Correlation of matrix metalloproteinase－2, －9, tissue inhibitor－1 of matrix metalloproteinase and CD44 variant 6 in head and neck cancer metastasis

This study aimed to explore the molecular mechanism in tumor invasion and metastasis. The expression of matrix metalloproteinase 2, 9 (MMP 2, MMP 9), tissue inhibitor 1 of matrix metalloproteinase (TIMP 1), cell adhesion molecule 44 variant 6 (CD44v6), HER2/neu and p53 was investigated in 154 patients with head and neck squamous cell carcinoma (SCC) by ABC and ImmunoMax immunohistochemical method. Their clinical relevance and correlation were analysed. The expression of MMP 2, MMP 9, TIMP 1, CD44v6, HER2/neu and p53 was found in cancer cells in 87.01%, 85.71%, 68.18%, 98.05%, 55.19% and 50.65% cases respectively. Linear regression and correlation analysis revealed that there was close positive relationship ( P <0.05) between the expression of MMP 2 and MMP 9, TIMP 1 and CD44v6, HER2/neu and MMP 9, MMP 2 and p53. Up regulation of MMP 2 was accompanied by advanced T stage ( P <0.01) . There was also a trend of MMP 2 expression being related with tumor metastasis. Increased expression of HER2/neu was found in patients with tumor recurrence( P <0.05). The expression of TIMP 1 was higher in laryngeal cancer than that in pharyngeal cancer, and higher in keratinizing and non keratinizing SCC than that in basaloid SCC( P <0.05). These findings suggested that MMP 2 and MMP 9, HER2/neu and MMP 9, MMP 2 and p53 had a coordinate function in aggression of tumor; that MMP 2 had a more important function than MMP 9 in tumor invasion and metastasis; and that HER2/neu might serve as a biomarker for poor prognosis in HNSCC.

Imbalance of matrix metalloproteinase-9 and matrix metalloproteinase tissue inhibitor-1 may contribute to hemorrhage in cerebellar arteriovenous malformations

In this study, we determined the expression levels of matrix metalloproteinase-2 and -9 and matrix metalloproteinase tissue inhibitor-1 and -2 in brain tissues and blood plasma of patients undergoing surgery for cerebellar arteriovenous malformations or primary epilepsy （control group）. Immunohistochemistry and enzyme-linked immunosorbent assay revealed that the expression of matrix metalloproteinase-9 and matrix metalloproteinase tissue inhibitor-1 was significantly higher in patients with cerebellar arteriovenous malformations than in patients with primary epilepsy. The ratio of matrix metalloproteinase-9 to matrix metalloproteinase tissue inhibitor-1 was significantly higher in patients with hemorrhagic cerebellar arteriovenous malformations compared with those with non-hemorrhagic malformations. Matrix metalloproteinase-2 and matrix metalloproteinase tissue inhibitor-2 levels were not significantly changed. These findings indicate that an imbalance of matrix metalloproteinase-9 and matrix metalloproteinase tissue inhibitor-I, resulting in a relative overabundance of matrix metalloproteinase-9, might be the underlying mechanism of hemorrhage of cerebellar arteriovenous malformations.

Observation on In Situ Hybridization and Immunocytochemistry of Matrix Metalloproteinases in Rat Pancreas

In situ hybridization and immunocytochemical techniques were employed to examine the expression of matrix metalloproteinases 1 (MMP 1) and to identify the pattern of its distribution in rat pancreas. The results indicated that the signal of MMP 1 mRNA and MMP 1 positive immunoreaction were detected in some fiberoblasts around interlobular ducts and exocrine cell in margin acinus of some lobules, but the signal of MMP 1 mRNA and MMP 1 positive immunoreaction could not be detected in most of other acinus and islets of pancreas. It is concluded that the expression of MMP 1 in above cells of rat might play an important role in acinar proliferation and differentiation of rat pancreatic tissues.

Herbal compound 861 regulates mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells

AIM： To identify the role of herbal compound 861 （Cpd 861） in the regulation of mRNA expression of collagen synthesis- and degradation-related genes in human hepatic stellate cells （HSCs）. METHODS： mRNA levels of collagen types I and III, matrix metalloproteinase 1 （MMP-1）, matrix metalloproteinase 2 （MMP-2）, membrane type-1 matrix metalloproteinase （MT1-MMP）, tissue inhibitor of metalloproteinase 1 （TIMP-1）, and transforming growth factor β1 （TGF-β1） in cultured-activated HSCs treated with Cpd 861 or interferon-γ, （IFN-γ,） were determined by real-time PCR. RESULTS： Both Cpd 861 and IFN-γ reduced the mRNA levels of collagen type Ⅲ, MMP-2 and TGF-β1. Moreover, Cpd 861 significantly enhanced the MMP-1 mRNA levels while down-regulated the TIMP-1 mRNA expression, increasing the ratio of MMP-1 to TIMP-1 to （6.3 ＋ 0.3）- fold compared to the control group. CONCLUSION： The anti-fibrosis function of Cpd 861 may be mediated by both decreased interstitial collagen sythesis by inhibiting the transcription of collagen type Ⅲ and TGF-β1 and increased degradation of these collagens by up-regulating MMP-1 and down-regulating TIMP-1 mRNA levels.

金属蛋白酶-2和金属蛋白酶组织抑制物-1与糖尿病肾病关系的实验研究

目的 观察链脲佐菌素 (STZ)实验性糖尿病大鼠肾脏基质金属蛋白酶 2 (MMP 2 )及金属蛋白酶组织抑制物 1(TIMP 1)蛋白的表达及功能、形态学改变 ,探讨MMP 2、TIMP 1在糖尿病肾病(DN)发生机制中的意义。 方法 2 0只雄性Wistar大鼠随机分为糖尿病组和正常对照组 ,分别于第1、2、4、6、8周测定尿白蛋白排泄率 (UAER) ,第 9周用Western印迹方法检测MMP 2、TIMP 1蛋白表达水平 ,电镜观察肾小球基底膜厚度 (GBMT)。 结果 糖尿病组较正常组TIMP 1蛋白表达明显增加 ,MMP 2蛋白表达显著降低 (P <0 .0 1)。 4、6、8周UAER显著增加 ,GBMT明显增厚 (P <0 .0 1)。电镜发现糖尿病组肾小球基底膜弥慢性增厚 ,局部有系膜细胞插入和双轨征。 结论 持续高血糖可使大鼠肾脏MMP 2下降 ,TIMP 1增加。导致肾小球基底膜增厚 ,UAER增加。MMP 2、TIMP 1失衡可能对糖尿病肾病功能和形态学改变有重要意义。

Construction of Antisense MT1-MMP Vector and Its Inhibitory Effects on Invasion of Human Ovarian Cancer Cells

Membrane-type 1 matrix metalloproteinase （MT1-MMP/MMP-14） plays crucial roles in tumor cell growth, invasion, and angiogenesis. To clarify whether the endogenously expressed MT1-MMP in metastatic human ovarian carcinoma cell lines SKOV3 plays a critical role in tumor cell invasiveness, antisense MT1-MMP cloned in eukaryotic expression vector pMMP14as was transferred into SKOV3 cells. 48h after transfection, decreased expression of endogenous MT1- MMP protein was detected in pMMP14as-transfected SKOV3 cells and the activation of pro-MMP2 was inhibited markedly. The mean percentage of invasive cells was （62. 50 ± 5. 30） % in pMMP14as-transfected cells, which was obviously less than that （97.20±6.90） % in the control. Thus, antisense MT1-MMP effectively inhibited the endogenous MT1-MMP expression and the invasiveness in SKOV3 cells, suggesting that MT1-MMP may be a therapeutic target molecule for human invaslve ovarian cancers.

Effect of shRNA Inhibiting HIF1α Gene on TIMP1 Expression in RPE Cells

Small hairpin RNA （shRNA） was used to silence the HIF1α gene in human retinal pigment epithelial cells （RPE） under hypoxia in order to observe the effect of gene silencing on the expression of matrix metalloproteinase tissue inhibitor 1 （TIMP1）. By using chemical hypoxic inducer CoCl2 to mimic RPE hypoxic environment, shRNA against the targeting region of HIF1α mRNA sequence was synthesized by a method of in vitro transcription, and the HIF1α was interfered in RPE cultured under hypoxia （induced by 150 μmol/L CoCl2 ）. RT-PCR was employed to detect the expression of HIF1α and TIMP1. The expression levels of HIF1α and TIMP1 were measured by using Western blotting. The results showed that after the RPE were transfected with specific shRNA against HIF1α mRNA, RT PCR revealed that under hypoxia, the efficacy of HIF1α gene silencing in RPE was 83.4 %. Western blotting revealed that the expression levels of HIF1α protein was dramatically dropped. In addition, RT-PCR results demonstrated that the expression of TIMP1 mRNA was decreased by 28.9 %, and the expression levels of TIMP1 protein were also significantly reduced by Western blotting. It was suggested that shRNA targeted against HIF1α mRNA could effectively silence the HIF1α gene, subsequently effectively inhibit the hypoxia-induced up-regulation of TIMP1.

Effects of (-)-epigallocatechin-3-gallate on expression of matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 in fibroblasts irradiated with ultraviolet A

Background It is known that ultraviolet irradiation can affect cellular function through a number of signaling pathways ( ) epigallocatechin 3 gallate (EGCG) is the major effective component in green tea and can offer protection from ultraviolet induced damage In this study, we investigated the protective mechanism of EGCG on human dermal fibroblasts damaged by ultraviolet A (UVA) in vitro Methods Transcription factor Jun protein levels were measured by Western blot Matrix metalloproteinase 1 (MMP 1) and tissue inhibitor of metalloproteinase 1 (TIMP 1) mRNA were studied by reverse transcription polymerase chain reaction (RT PCR) analysis in conjunction with computer assisted image analysis MMP 1 and TIMP 1 proteins were quantified by enzyme linked immunosorbent assay (ELISA) Results EGCG decreased transcription activity of Jun protein after induction by UVA Both the mRNA and protein levels of MMP 1 were increased by UVA irradiation, while no significant changes were observed in TIMP 1 levels The ratio of MMP 1 to TIMP 1 showed statistically significant differences compared with the control EGCG decreased the ratio of MMP 1 to TIMP 1 by inhibiting UVA induced MMP 1 expression ( P <0 05) Conclusion EGCG can protect human fibroblasts against UVA damage by downregulating the transcription activity of Jun protein and the expression of MMP 1 The ratio of MMP 1 to TIMP 1, rather than the levels of MMP 1 or TIMP 1 alone, may play a significant role in human skin photodamage

The expression of matrix metalloproteinases-9, transforming growth factor-β1 and transforming growth factor-β receptorⅠ in human atherosclerotic plaque and their relationship with plaque stability

Background Transforming growth factor beta (TGF β) and matrix metalloproteinases 9 (MMP 9) have been implicated in the pathogenesis of human atherosclerosis but their relationship during lesion progression are poorly understood The objective of this study was to investigate the expression of MMP 9, TGF β1 and TGF β receptor Ⅰ (TβR Ⅰ) in human atherosclerotic plaque and their relationship and plaque stability Methods Specimens of human coronary artery atherosclerotic plaques were obtained from 41 patients undergoing coronary endarterectomy, and were paraffin embedded, sectioned at 4 μm intervals then stained with haematoxylin and eosin They were divided into stable (with no or only little lipid core) and unstable plaque groups (with lipid core size>40%): the immunohistochemical staining were performed for MMP 9,TGF β1 and TβR Ⅰ Results The expression of MMP 9 in the unstable plaques was much higher than in the stable ones, but the expression of TGF β1 was higher in the stable plaques There was no similar significant difference for TβR Ⅰ Correlation analysis showed that there was a negative correlation between the expression of MMP 9 and TGF β1 ( r =-0 332, P =0 034 for average areal density; r =-0 373, P = 0 016 for average optical density) Conclusions There were close relationships between MMP 9, TGF β1 and plaque stability Enhanced production of MMP 9 may participate in the formation of unstable plaque, while TGF β1 maybe an important stabilizing factor in preventing transition into an unstable plaque phenotype

Photoprotective effect of the N-terminal 5-mer peptide analog P165 of amyloid precursor protein in human dermal fibroblasts

Background We showed in our previous study that the N-terminal 17-mer peptide of amyloid precursor protein （APP17-mer peptide）,an active peptide segment with trophic and antioxidative effects,protects skin fibroblasts against ultraviolet （UV） damage and downregulates matrix metalloproteinase 1 （MMP-1） expression.The aim of the current study was to explore the protective effects of P165,the N-terminal 5-mer peptide analog of amyloid precursor protein that is resistant to enzymolysis,on UVA-induced damage in human dermal fibroblasts （HDFs）.Methods HDFs were cultured in Dulbecco＆#39;s modified Eagle＆#39;s medium without and with P165 （concentrations were 1,10,and 100 μJmol/L）.Then,15 J/cm2 UVA irradiation was used to obtain the UV-irradiated model.Cell proliferation was analyzed using MTT kit.The collagen type Ⅰ and MMP-1 contents in cell lysate were determined by enzyme-linked immunosorbent assay （ELISA）.Fluorometric assays were performed to detect the formation of intracellular reactive oxygen species （ROS） in the cells.Results P165 significantly protected the HDFs against UVA-induced cytotoxicity.Compared with the UVA-irradiated control,1,10,and 100 μmol/L P165 elevated cell proliferation by 14.98% （P〈0.05）,17.52% （P〈0.01） and 28.34% （P〈0.001）,respectively.Simultaneously,10 and 100 μmol/L P165 increased collagen type Ⅰ content （both P〈0.05）.Moreover,P165 treatment （all concentrations） also markedly suppressed the UVA-induced MMP-1 expression （all P〈0.001）.P165 at 1,10,and 100 μmol/L also reduced UVA-induced ROS generation by 11.27%,13.69% （both P〈0.05）,and 25.48% （P〈0.001）,respectively.Conclusions P165 could protect the HDFs against UVA-induced photodamage,including cytotoxicity,and MMP-1 generation.Furthermore,it also increased the collagen type Ⅰ content in the cells.The inhibitory effect on intracellular ROS generation might be involved in these photoprotective effects.Thus,P165 may be a useful candidate in the prevention and treatment of skin photoaging.

Effect of Bushenhuoxue formula on interleukin-1 beta and discoidin domain receptor 2 levels in a rat model of osteoarthritis

OBJECTIVE:To determine the effects of Bushenhuoxue formula(BHF) on interleukin-1 beta(IL-1β),transforming growth factor beta 1(TGF-β1),discoidin domain receptor 2(DDR2) and matrix metalloproteinase-1(MMP-1) levels in a rat model of osteoarthritis(OA).METHODS:Sprague-Dawley rats were used to establish an OA model and subjected to various treatments over 6 weeks.Rats were treated with BHF,glucosamine sulfate(G5),or starch as a control.Serum levels of IL-1β and MMP-1 and joint fluid levels of IL-1 β were determined by means of ELISAs.We used immunohistochemistry to determine DDR2 levels in knee cartilage.Gene expression levels of MMP-1 in joint synovial tissue were assessed using reverse transcription polymerase chain reaction assays.RESULTS:Serum IL-1β levels were unchanged throughout the study.Levels of IL-1β in joint fluid and MMP-1 in sera from the BHF- and GS-treated groups were significantly reduced.DDR2 levels in knee cartilage were also significantly reduced in the BHF group.Expression of the MMP-1 gene was significantly reduced by BHF treatment.CONCLUSION:BHF might be beneficial in the inhibition and alleviation of local inflammatory responses and cartilage degeneration in OA.

APP17肽通过抑制细胞内ROS保护紫外线照射后人皮肤成纤维细胞(英文)

Objective Ultraviolet light (UV) is known to cause photoaging of skin.UV irradiation can damage proliferation capacity and induce collagenase in fibroblasts in the dermis.Many researchers have explored the potential photo-protective agents;however,no ideal agent has been widely accepted.Amyloid precursor protein 17-mer peptide (APP17-mer peptide),an active peptide segment,has been reported to be responsible for the trophic effect in clonal CNS neuronal line,fibroblast cell line and HaCat cells.The aim of this study was to explore the effects of APP17-mer peptide on cultured fibroblasts after ultraviolet irradiation.Methods Human skin fibroblasts were cultured in DMEM medium with or without APP17-mer peptide (concentrations ranging from 20μmol/L,40 μmol/L,to 80μmol/L).The cultured fibroblasts were exposed to a single UV irradiation,and the proliferation activity of fibroblasts was detected by a MTT assay.The expression of matrix metalloproteinase-1 (MMP-1) mRNA was analyzed quantitatively following real-time RT-PCR.The generation of intracellular reactive oxygen species (ROS) was measured with fluorescent quantitation method.Results A single exposure to UV irradiation depressed proliferation activity of fibroblasts compared with sham-irradiated control (P<0.05).40μmol/L and 80μmol/L APP17-mer peptide increased the cellular proliferation activity in UV irradiated and unirradiated fibroblasts (P<0.05),however,20μmol/L did not show such protective effects (P>0.05).A single exposure of fibroblasts to UV irradiation resulted in 1.78 fold up-regulation of MMP-1 mRNA compared with unirradiated sample (P<0.05),and 40μmol/L and 80μmol/L APP17-mer peptide decreased the expression of MMP-1 mRNA (P<0.05 and P<0.01,respectively).UV irradiation increased generation of ROS in cultured fibroblasts (P<0.05).40μmol/L APP17-mer peptide inhibited the generation of ROS in irradiated fibroblasts.Conclusions APP17-mer peptide can enhance proliferation activity of fibroblasts after exposure to UV irradiation;it can also inhibit MMP-1 mRNA expression and ROS generation induced by UV irradiation.Inhibition of ROS generation after UV irradiation might be involved in the protective mechanism of APP17 peptide on proliferation activity and collagenase induction in UV-irradiated fibroblasts.

All-trans retinoic acid in pulmonary vascular structural remodeling in rats with pulmonary hypertension induced by monocrotaline

Objective To determine whether all-trans retinoic acid (atR A) exerts an inhibitory effect on rats with pulmonary hypertension induced by monocrotaline.Methods All rats were given a single subcutaneous injection of either monocrotaline (60 mg/kg) or saline.Monocrotaline-injected rats received either atRA (30 mg· kg-1· day-1) or saline through oral-gastro intubation. On Days 7, 14, 21, and 28 respectively after monocrotaline injection, cardiovascular catheters were inserted to examine the mean pulmonary artery pressure of rats in each group. Meanwhile, the matrix metalloproteinase-1 (MMP-1) mRNA expression and hydroxyproline content in the main pulmonary artery were determined by RT-PCR and chromornetry, respectively.Results The mean pulmonary artery pressure of rats in the model group increased significantly on day 21 and reached a peak on Day 28 compared with the control group (25.7+4.3 mm Hg vs 15.1 ± 1.5 mm Hg and 38.5 ± 6.4 mm Hg vs 16.4 ± 2.0 mm Hg, P ＜ 0.01 ). MMP-1 mRNA overexpression was present on Day 14 (0.72 ± 0.15 vs 0.39 ± 0.08, P ＜ 0.01 ) and was rapidly down-regulated on Day 21 and 28 compared with Day 14, but was still higher than that in the control. The hydroxyoroline content of the main pulmonary artery dropped significantly on Day 14 (4.01 ± 1.13 μg/mg vs 5.10 ± 0.91 μg/mg, P ＜ 0.05)and increased significantly on Days 21 and 28 compared with the control, atRA inhibited the MMP-1 mRNA overexpression from Day 14 to Day 28 and reduced the hydroxyproline content (5.59 ± 0.70 μg/mg vs 7.96 ± 1.13 μg/mg and 7.77 ± 0.96 μg/mg vs 9.93 ± 1.27μg/mg, P ＜ 0.01 ) and the mean pulmonary artery pressure compared with the model group ( 19.6 ± 3.2 mm Hg vs 25.7 ± 4.3 mm Hg and 26.3 ± 4.6 mm Hg vs 38.5±6.4 mm Hg, P＜0.01).Conclusion atRA inhibits MMP-1 overexpression and the accumulation of collagen, which might elicit favorable geometric remodeling in rat pulmonary hypertension induced by monocrotaline.