MMP-2、TIMP-2、HER-2及p38MAPK在卵巢浆液性上皮性肿瘤组织中的表达及意义

目的探讨基质金属蛋白酶-2（MMP-2）及其组织抑制因子（TIMP-2）、HER-2、p38丝裂原蛋白激酶（p38MAPK）在卵巢浆液性上皮性肿瘤组织中的表达及意义。方法采用组织微阵列技术和免疫组化法检测MMP-2、TIMP-2、HER-2、p38及其活化体P—p38在25例浆液性腺癌、7例卵巢交界性肿瘤和17例卵巢浆液性囊腺瘤组织中的表达。结果MMP-2、TIMP-2和p38MAPK表达阳性率及HER-2的过表达率在卵巢浆液性腺癌及交界性肿瘤中明显高于浆液性囊腺瘤（P〈0．05）；p38表达阳性率和MMP-2／TIMP-2随着卵巢恶性肿瘤的发展逐步提高（P〈0．05），P—p38与肿瘤分期、大小、转移情况及患者年龄明显相关（P〈0．05）；MMP-2、HER-2和P—p38在卵巢浆液性腺癌中的表达两两相关（P〈0．05）。结论HER-2、p38MAPK、MMP-2和TIMP-2通路可能参与了卵巢癌的发生、侵袭和转移；p38MAPK的磷酸化可能成为卵巢浆液性肿瘤恶化的分子指标及卵巢癌治疗的分子靶标之-。     

CD147、MMP-2和JNK在卵巢上皮癌中的表达及意义

应用免疫组化SP法检测45例卵巢上皮癌、10例交界性卵巢上皮肿瘤、15例良性卵巢上皮肿瘤及15例正常卵巢组织中的CD147、基质金属蛋白酶2（MMP-2）和c-Jun氨基末端激酶（JNK）的表达。结果卵巢上皮癌CD147和MMP-2的阳性表达率均高于其他卵巢上皮组织中的表达（P均〈0.05）;CD147和MMP-2与临床分期有关（P〈0.01、〈0.05）;CD147和MMP-2表达及MMP-2和JNK间的表达均呈正相关（r=0.655、0.477,P〈0.01、〈0.05）。提示CD147、MMP-2和JNK在卵巢上皮癌的增殖和浸润过程中密切相关。     

卵巢浆液性囊腺癌组织中TRAIL的表达及意义

目的探讨肿瘤坏死因子相关凋亡诱导配体（TRAIL）在卵巢浆液性囊腺癌发生、发展中的作用。方法选择手术切除的卵巢浆液性囊腺癌标本56份（恶性组）,卵巢交界性浆液性囊腺瘤12份（交界性组）,卵巢浆液性囊腺瘤14份（良性组）,采用免疫组化PV法检测三组TRAIL表达,并分析其与卵巢浆液性囊腺癌临床病理特征的相关性。结果恶性组、交界性组、良性组TRAIL阳性表达率分别为28．6％、58．3％、92．9％,三组比较P均〈0．05。TRAIL阳性表达与卵巢浆液性囊腺癌临床分析相关,与组织分化程度无关。结论TRAIL在卵巢浆液性囊腺癌的恶性转化中可能起一定作用,TRAIL丢失可能与卵巢浆液性囊腺癌恶性进展有关。     

MMP-9和TIMP-3在异位子宫内膜及卵巢癌中的表达

采用免疫组化SP法检测基质金属蛋白酶（MMP）-9、基质金属蛋白酶的组织抑制剂（TIMP）-3在30例卵巢内膜异位囊肿、30例卵巢癌、30例卵巢良性肿瘤和20例正常卵巢组织中的表达。发现MMP-9在卵巢癌组织中阳性率最高，卵巢异位囊肿次之，均明显高于卵巢良性肿瘤和正常卵巢组（P〈0．01）；TIMP-3在异位子宫内膜及正常子宫内膜中的表达高于卵巢异位囊肿（P〈0．05）；卵巢癌与卵巢异位囊肿组MMP-9／TIMP-3〉1，其他各组MMP-9/TIMP-3〈1。提示MMPO和TIMP-3在卵巢内膜异位囊肿及卵巢癌中不同程度的异常表达并失去平衡，子宫内膜异位症与卵巢癌有类似的生物学基础，对卵巢异位囊肿应注意恶变的可能。     

TIMP-2、MMP-2和MT1-MMP在活化肝星状细胞中的表达及对细胞外基质合成分泌的影响

目的 研究金属蛋白酶组织抑制剂-2(TIMP-2)、基质金属蛋白酶-2(MMP-2)和膜型基质金属蛋白酶-1(MT1-MMP)在活化肝星状细胞(HSCs)中的表达,观察其对细胞外基质(ECM)合成分泌的影响.方法 原代分离培养大鼠HSCs活化后,分别给予40～160 pmol化学合成经修饰抗TIMP-2 siRNA进行干预,检测培养细胞上清液透明质酸(HA)、Ⅲ型前胶原(PCⅢ)和羟脯氨酸(Hyp)的含量,采用荧光实时定量PCR法检测TIMP-2、MMP-2、MT1-MMP、MMP-13、COL Ⅰ和COL Ⅲ mRNA的表达,western印迹检测TIMP-2、MT1-MMP和MMP-13蛋白表达及明胶酶谱法检测MMP-2蛋白表达.结果 应用化学合成经修饰抗TIMP-2 siRNA后,TIMP-2、MMP-2、MT1-MMP、COL Ⅰ和COL Ⅲ的表达明显降低,而MMP-13的表达则明显增加,培养细胞上清液中HA、PCⅢ和Hyp的含量也明显减少.结论 TIMP-2通过MT1-MMP介导MMP-2的活化,抑制TIMP-2的表达,MT1-MMP和MMP-2的表达随之降低,而HSCs合成分泌ECM也相应减少.     

成纤维细胞与肺癌细胞相互作用对膜型基质金属蛋白酶-1及基质金属蛋白酶-2表达的影响

目的 研究胚肺成纤维细胞对肺癌H460细胞膜型基质金属蛋白酶-1(MTl-MMP)、基质金属蛋白酶-2(MMP-2)表达的影响.方法 采用Western blot方法检测各组MT1-MMP的表达.取其上清,采用酶联免疫吸附法检测各组细胞培养液中活性MMP-2的浓度.结果 H460细胞、胚肺成纤维细胞单独培养时均有MT1-MMP表达,但混合培养后MT1-MMP表达增强(P＜0.05).H460细胞、胚肺成纤维细胞单独培养时MMP-2均有分泌,混合培养MMP-2分泌增强(P＜0.05).结论 胚肺成纤维细胞和肺癌H460细胞相互作用能通过上调MT1-MMP、MMP-2的表达从而促进肺癌的侵袭和转移,这可能为肺癌侵袭转移的一个重要机制.     

卵巢浆液性癌组织中C-kit、PDGFRβ的表达及意义

采用免疫组化法检测10例正常卵巢、18例良性卵巢浆液性囊腺瘤、16例交界卵巢浆液性囊腺瘤、42例卵巢浆液性囊腺癌组织中的C-kit、PDGFRβ蛋白。结果显示,C-kit在卵巢浆液性囊腺癌、交界性卵巢浆液性囊腺瘤、良性卵巢浆液性囊腺瘤中阳性表达率分别为61.9%、43.8%、11.1%,正常卵巢组织为阴性,前两者与后者相比,P均〈0.05;C-kit在卵巢浆液性癌中的表达与其组织学分级、FIGO分期有关,P均〈0.05。PDGFRβ在卵巢浆液性囊腺癌、正常卵巢组织、良性卵巢浆液性囊腺瘤、交界性卵巢浆液性囊腺瘤中的阳性表达率分别为83.3%、70.0%、77.8%、81.3%,四者相比,P均〉0.05;PDGFRβ蛋白在低分化与高分化卵巢浆液性囊腺癌中的表达相比,P〈0.05。认为C-kit和PDGFRβ在卵巢浆液性癌的发生、发展过程中有一定作用。     

MT1-MMP对人肝癌细胞浸润能力的影响

目的 观察膜型基质金属蛋白酶-1(MT1-MMP)对人肝癌细胞株浸润能力的影响,并初步探讨其作用机制.方法 将稳定表达MT1-MMP基因转染至HepG2细胞中,应用Western印迹法检测MT1-MMP蛋白表达;明胶酶谱分析法检测MMP-2酶原活性;体外侵袭实验检测细胞株浸润能力.结果 重组质粒转染株MT1-MMP蛋白表达水平明显高于空质粒组和未转染组(P＜0.01).明胶酶谱分析实验发现,MT1-MMP组同时检测到72000酶原形式和64000活性形式的MMP-2,另两组只检测到72000酶原形式的MMP-2.体外侵袭实验结果显示,MT1-MMP组细胞穿过基质胶(Matrigel)的细胞数目明显多于其他两组(P＜0.01).结论 MT1-MMP能显著增强肝癌细胞株的浸润能力,其机制主要是通过激活MMP-2-酶原,降解肿瘤周围的基质成分实现的.     

灵芝多糖对糖尿病大鼠肾组织MMP-2和TIMP-2表达的影响

目的观察基质金属蛋白酶-2（MMP-2）、基质金属蛋白酶抑制因子-2（TIMP-2）在糖尿病大鼠肾组织中的表达及灵芝多糖干预后的影响。方法腹腔注射链脲佐菌素（STZ）诱导制备糖尿病大鼠模型，分组给予不同剂量灵芝多糖（GLP，100、200、400mg／kg）进行治疗性灌胃。8w后，免疫组化和RT-PCR方法雄测各组大鼠肾皮质MMP-2、TIMP-2的表达。结果糖尿病组大鼠肾小球MMP-2的表达较正常对照组明显减少，TIMP-2明显升高（P〈0．01）；灵芝多糖各组较糖尿病组MMP-2表达升高（P〈0．01），TIMP-2表达减少（P〈0．01）。结论灵芝多糖通过调节MMP-2／TIMP-2的平衡，减少细胞外基质积聚，对糖尿病大鼠肾脏起保护作用。     

MMP-2、TIMP-2在脑膜瘤组织中的表达及临床意义

目的探讨基质金属蛋白酶-2（MMP-2）与基质金属蛋白酶组织抑制因子-2（TIMP-2）在脑膜瘤组织中的表达及意义。方法选择恶性脑膜瘤标本15份（恶性组）,良性脑膜瘤标本20份（良性组）,采用免疫组化sP法检测两组MMP-2和TIMP-2表达情况,采用spe蛐an等级相关分析法检测两指标的相关性。结果恶性组及良性组MMP-2阳性表达率分别为73．3％（11／15）、15％（3／20）,TIMP-2阳性表达率分别为13．3％（2／15）、80％（16／20）,两组比较P均〈0．01。脑膜瘤中MMP-2与TIMP-2表达呈负相关（r=-0．545,P〈0．01）。结论MMP-2可作为判断肿瘤恶变的指标,TIMP-2可作为肿瘤非恶变的指标,两指标检测可用于指导脑膜瘤的治疗及预后评估。     

The prognostic value of metalloproteinases and angiogenic factors in ovarian carcinoma

The objective of this study was to analyze the correlation between matrix metalloproteinases (MMPs) and angiogenic genes and survival in advanced-stage ovarian carcinomas. Primary and metastatic ovarian carcinomas from patients diagnosed with FIGO stage III-IV disease and followed up to 20 years were studied using mRNA in situ hybridization (ISH). Expression of MMP-2, MMP-9, membrane-type 1-MMP (MT1-MMP), the MMP inhibitor TIMP-2, vascular endothelial growth factor (VEGF), interleukin-8 (IL-8) and basic fibroblast growth factor (bFGF) was studied. MMP-2, MMP-9 and TIMP-2 mRNA was detected in both tumor and stromal cells, while MT1-MMP was largely confined to tumor cells. In univariate analysis of primary tumors, TIMP-2 and MMP-9 mRNA expression correlated with poor outcome. In metastatic lesions, mRNA expression of TIMP-2, MMP-2, and MT1-MMP correlated with poor survival. In a multivariate analysis of primary tumors, TIMP-2 expression in stromal cells (P=0.006) and MMP-9 expression in tumor cells (P=0.011) retained their predictive value. Intense expression of bFGF mRNA and weak expression of IL-8 mRNA was detected in both stromal and tumor cells in most cases, while VEGF mRNA expression was limited to a few cases. Angiogenic mRNA expression showed no correlation with disease outcome in survival analysis (P>0.05). We conclude that bFGF is the major angiogenic factor expressed in ovarian carcinoma at the mRNA level. MMP-2, MMP-9, MT1-MMP and TIMP-2 are valid markers of poor survival in advanced-stage ovarian carcinoma.     

Survivin gene expression in endometriosis

Survivin is a novel inhibitor of apoptosis and is expressed during fetal development and in cancer tissues, but its expression has not been reported in normal adult tissues or benign diseases. We investigated survivin gene and protein expression in a tumor-like benign disease, endometriosis, and correlated them with apoptosis and invasive phenotype of endometriotic tissues. Gene expression levels of survivin, matrix metalloproteinase (MMP)-2, MMP-9, and membrane type 1 (MT1)-MMP in 63 pigmented or nonpigmented endometriotic tissues surgically obtained from 35 women with endometriosis were compared with those in normal eutopic endometrium obtained from 12 women without endometriosis. Survivin, MMP-2, MMP-9, and MT1-MMP mRNA expression levels in clinically aggressive pigmented lesions were significantly higher than those in normal eutopic endometrium, and survivin gene expression in pigmented lesions was also higher than that in nonpigmented lesions (P < 0.05). There was a close correlation between survivin and MMP-2, MMP-9, or MT1-MMP gene expression levels in 63 endometriotic tissues examined (P < 0.01). Apoptotic cells detected by the dUTP nick-end labeling were rare in 11 ovarian endometriotic tissues, which showed positive immunohistochemical expression for survivin and MMPs. Our findings suggest that up-regulation of survivin and MMPs may cooperatively contribute to survival and invasion of endometriosis.     

Overexpression of MT3-MMP in hepatocellular carcinoma correlates with capsular invasion

BACKGROUND/AIMS: Extracellular matrix-degrading matrix metalloproteinases (MMPs) are invariably up-regulated in epithelial cancers and are key agonists of angiogenesis, invasion and metastasis. Recent studies have shown high levels of various MMPs, including MT1-MMP, MMP-1, MMP-2 and MMP-9, and their involvement in tumor progression in human hepatocellular carcinoma (HCC). However, the expression and role of MT3-MMP in HCC remains unclear. METHODOLOGY: We examined the immunohistochemical expression of MT3-MMP in surgically resected HCCs (n=58), hepatitis C virus (HCV) and hepatitis B virus (HBV)-related chronic hepatitis (n=34) and cirrhosis (n=24). RESULTS: MT3-MMP expression was observed in all non-cancerous liver tissues. In HCCs, 52% (30/58) of patients showed high MT3-MMP expression while the remaining 48% (28/58) of patients showed low expression. A clinicopathological survey demonstrated a significant correlation between high MT3-MMP expression and capsular invasion of carcinoma (p = 0.034) although there was no correlation between high MT3-MMP expression in HCC and overall survival or disease-free survival. CONCLUSIONS: MT3-MMP was expressed not only in chronic hepatitis and liver cirrhosis, but also in HCC, and high MT3-MMP expression correlated significantly with capsular invasion of carcinoma.     

Matrix metalloproteinase (MMP) expression by differentiated thyroid carcinoma of children and adolescents

The factor(s) that control metastasis of thyroid carcinoma are unknown, but the matrix metalloproteinases (MMPs) are excellent candidates. MMP-1, membrane-type-1 MMP (MT1-MMP), and tissue inhibitor of MMP-1 (TIMP-1) have all been implicated, but the site of production and importance are disputed. In vitro, normal thyroid cells secrete TIMP-1, while thyroid cancer cells secrete TIMP-1 and MMP-1. However, previous pathological studies identified MMP-1 and TIMP-1 only in the stroma surrounding thyroid carcinoma. These data suggest that thyroid carcinoma or tumor-associated inflammatory cells might secrete a factor(s) which stimulates MMP-1 or TIMP-1 expression by surrounding tissues. We hypothesized that MMP-1, MT1-MMP, and TIMP-1 would be directly expressed by thyroid carcinoma and might promote invasion or metastasis. We used immunohistochemistry to determine the expression of MMP-1, MT1-MMP, and TIMP-1 in 32 papillary thyroid carcinoma (PTC), 10 follicular thyroid carcinoma (FTC) and 13 benign thyroid lesions from children and adolescents. The intensity of staining was graded from absent (grade 0) to intense (grade 3). Average MMP-1 expression (mean relative intensity units+/-SE) was significantly greater among PTC (1.97+/-0.15; p=0.004) and FTC (2.2+/-0.25; p=0.006) compared to benign lesions (1.30+/-0.15); but there was no relationship between MMP-1 expression and invasion, metastasis, or recurrence. Expression of MT1-MMP and TIMP-1 was similar for benign and malignant lesions; but recurrent PTC expressed lower levels of TIMP-1 when compared to non-recurrent PTC (p=0.049). Only the expression of TIMP-1 correlated with the presence of tumor-associated lymphocytes (r=0.35, p=0.032). We conclude that MMP-1, MT1-MMP and TIMP-1 are all expressed by thyroid carcinoma and could be important in promoting recurrence.     

Expression of membrane-type matrix metalloproteinases in synovial tissue from patients with rheumatoid arthritis or osteoarthritis

We investigated the expression of membrane-type matrix metalloproteinase (MT-MMP) and matrix metalloproteinase (MMP) mRNAs in synovial tissue from patients with rheumatoid arthritis (RA, n = 5) or osteoarthritis (OA, n = 5) by Northern blot analysis. Northern analysis demonstrated strong expression of MT1-MMP, MT3-MMP, MMP-1, and MMP-3 and weak expression of MT2-MMP and MMP-8 in synovial tissue from patients with RA or OA. MT4-MMP was not detected. No significant difference was shown in the expression of MT-MMP mRNAs between RA and OA. Synovial tissue of RA or OA patients expressed MT-MMPs as well as MMPs. These results indicate that, in addition to MMPs, MT1-MMP, MT3-MMP, and probably MT2-MMP may play a role in the degradation of bone and cartilage matrix in RA and OA. Such information may provide a clue to the development of a novel therapeutic approach targeted on the prevention of joint destruction. Received: April 30, 2000 / Accepted: September 19, 2000     

Chronic hypoxia inhibits MMP-2 activation and cellular invasion in human cardiac myofibroblasts

Cardiac myofibroblasts are pivotal to adaptive remodelling after myocardial infarction (MI). These normally quiescent cells invade and proliferate as a wound healing response, facilitated by activation of matrix metalloproteinases, particularly MMP-2. Following MI these reparative events occur under chronically hypoxic conditions yet the mechanisms by which hypoxia might modulate MMP-2 activation and cardiac myofibroblast invasion have not been investigated. Human cardiac myofibroblasts cultured in collagen-supplemented medium were exposed to normoxia (20% O₂) or hypoxia (1% O₂) for up to 48 h. Secreted levels of total and active MMP-2 were quantified using gelatin zymography, TIMP-2 and membrane-associated MT1-MMP were quantified with ELISA, whole cell MT1-MMP by immunoblotting and immunocytochemistry and MT1-MMP mRNA with real-time RT-PCR. Cellular invasion was assessed in modified Boyden chambers and migration by scratch wound assay.In the human cardiac myofibroblast, MT1-MMP was central to MMP-2 activation and activated MMP-2 necessary for invasion, confirmed by gene silencing. MMP-2 activation was substantially attenuated by hypoxia (P < 0.001), paralleled by inhibition of myofibroblast invasion (P < 0.05). In contrast, migration was independent of either MT1-MMP or MMP-2. Reduced membrane expression of MT1-MMP (P < 0.05) was responsible for the hypoxic reduction of MMP-2 activation, with no change in either total MMP-2 or TIMP-2. In conclusion, hypoxia reduces MMP-2 activation and subsequent invasion of human cardiac myofibroblasts by reducing membrane expression of MT1-MMP and may delay healing after MI. Regulation of these MMPs remains an attractive target for therapeutic intervention.     

Expression of membrane type 1 matrix metalloproteinase, matrix metalloproteinase 2 and tissue inhibitor of metalloproteinase 2 in human cartilaginous tumors with special emphasis on mesenchymal and dedifferentiated chondrosarcoma

Membrane type 1 matrix metalloproteinase (MT1-MMP) has been identified as an activator of the proenzyme of matrix metalloproteinase 2 (MMP-2: gelatinase A), and has also been shown to play a crucial role in tumor invasion by activating proMMP2 in both lung and gastric carcinoma. The tissue inhibitor of metalloproteinase 2 (TIMP-2) plus the MT1-MMP complex also plays an important role in the activation of proMMP-2. In this study, the expressions of MT1-MMP, MMP-2 and TIMP-2 were evaluated in 10 enchondromas, 34 conventional chondrosarcomas, 5 clear-cell chondrosarcomas, 7 mesenchymal chondrosarcomas and 8 dedifferentiated chondrosarcomas. The expressions were immunohistochemically visualized on paraffin sections and the levels of expression were assessed semiquantitatively. The extent of staining was assessed by the extent score in order to determine the overall level of expression. The extent scores of MT1-MMP, MMP-2 and TIMP-2 in grade 2 chondrosarcoma were significantly higher than those in either enchondroma or grade 1 chondrosarcoma (P < 0.05). In conventional chondrosarcoma, significant correlations were found between the extent scores of MT1-MMP and MMP-2 (P < 0.001), MT1-MMP and TIMP-2 (P < 0.01), and MMP-2 and TIMP-2 (P < 0.01). The undifferentiated small round tumor cells of mesenchymal chondrosarcoma showed lower positive rates and extent scores for MT1-MMP (2/7, 0.7 ± 0.5) and MMP-2 (3/7, 0.7 ± 0.4) than for cartilaginous components of mesenchymal chondrosarcoma [MT1-MMP (4/7, 1.3 ± 0.5) and MMP-2 (7/7, 1.9 ± 0.3)] or conventional chondrosarcoma. In dedifferentiated chondrosarcoma, the extent scores of MT1-MMP, MMP-2 and TIMP-2 in low-grade cartilaginous components were not significantly different from those in conventional chondrosarcoma; however, the high-grade anaplastic components showed high extent scores for MT1-MMP, MMP-2 and TIMP-2, compared with the low-grade cartilaginous components of dedifferentiated chondrosarcoma or conventional chondrosarcoma. According to our results, the expression of MT1-MMP as well as that of MMP-2 or TIMP-2 demonstrated a significant correlation with the tumor grade in human cartilaginous tumors. Furthermore, the expressions of MT1-MMP, MMP-2 and TIMP-2 were also found to play a crucial role in invasion in the high-grade components of dedifferentiated chondrosarcoma. Received: 17 February 1999 / Accepted: 21 April 1999     

1,25-二羟维生素D3对人成骨细胞基质金属蛋白酶及其抑制因子的作用

目的　研究 1, 25 二羟维生素D3 [ 1α, 25 (OH)2D3 ]对人成骨细胞基质金属蛋白酶(MMP) 1、MMP 2、膜型基质金属蛋白酶 1 (MT1 MMP)、基质金属蛋白酶抑制因子 1 (TIMP 1 )的影响,探讨 1α, 25(OH)2D3 调节骨代谢作用机制。方法　人成骨细胞用 1α, 25 (OH)2D3 干预。Western杂交检测MT1 MMP蛋白质表达。MMP 1、MMP 2、TIMP 1的分泌及MMP 2的活性用ELISA检测。Northern杂交检测维生素D受体、MT1 MMPmRNA表达。结果　 1α, 25 (OH)2D3 对人成骨细胞MMP 1、MMP 2、TIMP 1表达无影响, 10-10 ~ 10-8 mol/L 1α, 25 (OH)2D3 干预诱导成骨细胞MT1 MMP表达呈剂量依赖性 (P值均 <0 05);促进MMP 2激活呈剂量依赖性 [MMP 2活性分别为(42 3 ± 8 6)、(64 4 ±11 4)、(93 5 ±9 9)μg/L, P值均<0 05]。结论　由于MT1 MMP在骨吸收过程中起着关键作用, 1α, 25(OH)2D3 可通过诱导成骨细胞MT1 MMP表达刺激骨吸收。