血管壁中的胶原蛋白，导致血管壁的脆性增加，从而促进斑块的破裂和血栓形成。

4. 关节疾病在关节炎和骨质疏松等关节疾病中，基质金属酶过度激活导致关节软骨的破坏和损伤，加速疾病的进展。