Downregulation Of MMP-9 Expression Research Articles

Regulation of MMP-9 expression by the A2b adenosine receptor and its dependency on TNF-α signaling

Macrophage- and vascular-derived matrix metalloproteinase (MMP)-9 plays an important role in neointima formation after vascular injury. The A2b adenosine receptor (A2bAR) elevates cyclic adenosine monophosphate and suppresses tumor necrosis factor-α (TNF-α) levels at baseline and after vascular injury. Considering the influences of TNF-α on MMP-9 expression and activity, here we examined the effect of the A2bAR on the expression of MMP-9 and its potential dependency on TNF-α. We applied protein activity and mRNA analyses of MMP-9 in macrophages derived from A2bAR knockout (KO) and TNF-α receptor KO mice. We employed guidewire-induced femoral artery injuries on A2bAR KO and control mice and analyzed by immunohistochemistry MMP-9 expression in the neointima area. MMP-9 activity is somewhat less in resident A2bAR KO macrophages compared with wild-type cells. However, MMP-9 is increased in activated macrophages from A2bAR KO when TNF-α is further elevated, or in wild-type cells after TNF-α treatment. In accordance, A2bAR activation downregulates MMP-9 expression in wild-type macrophages, which is ablated in TNF-α receptor KO cells. A greater vascular lesion after femoral artery injury in A2bAR KO mice is associated with elevated TNF-α levels and augmented MMP-9, compared to control mice. Ablation of the A2bAR in activated macrophages increases MMP-9. A2bAR activation reduces MMP-9 expression, which depends on TNF-α and could contribute to the protective role of A2bAR in a vascular injury model.

BAG3 (BCL2-associated athanogene 3) interacts with MMP-2 to positively regulate invasion by ovarian carcinoma cells

BAG3 (BCL2-associated athanogene 3) is a member of the BAG family proteins, which interact with and regulate Hsp70. Our aim in the present study was to correlate BAG3 expression in ovarian cancer with invasion and progression-free survival. We found that BAG3 interacts with MMP2 in cultured ovarian cancer cells, and that knockdown of BAG3 expression downregulated MMP2 expression and diminished cell motility and invasiveness. The incidence of BAG3 positivity was significantly higher at advanced clinical stages of ovarian cancer than at early stages. We suggest that BAG3 binds to MMP2 to positively regulate the process of cell invasion.

SiRNA-mediated silencing of the CXCR4 gene down-regulates MMP-9 expression in EC-9706 cells

目的：探讨在食管鳞癌EC-9706细胞中沉默CXCR4基因对MMP-9基因表达的影响,为阐明CXCR4基因在食管鳞癌侵袭转移中的作用提供实验依据.方法：化学合成2条靶向CXCR4基因的siRNA1和siRNA2,同时设立荧光标记阴性对照和空白对照.脂质体法转染入EC-9706细胞,荧光显微镜下观察转染效率.转染48h后,半定量RT-PCR检测各组细胞CXCR4和MMP-9基因mRNA表达的变化,Western blot检测各组细胞CXCR4和MMP-9基因蛋白表达的变化,侵袭小室检测各组细胞穿膜细胞数的变化,MTT检测各组细胞的A值.结果：与阴性对照和空白对照相比,转染CXCR4siRNA1和siRNA2组细胞CXCR4mRNA和蛋白的表达明显降低,差异具有统计学意义（P〈0.05）;同时与阴性对照和空白对照相比,转染CXCR4siRNA1和siRNA2组细胞MMP-9mRNA和蛋白的表达同样明显降低,差异具有统计学意义（P〈0.05）;转染CXCR4siRNA1和siRNA2组细胞穿膜细胞数与阴性对照和空白对照相比明显下降,差异具有统计学意义（P〈0.05）;MTT结果显示转染CXCR4siRNA1和siRNA2组细胞增殖能力下降,与阴性对照和空白对照相比差异具有统计学意义（P〈0.05）.结论：在食管鳞癌EC9706中存在CXCR4基因对MMP-9基因的调控作用,CXCR4基因有可能通过调控MMP-9基因的表达参与食管鳞癌细胞的浸润转移,CXCR4有可能成为食管鳞癌基因治疗的有效靶点.

SLUG promotes prostate cancer cell migration and invasion via CXCR4/CXCL12 axis

BackgroundSLUG is a zinc-finger transcription factor of the Snail/Slug zinc-finger family that plays a role in migration and invasion of tumor cells. Mechanisms by which SLUG promotes migration and invasion in prostate cancers remain elusive.MethodsExpression level of CXCR4 and CXCL12 was examined by Western blot, RT-PCR, and qPCR analyses. Forced expression of SLUG was mediated by retroviruses, and SLUG and CXCL12 was downregulated by shRNAs-expressing lentiviruses. Migration and invasion of prostate cancer were measured by scratch-wound assay and invasion assay, respectively.ResearchWe demonstrated that forced expression of SLUG elevated CXCR4 and CXCL12 expression in human prostate cancer cell lines PC3, DU145, 22RV1, and LNCaP; conversely, reduced expression of SLUG by shRNA downregulated CXCR4 and CXCL12 expression at RNA and protein levels in prostate cancer cells. Furthermore, ectopic expression of SLUG increased MMP9 expression and activity in PC3, 22RV1, and DU-145 cells, and SLUG knockdown by shRNA downregulated MMP9 expression. We showed that CXCL12 is required for SLUG-mediated MMP9 expression in prostate cancer cells. Moreover, we found that migration and invasion of prostate cancer cells was increased by ectopic expression of SLUG and decreased by SLUG knockdown. Notably, knockdown of CXCL12 by shRNA impaired SLUG-mediated migration and invasion in prostate cancer cells. Lastly, our data suggest that CXCL12 and SLUG regulate migration and invasion of prostate cancer cells independent of cell growth.ConclusionWe provide the first compelling evidence that upregulation of autocrine CXCL12 is a major mechanism underlying SLUG-mediated migration and invasion of prostate cancer cells. Our findings suggest that CXCL12 is a therapeutic target for prostate cancer metastasis.

Shikonin inhibits tumor invasion via down‐regulation of NF‐κB‐mediated MMP‐9 expression in human ACC‐M cells

The aim of this study was to examine the anti-invasion effect of Shikonin on human high-metastatic adenoid cystic carcinoma (ACC-M) cells and to explain the possible molecular mechanism involved. The ACC-M cells were treated with Shikonin (0, 2.5, 5, 10 μM) for 24 h. The protein levels and gelatinolytic activities of MMP-2 and MMP-9 were analyzed using Western blot and Gelatin zymography test, respectively. Matrigel invasion assays were used to investigate tumor invasive potential and electromobility shift assays were used to determine the activity of NF-κB. The invasiveness of ACC-M cells was reduced in a dose dependent manner following 24-h treatment of up to 10 μM of the Shikonin at which concentration no cytotoxicity occurred. The protein levels and gelatinolytic activities of MMP-9 were significantly suppressed by increasing Shikonin concentrations. The down-regulation of MMP-9 appeared to be via the inactivation of NF-κB as the treatment with Shikonin suppressed the protein level of phosphate-IkBa, which was accompanied by a decrease in DNA-binding level of the factor. Shikonin inhibits tumor invasion via downregulation of MMP-9 expression in ACC-M cells. Pharmacologic inhibition of the NF-κB-mediated MMP-9 expression by Shikonin might be a powerful treatment option for ACC patients in future.

Inhibitive effect of celecoxib on the adhesion and invasion of human tongue squamous carcinoma cells to extracellular matrix via down regulation of MMP-2 expression

The effect of cyclooxygenase 2 (COX-2) and its inhibitor on metastasis and invasion of tumor and its relationship with matrix metalloproteinase are getting more and more attention. Down regulation of matrix metalloproteinase (MMP) expression could inhibit the invasion and adhesion ability of cancer cells. Our study observed the inhibitive effect of selective COX-2 inhibitor, celecoxib, on adhesion and invasion ability of human tongue squamous carcinoma cell line Tca8113 and its relationship with MMP-2 secretion. COX-2 expression of Tca8113 cell was detected by Western blot analysis. Ratio of COX-2 positive Tca8113 cell was determined by flow cytometry analysis. MMP-2 and PGE2 secretion was measured by ELISA. Invasion of a Matrigel-coated membrane by Tca8113 cells was quantitatively assessed by counting migrated cells. Results showed that COX-2 expression was positive in Tca8113 cell and could be significantly inhibited by celecoxib. Celecoxib could inhibit PGE2 production of Tca8113 cell in a dose-dependent manner, down-regulate MMP-2 secretion of Tca8113 cell, and at the same time significantly inhibit invasion and adhesion ability of these cells. Adding extrinsic PGE2 could antagonize the inhibitive effect of celecoxib not only on MMP-2 secretion of Tca8113 cell, but also on invasion and adhesion ability of the cells. The results suggested that celecoxib had chemoprevention effect on invasion and metastasis of oral carcinoma.

Downregulation of Matrix Metalloproteinase–2 in Corneal Fibroblasts by Interleukin-1 Receptor Antagonist Released from Corneal Epithelial Cells

The authors previously showed that the expression of various junctional proteins in corneal epithelial cells or corneal fibroblasts (CFs) is regulated by the presence of the other cell type in a coculture system. In this study, the effect of corneal epithelial cells on the expression of matrix metalloproteinases (MMPs) in CFs was studied. Human CFs and simian virus 40-transformed human corneal epithelial (HCE) cells were cultured on opposite sides of a collagen vitrigel membrane. Expression of MMPs in CFs was examined by reverse transcription-polymerase chain reaction and immunoblot analyses. The amounts of MMP-2 mRNA and protein in CFs were decreased by the presence of HCE cells. HCE cells had no effect on the expression of MMP-1 in CFs. HCE cells released interleukin (IL)-1 receptor antagonist (IL-1RA) into the culture medium, and depletion of IL-1RA in HCE cells by RNA interference largely abolished the effect of these cells on MMP-2 expression in CFs. The downregulation of MMP-2 expression in CFs by HCE cells was blocked by an inhibitor of signaling by the mitogen-activated protein kinase (MAPK) ERK (PD98059) but was not affected by those of signaling by the MAPKs p38 (SB203580) or JNK (SP600125). Corneal epithelial cells downregulated expression of MMP-2 in CFs in a manner dependent at least in part on the release of IL-1RA from the former cells. This effect might contribute to the attenuation of corneal stromal remodeling by corneal epithelial cells.

Effects of recombinant human endostatin and docetaxel on MMP and its following anti-neoplastic effect under different administration sequences

背景与目的通过药物干预性动物实验，观察重组人血管内皮抑素（rh-endostatin）与多西紫杉醇不同顺序联合用药后MMP-2及相关因子的变化及其对移植瘤生长与血管生成的影响，探索两药联合的作用机制和最佳抗肿瘤方案。方法建立肺腺癌A549荷瘤裸鼠模型，分两个阶段进行实验。第一阶段：将荷瘤小鼠随机分为重组人血管内皮抑素组（重组人血管内皮抑素400 μg·d-1，d1-d14）和多西紫杉醇组（多西紫杉醇10 mg·kg-1·3d-1，d1-d14）；第二阶段：将荷瘤小鼠随机分为同时用药组（重组人血管内皮抑素400 μg·d-1、d1-d35，多西紫杉醇10 mg·kg-1·3d-1、d1-d19）、先重组人血管内皮抑素组（重组人血管内皮抑素400 μg·d-1、d1-d35，多西紫杉醇10 mg·kg-1·3d-1、d16-d34）和模型组。实验中动态测量移植瘤体积，结束后以免疫组化方法检测MMP-2、TIMP-2、EMMPRIN表达并计数微血管密度（MVD）。结果两单药组比较，重组人血管内皮抑素组MMP-2和EMMPRIN表达下调较多西紫杉醇组（P=0.024, P=0.081）明显，两组间TIMP-2表达无明显差异（P > 0.05）。联合组用药结束时，同时用药组与先重组人血管内皮抑素组的移植瘤体积小于模型组（P < 0.001, P=0.003），且MMP-2表达均明显下调、微血管数减少（P < 0.05），但同时用药组对肿瘤生长的抑制较先重组人血管内皮抑素组明显；与模型组相比，同时用药组TIMP-2上调（P=0.001）、EMMPRIN下调（P=0.018），先重组人血管内皮抑素组未见相似结果。结论同时用药方案可以从TIMP-2、EMMPRIN两个环节下调MMP-2的表达，从而更好地抑制肿瘤生长。

Inhibitory Effect of Panduratin A on c-Jun N-Terminal Kinase and Activator Protein-1 Signaling Involved in Porphyromonas gingivalis Supernatant-Stimulated Matrix Metalloproteinase-9 Expression in Human Oral Epidermoid Cells

Porphyromonas gingivalis, a type of Gram-negative periodontopathogen, causes periodontal disease by activating intracellular signaling pathways that produce excessive inflammatory responses such as matrix metalloproteinases (MMPs). Recently, we reported that panduratin A, a chalcone compound isolated from Kaempferia pandurata ROXB., caused the decreased levels of MMP-9 secretion, protein, and gene expression in human oral epidermoid KB cells exposed to P. gingivalis supernatant. In this study, we clarified if mitogen-activated protein kinase (MAPK) signaling mediated MMP-9 expression by examining the effect of specific MAPK inhibitors, i.e. U0126, SB203580, and SP600125, on P. gingivalis supernatant-stimulated MMP-9 expression in KB cells. We next elucidated the molecular mechanism by which panduratin A attenuated signaling pathways involved in MMP-9 expression by performing gelatin zymography, Western blotting, reverse transcription-polymerase chain reaction, and promoter assays. Exposure of KB cells to P. gingivalis supernatant up-regulated the expression of MMP-9 protein and gene, and activation of activator protein-1 (AP-1) element, MAPK phosphorylation (extracellular signal-related kinase 1/2 (ERK1/2), p38, and c-Jun N-terminal kinase (JNK)), and transcription factors (Elk1, c-Jun, and c-Fos). A JNK inhibitor (SP600125) significantly attenuated MMP-9 gene expression and AP-1 activity in KB cells in response to P. gingivalis supernatant. Similar to SP600125, panduratin A was found to strongly suppress the level of phosphorylated JNK and block AP-1 activity in P. gingivalis supernatant-stimulated KB cells. In summary, JNK and AP-1 are the major signaling for P. gingivalis supernatant-stimulated MMP-9 expression in KB cells, and panduratin A markedly down-regulates MMP-9 expression through inhibition of these signaling.

Behavioral stress accelerates plaque pathogenesis in the brain of Tg2576 mice via generation of metabolic oxidative stress

Alzheimer's disease (AD) is a progressive neurodegenerative disease caused by genetic and non-genetic factors. Most AD cases may be triggered and promoted by non-genetic environmental factors. Clinical studies have reported that patients with AD show enhanced baseline levels of stress hormones in the blood, but their physiological significance with respect to the pathophysiology of AD is not clearly understood. Here we report that AD mouse models exposed to restraints for 2 h daily on 16 consecutive days show increased levels of beta-amyloid (Abeta) plaque deposition and commensurable enhancements in Abeta(1-42), tau hyperphosphorylation, and neuritic atrophy of cortical neurons. Repeated restraints in Tg2576 mice markedly increased metabolic oxidative stress and down-regulated the expression of MMP-2, a potent Abeta-degrading enzyme, in the brain. These stress effects were reversed by blocking the activation of the hypothalamus-pituitary-adrenal gland axis with the corticotropin-releasing factor receptor antagonist NBI 27914, further suggesting that over-activation of the hypothalamic-pituitary-adrenal axis is required for stress-enhanced AD-like pathogenesis. Consistent with these findings, corticosteroid treatments to cultured primary cortical neurons increased metabolic oxidative stress and down-regulated MMP-2 expression, and MMP-2 down-regulation was reversed by inhibition of oxidative stress. These results suggest that behavioral stress aggravates AD pathology via generation of metabolic oxidative stress and MMP-2 down-regulation.

TGF-beta1 Targeting on MMP—Expression in Keloid Fibroblasts

Problem Transforming growth factor-beta1 (TGF-beta1) has been identified as an important regulator of wound healing. Recent developments in molecular therapy offer exciting prospects for the modulation of wound healing, specifically those targeting TGF-beta1. The purpose of this study was to analyze the effect of TGF-beta1 targeting on the expression of matrix metallo-proteinases (MMP-2 and MMP-9) in fibroblasts cultured from earlobe keloids. Methods The expression of MMP-2 and MMP-9 in tissue samples from keloids was investigated by immunohistochemistry. The effect of TGF-beta1 targeting using antisense oligonucleotides on the expression of MMPs in keloid-derived fibroblasts was analyzed by ELISA and multiplex RT-PCR. Results Immunohistochemical studies demonstrated an increased expression of MMP-2 and MMP-9 proteins in tissue samples from keloids compared to normal human skin. The treatment of human keloid-derived fibroblasts with TGF-beta1 antisense oligonucleotides (ONDs) in vitro efficiently down-regulated MMP-9 expression but not the MMP-2 expression. In the supernatans of keloid-derived fibroblasts, the TGF-beta1 antisense therapy showed a decreased secretion level of both MMP-9 and MMP-2. So other than in MMP-9, the synthesis of MMP-2 does not seem to be influenced by TGF-beta1 anti-sense ONDs whereas the secretion of MMP-2, similar to that of MMP-9, is significantly decreased in the supernatans of keloid-derived fibroblasts. Conclusion TGF-beta1 antisense OND technology may be a potential therapeutic option for the inhibition of proteolytic tissue destruction in keloids. The results should be regarded as a further implication for the possible treatment of keloids. Significance The results should be regarded as a further implication for the possible treatment of keloids.

L-Carnosine Inhibits Metastasis of SK-Hep-1 Cells by Inhibition of Matrix Metaoproteinase-9 Expression and Induction of an Antimetastatic Gene, nm23-H1

Antioxidants have been suggested to inhibit the expression of matrix metalloproteinases (MMPs), especially MMP-9, which plays a critical role in tumor metastasis. Because of its antioxidant activity and the ability to chelate divalent cations, L-carnosine (LC) was tested for inhibition of MMP-9 in a highly invasive hepatocarcinoma, SK-Hep-1 cells. We found that LC (50–1,000 μ M) did not directly inhibit the activity of MMP-9 in a cell-free system. However, LC significantly inhibited the expression and activity of MMP-9 protein in SK-Hep-1 cells [inhibitory concentration of 50% (IC 50 )| = 105 and 63 μ M, respectively). Whereas LC did not inhibit the viability of SK-Hep-1 cells at concentrations up to 1,000 μ M within 3 days of incubation, this dipeptide significantly inhibited cell migration (IC 50 = 82 μ M) and invasion (IC 50 = 113 μM). LC significantly (P < 0.05) and dose dependently enhanced the expression of an antimetastatic gene, nonmetastatic cells 1, protein (nm23)-H1, at both protein and messenger ribonucleic acid (mRNA) levels. MMP-9 activity inversely correlated significantly with the expression of protein (r 2 = 0.77, P < 0.001) and mRNA (r 2 = 0.65, P < 0.001) of nm23-H1 in LC-treated cells. Thus, LC can inhibit the migration and invasion of SK-Hep-1 cells, and the effect is likely associated with upregulation of nm23-H1 and downregulation of MMP-9 expression.

Cannabinoids Inhibit Glioma Cell Invasion by Down-regulating Matrix Metalloproteinase-2 Expression

Cannabinoids, the active components of Cannabis sativa L. and their derivatives, inhibit tumor growth in laboratory animals by inducing apoptosis of tumor cells and impairing tumor angiogenesis. It has also been reported that these compounds inhibit tumor cell spreading, but the molecular targets of this cannabinoid action remain elusive. Here, we evaluated the effect of cannabinoids on matrix metalloproteinase (MMP) expression and its effect on tumor cell invasion. Local administration of Delta(9)-tetrahydrocannabinol (THC), the major active ingredient of cannabis, down-regulated MMP-2 expression in gliomas generated in mice, as determined by Western blot, immunofluorescence, and real-time quantitative PCR analyses. This cannabinoid-induced inhibition of MMP-2 expression in gliomas (a) was MMP-2-selective, as levels of other MMP family members were unaffected; (b) was mimicked by JWH-133, a CB(2) cannabinoid receptor-selective agonist that is devoid of psychoactive side effects; (c) was abrogated by fumonisin B1, a selective inhibitor of ceramide biosynthesis; and (d) was also evident in two patients with recurrent glioblastoma multiforme. THC inhibited MMP-2 expression and cell invasion in cultured glioma cells. Manipulation of MMP-2 expression by RNA interference and cDNA overexpression experiments proved that down-regulation of this MMP plays a critical role in THC-mediated inhibition of cell invasion. Cannabinoid-induced inhibition of MMP-2 expression and cell invasion was prevented by blocking ceramide biosynthesis and by knocking-down the expression of the stress protein p8. As MMP-2 up-regulation is associated with high progression and poor prognosis of gliomas and many other tumors, MMP-2 down-regulation constitutes a new hallmark of cannabinoid antitumoral activity.

Interplay of angiotensin II and angiotensin 1–7 in the regulations of matrix metalloproteinases of human cardiocytes

Angiotenisn 1–7 (Ang 1–7) is considered to play an opposing role to angiotensin II (Ang II). However, the modulation of MMPs and TIMPs by Ang 1–7 is largely unclear in cardiocytes, and the counteractive effects of Ang 1–7 on Ang II‐mediated MMPs and TIMPs expression need to be identified. We examined the mRNA expression of MMPs and TIMPs in human cardiac fibroblasts (HCF) and cardiac myocytes (HCM) stimulated by Ang II or Ang 1–7, and analyzed the antagonistic effects of Ang 1–7 to Ang II. Ang II decreased transcript expression of MMP‐1, MMP‐2, TIMP‐1, TIMP‐2 and TIMP‐3, but downregulated MMP‐9 expression in the HCF cells. Transcript expression of MMP‐9 and TIMP‐2 were downregulated by Ang 1–7 in the cells. In the HCM cells, Ang II induced MMP‐1 and MMP‐9 expression, but all of examined MMPs and TIMPs, except MMP‐9, were markedly decreased by Ang 1–7. Ang 1–7 could counteract the effects of Ang II‐mediated regulations on MMP‐1, MMP‐9 and TIMP‐1 in the HCF cells. The regulations of all examined MMPs by Ang II were reversed to basal expression by Ang 1–7 in the HCM cells. Opposite regulation in the examined MMPs and TIMPs expression was found between Ang II and Ang 1–7, and the antagonistic effects of Ang 1–7 to Ang II were also identified. These results might raise a potential role of Ang 1–7 to attenuate Ang II‐induced ECM remodeling in heart. This work was supported by National Science Council (NSC 95‐2313‐B‐009‐002‐MY3), Taiwan.

The change of matrix metalloproteinases-9 expression in newborn rabbits with oxygen inhalation and ventilator-induced lung injury

目的 观察新生免机械通气肺损伤对基质金属蛋白酶(MMP)-9表达的影响,研究肺损伤的分子机制.方法 将108只新生兔按2×3×3析因设计随机分配在高浓度氧组、低浓度氧组,每组又分为高吸气峰压、中吸气峰压、低吸气峰压共六组进行机械通气并与正常未通气组6只进行对照,通气后1、3、6 h 3个时间点处死取肺,行肺灌洗液细胞计数、沉渣涂片细胞分类,测定肺湿干重比、病理切片MMP-9免疫组织化学染色,ELISA法检测肺组织匀浆中MMP-9的浓度.结果 (1)高氧通气时肺组织MMP-9的表达低于低氧通气,差异有显著性(F=15.660,P=0.000),不同吸气峰压组间MMP-9的表达差异无显著性.(2)与对照组比,高吸气峰压组MMP-9有下调趋势,中、低吸气峰压组接近正常,随通气进行,MMP-9表达上调,以6 h表达最强.(3)MMP-9表达水平与灌洗液中白细胞计数、巨噬细胞数、肺湿干重比呈正相关.结论 新生免机械通气6 h内,高氧通气能下调MMP-9表达,机械拉伸对MMP-9表达的影响不大,MMP-9的表达与肺损伤相关。

Hydrogen peroxide-mediated downregulation of matrix metalloprotease-2 in indomethacin-induced acute gastric ulceration is blocked by melatonin and other antioxidants

Gastric mucosal damage is directly associated with extracellular matrix degradation in which matrix metalloproteinases (MMPs) play a crucial role. Remodeling of connective tissues and loss of tissue integrity due to the action of MMPs are reported in several inflammatory diseases, including gastric ulcer. Indomethacin-induced gastric ulceration involves the generation of reactive oxygen species (ROS) and a reduction in MMP-2 transcription and translation. Our aim was to identify the mechanism for suppression of MMP-2 activity by ROS during acute ulceration and further to examine the possible actions of antioxidants, especially melatonin, during healing. Melatonin ( N-acetyl-5-methoxytryptamine) blocked hydroxyl radical and nitrite anion generation, protein oxidation, mucosal cell disruption, and MMP-2 downregulation. In addition, suppression of MMP-2 activity by H 2O 2 in a dose- and time-dependent manner in vitro is blocked by melatonin, omeprazole, and curcumin. We observed that melatonin and other antioxidants (e.g., curcumin and omeprazole) offered gastroprotection in vivo by upregulation of suppressed MMP-2 expression and activity at the level of secretion and synthesis. Moreover, antioxidants reversed the suppression of MMP-2 expression by upregulation of MT1-MMP and downregulation of TIMP-2. Hence, we hypothesize that antioxidants exerted protection against H 2O 2-mediated inactivation and downregulation of MMP-2 expression during onset of indomethacin-induced ulceration.

Carboxy derivatized glucosamine is a potent inhibitor of matrix metalloproteinase-9 in HT1080 cells

Experimental evidences have confirmed that matrix metalloproteinases (MMPs) play a fundamental role in a wide variety of pathologic conditions and recent advances in medicinal chemistry approach to the design of MMP inhibitors with desired structural and functional properties. Among MMPs, MMP-9 has demonstrated to play a major role in the establishment of metastases and it is substantially increased in the majority of malignant tumors. Inhibition of MMP-9 is thought to have a therapeutic benefit to cancer. Results of this study present a novel synthetic MMP-9 inhibitor that downregulates MMP-9 expression level in HT1080, human fibrosarcoma cells.

Curcumin, a Potential Inhibitor of MMP-2 in Human Laryngeal Squamous Carcinoma Cells HEp2

Curcumin (diferuloylmethane) has been widely studied for its tumor inhibiting and anticarcino-genic properties. In the present communication, we studied the effect of curcumin on matrix-metalloproteinase-2 (MMP-2), integrin receptors, and focal adhesion kinase (FAK) in human laryngeal cancer cells, HEp2. HEp2 cells were treated with curcumin (5 microM) for 30 days and then grown without curcumin for 28 days. Effect of curcumin on MMP-2 expression and activity and on membrane type matrixmetalloproteinase-1 (MT1-MMP), FAK, and integrin receptors was studied by zymography, Western blot, ELISA, RT-PCR, and cell adhesion assay. Treatment of HEp2 cells with curcumin downregulated MMP-2 expression and activity and expression of integrin receptors, FAK, and MT1-MMP to almost background levels. MMP-2 (but not MMP-9) mRNA expression was abolished on curcumin treatment, indicating specific inhibition of MMP-2. Invasive potential of HEp2 cells was also significantly reduced. After drug withdrawal, expression of MMP-2, integrin receptors, MT1-MMP, and FAK returned to control levels. However, MMP-2 activity in serum free medium remained low. Downregulation of integrin receptors and low levels of FAK may hinder integrin-mediated signal transduction, preventing upregulation of MMP-2 activity. Reduction of MMP-2 activity and inhibition of HEp2 cell invasion by curcumin strongly indicate the potential of curcumin as an inhibitor of tumor cell invasion and metastasis.

Constitutive activation of the MAPkinase p38 is critical for MMP-9 production and survival of B-CLL cells on bone marrow stromal cells.

In the present work we investigated the role and biological significance of mitogen activated protein kinases (MAPK) in B-cell chronic lymphocytic leukaemia (B-CLL). The MAPK p38 was constitutively activated in B-CLL, but not in normal peripheral B cells. In addition, we demonstrated that the upstream kinases of p38, MKK3/6 were also constitutively activated in B-CLL cells. Furthermore, we determined by EMSA that the p38 MAP kinase pathway was not linked to the constitutive high expression of NF-kappaB, a critical survival factor of B-CLL cells. Recently, it has been shown that serum levels of angiogenic factors like VEGF, bFGF and MMP-9 are elevated in the serum of CLL patients and correlate with an unfavorable prognosis. We showed that the constitutive expression of MMP-9 was dependent on p38-activity and inhibition of p38 strongly downregulated MMP-9 expression. Coculture of B-CLL cells and stromal cells can protect spontaneous apoptosis of leukemic B cells. To determine the role of permanently activated p38 and MMP-9 expression, we cocultured B-CLL cells with bone marrow stromal cells. Survival of B-CLL cells on stroma was severely impaired when p38 was inhibited. Furthermore, blockade of MMP-9 activity also antagonized the antiapoptotic effect of stromal cells.

Molecular mechanism for activation and regulation of matrix metalloproteinases during bacterial infections and respiratory inflammation

Matrix metalloproteinases (MMPs) are critical mediators of tissue remodeling. Inappropriate regulation of MMPs causes many pathological events, including microbial invasion and inflammatory tissue damage. Some of the bacterial exoproteinases can effectively activate pro-MMPs (inactive zymogens) via limited proteolysis around their autoinhibitory domains. In addition, overproduction of nitric oxide (NO) may contribute to respiratory inflammation via the formation of reactive nitrogen species (RNS). Several studies have identified regulatory properties of NO/RNS on biomolecules due to functional modification of their cysteine residues. In fact, NO/RNS can mediate activation and expression of MMPs, because RNS can interact with a cysteine switch in the autoinhibitory domain, thus converting proMMPs into their active forms without proteolysis. Many studies have indicated that NO/RNS can participate in expression of various genes that affect immune-inflammatory responses, including MMPs. Although NO in some cases upregulates MMPs, S -nitrosothiols downregulate MMP-9 expression by suppressing the NF-kappaB pathway. While microbial proteinases cause excessive activation of MMPs and contribute to microbial pathogenesis, NO/RNS may modulate expression and activation of MMPs as well as various inflammatory mediators, depending on the redox status at sites of inflammation. Therefore, appropriate regulation of MMPs may be of potential therapeutic value for various infections and inflammatory lung diseases.