运动负荷对骨基质有机成分的影响及其可能机制

骨基质的有机成分主要为骨Ⅰ型胶原基质金属蛋白酶,该酶是细胞外基质降解的重要酶类;基质金属蛋白酶抑制因子则是基质金属蛋白酶活性的抑制剂,它们均为骨代谢过程中的重要标志性物质。本文通过查阅文献资料,对生理、部分病理状态及运动干预条件下,骨基质的Ⅰ型胶原和基质金属蛋白酶及其抑制因子的变化情况进行综述,并对其变化的机制予以阐释。     

缺氧对肺动脉内皮细胞MMP-2、MMP-9表达的影响

目的观察缺氧对培养的猪肺动脉内皮细胞基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)的表达的影响.方法采用RT-PCR、Western blot、底物胶电泳(酶谱图)、免疫细胞化学等方法.结果肺动脉内皮细胞缺氧24h可使MMP-2的mRNA表达和蛋白分泌减少,酶活性减弱,与常氧组比较有显著性差异, MMP-9无明显变化.结论缺氧时MMP-2降低,溶解细胞外基质的能力降低,可能是缺氧性肺血管构型重组的机制之一.     

MMP-2及MMP-9与主动脉疾病相关性的研究进展

基质金属蛋白酶是一类可降解细胞外基质的蛋白酶,基质金属蛋白酶-2和-9为明胶酶,可降解细胞外基质中的胶原蛋白及弹性蛋白,其动态平衡对维持细胞外基质的稳定具有重要意义。主动脉的细胞外基质是主动脉中层重要的组成部分,细胞外基质成分的改变可导致主动脉中层结构的损伤,在主动脉疾病的发生、发展过程中起着重要作用。主动脉基质金属蛋白酶-2和-9的表达失衡可引起主动脉中层细胞外基质的降解,导致主动脉中层结构的损伤,从而促进主动脉疾病的发生。同时,主动脉疾病也可导致血浆中MMP-2、MMP-9浓度的升高。本文对近年来基质金属蛋白酶与主动脉疾病相关性的研究及进展作一综述,为心血管疾病发生机制的研究和治疗提供文献依据。     

基质金属蛋白酶与心肌重塑

细胞外基质参与和促进了心肌重塑的过程,基质金属蛋白酶是调节细胞外基质重要的酶,基质金属蛋白酶在心肌重塑过程表达变化可分为三个时相,其活性受到信号传导途径、炎症因子和活性氧/活性氮的调节,基质金属蛋白酶可能作为心肌梗塞等疾病治疗的靶标     

口腔角质形成细胞与成纤维细胞共培养对胶原代谢的影响

目的:了解口腔角质形成细胞与成纤维细胞共同培养对胶原代谢的影响.方法:对口腔粘膜角质形成细胞和成纤维细胞进行了共培养,检测其胶原分泌水平、基质金属蛋白酶的含量及活性、基质金属蛋白酶抑制剂的水平.结果:只有共同培养组才能检测到MMP-9;共同培养组的活化型MMP-2水平与单独培养组相比无显著差异.角质形成细胞与成纤维细胞共同培养组的TIMP-1水平及胶原水平均明显高于单独培养组.结论:口腔角质形成细胞与成纤维细胞共同培养可能主要通过改变基质金属蛋白酶抑制剂的水平而影响胶原代谢.     

基质金属蛋白酶-9及其抑制剂mRNA在吸烟大鼠肺组织中的表达

目的 研究吸烟大鼠肺组织和肺泡巨噬细胞中基质金属蛋白酶9(MMP-9)和金属蛋白酶组织抑制剂-1(TIMP-1)基因的表达,探讨其在细胞外基质重塑中的作用。方法 建立吸烟大鼠模型,随机分为对照组和吸烟1、2、3、4、5及6月组(每组10只),用原位杂交技术检测肺组织和肺泡巨噬细胞MMP-9和TIMP-1 mRNA表达,用免疫组织化学技术观察Ⅳ型胶原在肺内的表达。结果 吸烟组肺组织和肺泡巨噬细胞MMP-9 mRNA的表达逐渐上升,至吸烟6月时均达高峰;而TIMP-1 mRNA的表达渐上升,至吸烟3～4月时达高峰,后逐步下降;肺组织Ⅳ型胶原的表达在吸烟3月时达高峰,然后渐降。结论 MMP-9／TIMP-1的动态平衡在吸烟大鼠肺气肿模型肺组织的细胞外基质重塑中有重要作用。     

牛黄对小鼠口腔成纤维细胞功能的调节作用

探讨牛黄对原代小鼠口腔成纤维细胞功能的影响,揭示其在溃疡愈合过程中的作用及机制。本试验采用MTT法、氯胺-T法、明胶酶谱分析和酶联免疫反应测定了牛黄对小鼠口腔成纤维细胞增殖、胶原沉积、金属蛋白酶-2、-9活性和基质金属蛋白酶抑制因子-1合成的影响。结果表明牛黄能显著抑制小鼠口腔成纤维细胞的增殖、胶原沉积和金属蛋白酶-2活性,同时也极显著(P<0.01)抑制基质金属蛋白酶抑制因子-1的产生。结果提示牛黄在溃疡愈合过程中不具生肌作用,可能通过抗炎促进溃疡愈合;其抑制胶原合成的机制可能与极显著抑制基质金属蛋白酶抑制因子-1有关。     

血小板、基质金属蛋白酶与肿瘤转移

血小板除参与正常的止血过程外还具有很多病理和生理作用。血小板活化后可以分泌基质金属蛋白酶（matrix metalloproteinases,MMPs）。MMPs属于Zn^2＋和Ca^2＋依赖的内肽酶家族,能特异性与细胞外基质成分相结合并降解细胞外基质。MMPs降解基底膜中的主要成分Ⅳ型胶原,是肿瘤转移发生必不可少的关键步骤。血小板能够与肿瘤细胞结合并促进肿瘤转移,而MMPs在血小板促进肿瘤转移过程中发挥了重要的作用。     

人基质金属蛋白酶-2的表达、纯化和性质鉴定

PCR方法扩增人基质金属蛋白酶 2 (MMP 2 )不含信号肽的表达序列 ,酶切和测序鉴定正确后 ,构建酵母重组表达质粒pPIC9 MMP 2 ,电击法转化毕赤酵母 (Pichiapastoris)细胞得到阳性克隆 ,甲醇诱导获得含大量基质金属蛋白酶 2的培养上清 ,经SephacrylS 2 0 0纯化后 ,纯度达到电泳纯。明胶酶谱和SDS PAGE分析说明重组MMP 2能够降解明胶和IV型胶原 ,表明重组蛋白具有与天然MMP 2相似的底物特异性。糖基化分析和SDS PAGE表明 ,表达产物的分子量约为 5 0kD ,重组MMP 2的C 末段可能发生了降解。     

基质金属蛋白酶及其组织抑制剂研究进展

基质金属蛋白酶家族是细胞外基质降解过程中的重要酶类,组织金属蛋白酶抑制剂是基质金属蛋白酶的天然抑制物。研究证实,细胞外基质中基质金属蛋白酶及其组织抑制剂的失衡与多种病理机制有关,尤其与肿瘤的侵袭和转移密切相关。本就基质金属蛋白酶及其组织抑制剂的性质、结构以及功能进行了综述。     

Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-3 are key regulators of extracellular matrix degradation by mouse embryos

Embryo implantation in humans and rodents is a highly invasive yet tightly controlled process involving extracellular matrix (ECM) degradation. Matrix metalloproteinase 9 (MMP-9) has been implicated as the major facilitator of this ECM degradation. MMP-9 is expressed by the embryo's trophoblast cells, whereas tissue inhibitor of metalloproteinases 3 (TIMP-3) is expressed by the maternal uterine cells immediately adjacent to the trophoblast. We examined the functional roles of MMP-9 and TIMP-3 during in vitro ECM degradation by mouse embryos. Blastocysts were treated with either MMP-9 antisense or sense oligonucleotides and incubated on an ECM gel. The extent of ECM degradation exhibited by the blastocysts due to proteinase secretion was quantified. Embryos exposed to MMP-9 antisense oligonucleotides exhibited reduced ECM-degrading activity as compared with controls, and this reduced activity was correlated with the level of MMP-9 secreted by the embryos. The functional role of TIMP-3 was then examined by incubating blastocysts on an ECM gel that had been impregnated with various amounts of TIMP-3. In a dose-dependent manner, increases in TIMP-3 resulted in a reduction in ECM degradation and were correlated with diminished MMP-9 activity. These results provide important functional evidence that in vitro ECM degradation is regulated by embryo-derived MMP-9 and ECM-derived TIMP-3.     

Matrix metalloproteinase expression and activity in trophoblast-decidual tissues at organogenesis in CF-1 mouse

During early placentation, matrix metalloproteinases (MMPs) play important roles in decidualization, trophoblast migration, invasion, angiogenesis, vascularization and extracellular matrix (ECM) remodeling of the endometrium. The aim of our study was to analyze the localization, distribution and differential expression of MMP-2 and -9 in the organogenic implantation site and to evaluate in vivo and in vitro decidual MMP-2 and -9 activities on day 10 of gestation in CF-1 mouse. Whole extracts for Western blotting of organogenic E10-decidua expressed MMP-2 and -9 isoforms. MMP-2 immunoreactivity was found in a granular and discrete pattern in ECM of mesometrial decidua (MD) near maternal blood vessels and slightly in non-decidualized endometrium (NDE). Immunoexpression of MMP-9 was also detected in NDE, in cytoplasm of decidual cells and ECM of vascular MD, in trophoblastic area and in growing antimesometrial deciduum. Gelatin zymography showed that MMP-9 activity was significantly lower in CM compared to the active form of direct (not cultured) and cultured decidua. The decidual active MMP-9 was significantly higher than the active MMP-2. These results show differential localization, protein expression and enzymatic activation of MMPs, suggesting specific roles for MMP-2 and MMP-9 in decidual and trophoblast tissues related to organogenic ECM remodeling and vascularization during early establishment of mouse placentation.     

PAI-1 promotes extracellular matrix deposition in the airways of a murine asthma model

Dysregulation of matrix metalloproteinases (MMPs) and ineffective fibrinolysis are associated with the deposition of extracellular matrix (ECM). We hypothesized that elevated plasminogen activator inhibitor (PAI)-1 promotes ECM deposition in the asthmatic airway by inhibiting MMP-9 activity and fibrinolysis. Degree of airway inflammation was similar in PAI-1(-/-) and wild type (WT) mice after ovalbumin (OVA) challenge. PAI-1 production, deposition of collagen and fibrin, and MMP-9 activity in the lung tissue or airways were greater after OVA challenge compared with saline challenge. However, in PAI-1(-/-) mice, collagen deposition was 2-fold less, fibrin deposition was 4-fold less, and MMP-9 activity was 3-fold higher. This is the first direct evidence that the plasmin system regulates ECM deposition in the airways of a murine asthma model, independently of the effect of PAI-1 on inflammatory cells. The results suggest that the PAI-1-dependent inhibition of MMP-9 activity and fibrinolysis is a major mechanism by which ECM deposition occurs.     

Airway epithelial cell migration dynamics. MMP-9 role in cell-extracellular matrix remodeling.

Cell spreading and migration associated with the expression of the 92-kD gelatinase (matrix metalloproteinase 9 or MMP-9) are important mechanisms involved in the repair of the respiratory epithelium. We investigated the location of MMP-9 and its potential role in migrating human bronchial epithelial cells (HBEC). In vivo and in vitro, MMP-9 accumulated in migrating HBEC located at the leading edge of a wound and MMP-9 expression paralleled cell migration speed. MMP-9 accumulated through an actin-dependent pathway in the advancing lamellipodia of migrating cells and was subsequently found active in the extracellular matrix (ECM). Lamellipodia became anchored through primordial contacts established with type IV collagen. MMP-9 became amassed behind collagen IV where there were fewer cell-ECM contacts. Both collagen IV and MMP-9 were involved in cell migration because when cell-collagen IV interaction was blocked, cells spread slightly but did not migrate; and when MMP-9 activation was prevented, cells remained fixed on primordial contacts and did not advance at all. These observations suggest that MMP-9 controls the migration of repairing HBEC by remodeling the provisional ECM implicated in primordial contacts.     

Modulatory effect of interleukin-1α on expression of structural matrix proteins,MMPs and TIMPs in human cardiac myofibroblasts: Role of p38 MAP kinase

The proinflammatory cytokine interleukin-1 (IL-1) elicits catabolic effects on the myocardial extracellular matrix (ECM) early after myocardial infarction but there is little understanding of its direct effects on cardiac myofibroblasts (CMF), or the role of p38 mitogen-activated protein kinase (MAPK). We used a focused RT-PCR microarray to investigate the effects of IL-1α on expression of 41 ECM genes in CMF cultured from different patients, and explored regulation by p38 MAPK.IL-1α (10 ng/ml, 6 h) had minimal effect on mRNA expression of structural ECM proteins, including collagens, laminins, fibronectin and vitronectin. However, it induced marked increases in expression of specific ECM proteases, including matrix metalloproteinases MMP-1 (collagenase-1), MMP-3 (stromelysin-1), MMP-9 (gelatinase-B) and MMP-10 (stromelysin-2). Conversely, IL-1α reduced mRNA and protein expression of ADAMTS1, a metalloproteinase that suppresses neovascularization. IL-1α increased expression of TIMP-1 slightly, but not TIMP-2. Data for MMP-1, MMP-2, MMP-3, MMP-9, MMP-10 and ADAMTS1 were confirmed by quantitative real-time RT-PCR. Tumor necrosis factor-alpha (TNFα), another important myocardial proinflammatory cytokine, did not alter expression of these metalloproteinases. IL-1α strongly activated the p38 MAPK pathway in human CMF. Pharmacological inhibitors of p38-α/β (SB203580) or p38-α/β/γ/δ (BIRB-0796) reduced MMP-3 and ADAMTS1 mRNA expression, but neither inhibitor affected MMP-9 levels. MMP-1 and MMP-10 expression were inhibited by BIRB-0796 but not SB203580, suggesting roles for p38-γ/δ.In summary, IL-1α induces a distinct pattern of ECM protein and protease expression in human CMF, in part regulated by distinct p38 MAPK subtypes, affirming the key role of IL-1α and CMF in post-infarction cardiac remodeling.     

脓毒症患儿内生吗啡动态变化及临床意义

目的:检测脓毒症患儿血清内生吗啡(EM)动态变化,探讨脓毒症患儿血清EM水平的临床意义。方法:选择2017年6月-2018年2月华中科技大学同济医学院附属同济医院收治的30例伴有严重脓毒症或脓毒症休克的患儿作为脓毒症组,30例全身炎症反应综合征(SIRS)患儿作为SIRS组,30例健康儿童作为对照组。脓毒症组、SIRS组及对照组分别于入组后第1、3、6、9天抽取外周静脉血,采用免疫荧光法检测血清降钙素原(PCT)水平,采用酶联免疫吸附试验(ELISA)测定血清EM水平,采用流式细胞术(FCM)检测细胞免疫功能。结果:SIRS组、脓毒症组患儿的第1、3、6天血清PCT水平显著高于对照组,且随时间延长PCT水平逐渐降低,至第9天降至正常水平(P0.05),而脓毒症组与SIRS组间血清PCT水平无统计学差异(P0.05)。脓毒症组第1、3、6天血清EM水平均高于SIRS组(P0.05),第9天两组血清EM水平比较差异无统计学意义(P0.05)。脓毒症组EM水平随时间延长而降低,至第9天降至SIRS组的水平(P0.05)。与SIRS组相比,第1天脓毒症组的CD3~+T细胞数量增多(P0.05),两组CD4~+、CD8~+T细胞数量、CD4~+/CD8~+比例比较无统计学差异(P0.05)。结论:脓毒症患儿中血清EM水平较高,有可能作为诊断脓毒症特异性较高的生物标志物。     

灯盏细辛口服液对急性冠状动脉综合征患者外周血C-反应蛋白、MCP-1 和MMP-9 水平的影响

目的:探讨灯盏细辛口服液对急性冠状动脉综合征患者外周血C-反应蛋白、1-甲基环丙烯和9-基质金属蛋白酶水平的影响。方法:选取我院心内科已确诊为急性冠状动脉综合征的患者110例,随机分为实验组和对照组,对照组行常规药物治疗,实验组在对照组的基础上口服灯盏细辛口服液。比较两组患者治疗前后外周血C-反应蛋白变化情况及MCP-1及MMP-9水平及临床疗效。结果:与治疗前相比,两组患者治疗后MCP-1、MMP-9及C-反应蛋白水平降低(P0.05),与对照组相比,实验组患者MCP-1、MMP-9及C-反应蛋白较低(P0.05);与对照组相比,实验组总有效率较高(P0.05)。结论:灯盏细辛口服液对急性冠状动脉综合征患者具有较好的疗效,这可能与其降低外周血中C-反应蛋白、MMP-9和MCP-1水平具有一定的关系。     

Immunocharacterization of matrix metalloproteinase-2 and matrix metalloproteinase-9 in canine transmissible venereal tumors

Matrix metalloproteases (MMPs) are endogenous proteases that are responsible for degradation of extracellular matrix (ECM) proteins and cell surface antigens. The breakdown of ECM participates in the local invasion and distant metastases of malignant tumors. Canine transmissible venereal tumor (CTVT) is a naturally occurring contagious round cell neoplasm of dogs that affects mainly the external genitalia of both sexes. CTVT generally is a locally invasive tumor, but distant metastases also are common in puppies and immunocompromised dogs. We investigated the immune expressions and activities of MMP-2 and MMP-9 in CTVT. The presence of these enzymes in tumor cells and tissue homogenates was demonstrated by immunohistochemistry and western blotting. We used gelatin substrate zymography to evaluate the activities of MMP-2 and MMP-9 enzymes in tumor homogenates. We found that tumor cells expressed both MMP-2 and MMP-9. Electrophoretic bands corresponding to MMP-9 and MMP-2 were identified in immunoblots and clear bands that corresponded to the active forms of MMP-2 and MMP-9 also were detected in gelatin zymograms. Our study is the first detailed documentation of MMPs in CTVT.