BACKGROUND: Matrix metalloproteinases (MMPs) have been implicated in the remodelling of extracellular matrix (ECM), including basement membrane. ECM remodelling is associated with pathological processes, including hepatic fibrosis, tumor invasion and metastasis. Tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 were known to inhibit MMP-9 and MMP-2, respectively. In the present study, we examined the expression of TIMP-1 and TIMP-2 in surgical specimen pairs of hepatocellular carcinoma and nontumoral liver and the correlation between their expression and clinicopathological characteristics. METHODS: The localization of both transcripts and protein of TIMP-1 and TIMP-2 was studied by using in situ hybridization and immunohistochemistry. RESULTS: TIMP-1 and TIMP-2 mRNA transcripts were found in tumor cells, hepatocyte, sinusoidal cells, endothelial cells and stromal cells. Signal intensity of TIMP-1 was stronger than that of TIMP-2. The results of immunohistochemical stainings were concordant with those obtained by in situ hybridization. Expression of TIMP-1 and TIMP-2 was observed in tumorous tissue, in nontumorous tissue and in the portions of the tumors adjacent to the capsules. However, a clear difference in TIMP-1 and TIMP-2 mRNA expression was not observed among the three tissue types. The intensity of TIMP-2 expression was generally weaker than that of TIMP-1, and the intensity of TIMP-1 and TIMP-2 mRNA expression did not correlate with variable clinicopathological characteristics. CONCLUSION: TIMPs was expressed in tumor cells and many cell types of the nontumoral liver. Further investigations for TIMPs' unknown functional role are needed.
Adult ; Aged ; Carcinoma, Hepatocellular/pathology ; Carcinoma, Hepatocellular/metabolism* ; Female ; Human ; Immunohistochemistry ; Liver Neoplasms/pathology ; Liver Neoplasms/metabolism* ; Male ; Middle Age ; RNA, Messenger/analysis ; Tissue Inhibitor-of Metalloproteinase-2/physiology ; Tissue Inhibitor-of Metalloproteinase-2/genetics* ; Tissue Inhibitor-of Metalloproteinase-2/analysis ; Tissue-Inhibitor of Metalloproteinase-1/physiology ; Tissue-Inhibitor of Metalloproteinase-1/genetics* ; Tissue-Inhibitor of Metalloproteinase-1/analysis

OBJECTIVEThis study aimed to investigate the effects of in vitro continuous passaging on the morphological phenotype and differentiation characteristics of mouse hyaline chondrocytes, as well as on the balance of the extracellular matrix (ECM).

METHODSEnzymatic digestion was conducted to isolate mouse hyaline chondrocytes, which expanded over five passages in vitro. Hematoxylin-eosin stain was used to show the changes in chondrocyte morphology. Semi-quantitative polymerase chain reaction was performed to analyze the mRNA changes in the marker genes, routine genes, matrix metalloproteinases (MMPs), and tissue inhibitors of MMPs (TIMPs) in chondrocytes. Zymography was carried out to elucidate changes in gelatinase activities.

RESULTSAfter continuous expansion in vitro, the morphology of round or polygonal chondrocytes changed to elongated and spindled shape. The expression of marker genes significantly decreased (P < 0.05), and it was almost negatively expressed by P5 chondrocytes. By contrast, the down regulation of routine genes was insignificant. The gene expression levels of MMPs and TIMPs both decreased (P < 0.05), but the change in MMP-1 and TIMP-1 was not statistically significant (P > 0.05). Meanwhile, the ratio of MMPs/TIMPs was altered. At the protein level, the activities of gelatinases decreased after passaging, especially for P4 and P5 chondrocytes (P < 0.05).

CONCLUSIONSerially passaged chondrocytes dedifferentiated and lost specific phenotypic characteristics during in vitro expansion culture. Simultaneously, the anabolism and catabolism of the cartilage ECM became uncontrollable and led to the imbalance of ECM homeostasis. When hyaline chondrocytes are applied in research on relevant diseases or cartilage tissue engineering, P0-P2 chondrocytes should be used.

Animals ; Cartilage ; Cell Differentiation ; Cells, Cultured ; Chondrocytes ; physiology ; Cytoskeleton ; Extracellular Matrix ; Gelatinases ; Gene Expression ; Hyalin ; physiology ; Matrix Metalloproteinase 1 ; Matrix Metalloproteinases ; Mice ; RNA, Messenger ; Tissue Engineering ; Tissue Inhibitor of Metalloproteinase-1 ; Tissue Inhibitor of Metalloproteinases

OBJECTIVETo investigate the roles of microfilaments in the expression of collagenase and TIMP-1 in normal and hypertrophic scar.

METHODSCell culture and Northern blot hybridization methods were used in this study.

RESULTSAfter the disruption of microfilaments with cytochalasin B, mRNA contents of collagenase and TIMP-1 both increased significantly in normal and hypertrophic scar fibroblasts.

CONCLUSIONThe result suggest that the microfilament cytoskeleton may regulate the synthesis and degradation of ECM, which may be a mechanism of gene regulation during wound healing.

Actin Cytoskeleton ; physiology ; Cells, Cultured ; Collagenases ; genetics ; Fibroblasts ; metabolism ; Humans ; RNA, Messenger ; analysis ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; Wound Healing ; physiology

OBJECTIVETo study the anti-invasion effect of SEPT7 gene on U251MG glioma cells and its possible molecular mechanism.

METHODSRecombinant adenovirus vector carrying SEPT7 gene (rAd5-SEPT7) was transduced to human glioma cell line U251MG, and empty adenovirus vector was used as control. Tumor invasion was examined by Transwell method and 3 D-Matrigel assay, and tumor cell migration by wound-healing method and 2 D-Matrigel assay. Three major molecular events associated with cell motility and migration, including changes of expression in MMP2, MMP9, MT1-MMP, TIMP1 and TIMP2, the alteration of integrin alpha(v)beta(3) expression, and the structural change of cytoskeleton protein, tubulin-alpha, in U251 cells transduced with rAd5-SEPT7 were studied by Western blotting, immunofluorescence and laser scanning confocal microscope, respectively.

RESULTSThe invasive and migratory capabilities of cells transduced with rAd5-SEPT7 were inhibited. The expression of extracellular matrix metalloproteinases MMP-2, MMP-9, MT1-MMP and integrin alpha(v)beta(3) was significantly decreased, while the expression of matrix metalloproteinase inhibitor TIMP1, TIMP2 was upregulated. Intracellular cytoskeleton protein-tubulin-alpha in U251 cells exhibited prominent morphological changes which including the appearance of distortion and aggregation resulting from redistribution of tubulin-alpha, and this feature of alteration was similar to the tubulin-alpha structure in normal non-tumor cells.

CONCLUSIONSEPT7 gene can inhibit the invasion and migration ability of U251 glioma cells. Its molecular mechanism may include that SEPT7 gene reverses the imbalanced state of MMPs/TIMPs, downregulates the expression of integrin alpha(v)beta(3) and alters the structure of tubulin-alpha of U251MG glioma cells. It is suggested that SEPT7 gene could be a good candidate for gene therapy of gliomas.

Adenoviridae ; genetics ; Blotting, Western ; Cell Cycle Proteins ; genetics ; physiology ; Cell Line, Tumor ; Cell Movement ; genetics ; Genetic Vectors ; genetics ; Glioma ; metabolism ; pathology ; Humans ; Integrin alphaVbeta3 ; metabolism ; Matrix Metalloproteinase 14 ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Microscopy, Confocal ; Neoplasm Invasiveness ; genetics ; Septins ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism ; Tissue Inhibitor of Metalloproteinase-2 ; metabolism

BACKGROUNDAs a novel molecular markerof non-small cell lung cancer (NSCLC), PRDI-BF1 and RIZ homology domain containing protein 14 (PRDM14) is over-expressed in NSCLC tumor tissues. Extracellular matrix degradation mediated by the balance between matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) is one of the most important mechanism in lung cancer metastasis. This study aimed to determine if PRDM14 promoted the migration of NSCLC cells through extracellular matrix degradation mediated by change of MMP/TIMP expression.

METHODSThe expression of PRDM14 was down-regulated in human cell line A 549 after transfection with lentiviral vector-mediated short-hairpin ribonucleic acids (shRNAs) which targeted the PRDM14 promoter. Cellular migration of shRNA-infected cells was detected by a scratch wound healing assay and transwell cell migration assay. Expression levels of MMP1, MMP2, TIMP1, and TIMP2 were measured by quantitative real-time polymerase chain reaction (RT-PCR).

RESULTSMigration of PRDM14-shRNA-infected cells was significantly inhibited relative to control cells as measured by the scratch wound healing (P < 0.05) and transwell cell migration assays (P < 0.01). The expression of MMP1 in A549 cells infected by PRDM14-shRNA was down-regulated significantly (P < 0.01), whereas the expression of TIMP1 and TIMP2 was up-regulated significantly (P < 0.01).

CONCLUSIONSPRDM14 accelerates A549 cells migration in vitro through extracellular matrix degradation. PRDM14 is considered as a potential therapeutic target in metastatic NSCLC.

Carcinoma, Non-Small-Cell Lung ; metabolism ; Cell Line, Tumor ; Cell Movement ; genetics ; physiology ; Extracellular Matrix ; metabolism ; Humans ; Matrix Metalloproteinase 1 ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; Neoplasm Metastasis ; genetics ; Repressor Proteins ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism ; Tissue Inhibitor of Metalloproteinase-2 ; metabolism

OBJECTIVETo study the effect of the cultured supernatant of human silicotic alveolar macrophages (AM) on the expression of matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in human lung fibroblasts (FB).

METHODSHuman alveolar macrophages were collected from a silicotic patients by bronchoalveolar lavage and exposed to SiO(2), then the cultured supernatant were incubated with human fetal lung fibroblasts for 6, 12, 18, 24, 36, 48 h. The immunocytochemical method was used to detect the level of expression of MMP-1 and TIMP-1 in lung fibroblasts.

RESULTSThe expression of MMP-1 in FB in 24 h incubation was lower in cultured supernatant of silicotic AM unexposed to SiO(2) than in blank control [integrated OD (IOD)]: 0.103 +/- 0.014 vs 0.133 +/- 0.023), while the expression of TIMP-1 was higher (IOD: 0.108 +/- 0.012 vs 0.065 +/- 0.006). The expression of MMP-1 in FB in cultured supernatant of AM exposed to SiO(2) for 24 h was further decreased (IOD: 0.062 +/- 0.008 vs 0.133 +/- 0.023), while that of TIMP-1 was further increased (IOD: 0.143 +/- 0.015 vs 0.065 +/- 0.006).

CONCLUSIONSiO(2) may affect the expression of MMP-1 and TIMP-1 system through AM mediation and participate in the formation of lung fibrosis.

Cells, Cultured ; Coculture Techniques ; Fibroblasts ; drug effects ; metabolism ; Humans ; Lung ; cytology ; Macrophages, Alveolar ; physiology ; Male ; Matrix Metalloproteinase 1 ; metabolism ; Middle Aged ; Silicon Dioxide ; pharmacology ; Silicosis ; pathology ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism

OBJECTIVEExtracellular matrix (ECM) deposition is a major reason of pulmonary fibrosis in hyperoxia-induced lung injury. However, the relevant mechanism has not been identified. This study examined the gene expressions of matrix metalloproteinases-8 (MMP-8, a catabolic enzyme of type I collagen) and tissue inhibitor of metalloproteinases-1 (TIMP-1) in neonatal rats with hyperoxia-induced pulmonary injury in order to explore the role of MMP-8 and TIMP-1 in pulmonary fibrosis.

METHODSEighty term newborn rats were randomly exposed to hyperoxia (FiO2=0.90, hyperoxia group)and to room air (FiO2=0.21, control group)(n=40 each). Lung injury was induced by hyperoxia exposure. The content of type I collagen and the expressions of type I collagen protein and MMP-1 mRNA and TIMP-1 mRNA were assayed with enzyme linked immunoadsorbent (ELISA), immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) respectively on days 1, 3, 7, 14 and 21 after exposure.

RESULTSThe content of type I collagen and the expression of type I collagen protein in the hyperoxia group were statistically higher than those in the control group at 14 and 21 days post-exposure. The MMP-8 mRNA expression decreased while the TIMP-1 mRNA expression increased significantly in the hyperoxia group as compared to the control group at 14 and 21 days post-exposure.

CONCLUSIONSHyperoxia exposure down-regulates MMP-8 mRNA expression and up-regulates TIMP-1 mRNA expression. This results in a reduction of ECM degradation, thereby ECM deposition occurs in lung tissue, which may be an important mechanism of pulmonary fibrosis following hyperoxia-induced lung injury.

Animals ; Animals, Newborn ; Chronic Disease ; Collagen Type I ; analysis ; genetics ; Female ; Hyperoxia ; complications ; Male ; Matrix Metalloproteinase 8 ; genetics ; physiology ; Pulmonary Fibrosis ; etiology ; RNA, Messenger ; analysis ; Rats ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; physiology

PURPOSE: To investigate the changes and correlations of the serum inflammation factors levels and left ventricular (LV) structure and function in patients with acute ST segment elevation myocardial infarction (STEMI). MATERIALS AND METHODS: A prospective study was performed on 70 STEMI patients and 70 control subjects. Serum levels of interleukin-6 (IL-6), soluble CD40 ligand (sCD40L), metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) were measured by sandwich enzyme-linked immunosorbent assay (ELISA), and cardiac structure and function were assessed by echocardiography at admission and 3-year follow-up. RESULTS: We found that the levels of serum IL-6, sCD40L and MMP-9 increased steadily among control subjects, remote myocardial infarction and acute STEMI patients, and the level of TIMP-1 elevated remarkly at 3-year follow-up visit in STEMI. The admission level of serum MMP-9 positively correlated with LV end-diastolic and end-diastole volume (r=0.294, p=0.022; r=0.269, p=0.036, respectively), and TIMP-1 positively correlated with E/A ratio (r=0.278, p=0.044) at 3-year follow-up. CONCLUSION: The study indicates that admission levels of serum MMP-9 and TIMP-1 closely correlated with left ventricular structure and function, which may be involved in the process of post-infarction remodeling of myocardium.
Aged ; CD40 Ligand/blood ; Female ; Humans ; Interleukin-6/blood ; Male ; Matrix Metalloproteinase 9/blood ; Middle Aged ; Myocardial Infarction/*blood/*physiopathology ; Prospective Studies ; Tissue Inhibitor of Metalloproteinase-1/blood ; Ventricular Remodeling/*physiology

OBJECTIVETo study the influence of hepatocyte growth factor (HGF) antibody on the lung expression level of matrix metalloproteinases-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1).

METHODSThirty male Wistar rats were randomly divided into 3 groups: control group, model group, and intervention group. Endotoxin was intratracheally infused in the model and intervention groups. HGF antibody was injected in the rats of the intervention group from day 1 to day 14, while the same volume of saline was injected in the control group. The rats were sacrificed on day 28 after endotoxin treatment. The amounts of MMP-9 mRNA and TIMP-1 mRNA were measured by reverse transcription-polymerase chain reaction, and protein expression levels of MMP-9 and TIMP-1 were measured by immunohistochemistry.

RESULTSIn the model group, both mRNA and protein expression levels of TIMP-1 were significantly increased, the same as MMP-9. In the intervention group, the increase of TIMP-1 was remarkably reduced compared with the model group, while the mRNA and protein expression levels of MMP-9 were still increased.

CONCLUSIONHGF activity may accelerate the repair of lung injury through contrary regulating the expression levels of TIMP-1 and MMP-9.

Acute Lung Injury ; metabolism ; pathology ; Animals ; Antibodies ; immunology ; Hepatocyte Growth Factor ; physiology ; Male ; Matrix Metalloproteinase 9 ; analysis ; genetics ; RNA, Messenger ; analysis ; Rats ; Rats, Wistar ; Tissue Inhibitor of Metalloproteinase-1 ; analysis ; genetics

OBJECTIVETo study the effect of laminarin polysaccharide (LP) on the activity of matrix metalloproteinase of photoaging skins.

METHODKunming SPF mice were prepared with back hair shaved, and randomly divided into the control group, the model group, the LP low does group (LP-L, 1 mg x kg(-1)), the LP high dose group (LP-H, 5 mg x kg(-1)) and the Vit E (100 mg x kg(-1)) group. They were abdominally injected with drugs twice on a daily basis. Except for the control group, all groups were exposed to ultraviolet rays for 1 hour every day, five times on a weekly basis, with accumulated exposure dose of UVB being 21.60 J x cm(-2) and accumulated exposure dose of UVA being 84.02 J x cm(-2). Eight weeks later, exposed back skins were collected to detect thickness of dermis by HE stain, content of hydroxyproline (Hyp) by chemical colorimetry, and serum MMP-1 and TIMP-1 content by ELISA. In addition, matrix metalloproteinase-1 (MMP-1) mRNA and relative content of tissue inhibitor of metalloproteinase-1 (TIMP1) mRNA was analyzed with Real-time PCR.

RESULTCompared with the model group, the LP-H group could significantly increase the thickness of dermis, skin Hyp content and serum TIMP-1 level, and decrease relative content of MMP-1 mRNA in skin and MMP-1 content in serum.

CONCLUSIONLP can regulate the metabolism of collagen photoaging skins by adjusting the activity of matrix metalloproteinase.

Animals ; Female ; Glucans ; Matrix Metalloproteinase 13 ; biosynthesis ; genetics ; metabolism ; Mice ; Plant Extracts ; chemistry ; pharmacology ; Plants, Medicinal ; chemistry ; Polysaccharides ; chemistry ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Skin Aging ; drug effects ; physiology ; radiation effects ; Tissue Inhibitor of Metalloproteinase-1 ; biosynthesis ; genetics ; metabolism ; Ultraviolet Rays