OBJECTIVETo investigate the effects of adenovirus-delivered angiopoietin-1 siRNA (Ad. Ang-1siRNA) on the expression of matrix metalloproteinase-2, 9 (MMP-2, 9) and tissue inhibitor of metallopro-teinase-1 (TIMP-1) in rats with acute lung injury (ALI) induced by phosgene (Psg).

METHODSWe first established a rat model of Psg-induced acute lung injury (ALI). The rats were randomly divided into 6 groups: air control group with exposure to air, air+adenovirus (air+Ad) group with caudal vein injection of 1×10(8) pfu/ml adenovirus 1 h after air exposure, air+Ad/Ang1 group with caudal vein injection of 1×10(8) pfu/ml Ad.Ang-1siRNA 1 h after air exposure, Psg group with exposure to 8.33 mg/L Psg (purity 100%, of the same volume as the inhaled air in the air control group) for 5 min, Psg+Ad group with caudal vein injection of 1×10(8) pfu/ml adenovirus 1 h after exposure to the same dose of Psg, and Psg+Ad/Ang1 group with caudal vein injection of 1×10(8) pfu/ml Ad.Ang-1siRNA 1 h after exposure to the same dose of Psg. Serum, bronchoalveolar lavage fluid (BALF), and lung tissue were collected 36 h after exposure. The protein expression of Ang-1, MMP-2, 9, and TIMP-1 in serum and BALF was determined by double-antibody sandwich ELISA. RT-PCR was used to determine the mRNA levels of Ang-1, MMP-2, 9, and TIMP-1 in lung tissue. The protein expression of MMP-2, 9 and TIMP-1 in lung tissue was determined by Western blot.

RESULTSA rat model of Psg-induced ALI was successfully established. The levels of MMP-2, 9 in serum, BALF, and lung tissue were significantly increased in the Psg group and Psg+Ad/Ang1 group as compared with the control group (P<0.01); no significant change was observed in serum TIMP-1 protein expression (P>0.05); interestingly, TIMP-1 protein expression in BALF and lung tissue was significantly increased (P<0.01). Compared with the Psg group, the Psg+Ad/Ang1 group showed a significant decrease in MMP-2, 9 expression in BALF, serum, and lung tissue (P<0.05), but no significant change in protein expression of TIMP-1 was discovered (P>0.05).

CONCLUSIONAd.Ang-1siRNA has a potential beneficial effect in rats with Psg-induced ALI through inhibition of MMP-2, 9 expression, but has no significant effect on the expression of TIMP-1.

Acute Lung Injury ; chemically induced ; metabolism ; Adenoviridae ; genetics ; Angiopoietin-1 ; physiology ; Animals ; Bronchoalveolar Lavage Fluid ; Chemical Warfare Agents ; toxicity ; Disease Models, Animal ; Lung ; metabolism ; Matrix Metalloproteinase 2 ; genetics ; Matrix Metalloproteinases ; metabolism ; Phosgene ; toxicity ; RNA, Messenger ; genetics ; RNA, Small Interfering ; Rats ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism

OBJECTIVETo observe P38 mitogen-activated protein kinase (P38 MAPK) and matrix metalloproteinase-2 (MMP-2) mRNA expression level changes in neonatal rats with hyperoxia-induced lung injury,and to investigate the influence of P38 MAPK activation on MMP-2 mRNA expression.

METHODSThirty-six Sprague-Dawley (SD) rats were randomly divided into three groups: air control, hyperoxia and SB203580-treated hyperoxia (n=12). The rats were sacrificed on the 3rd and 7th days and the lungs were removed. Hematoxylin-eosine staining was used to observe the pathological changes in lung tissues.

RESULTSCompared with the air and SB203580-treated groups, levels of P38 MAPK and MMP-2 mRNA significantly increased in the hyperoxia group (P<0.01).

CONCLUSIONSExpression of P38 MAPK increases in neonatal rats with hyperoxia-induced acute lung injury and this may play a role in control of the expression of MMP-2 mRNA.

Animals ; Animals, Newborn ; Female ; Hyperoxia ; complications ; Lung ; pathology ; Lung Injury ; metabolism ; Male ; Matrix Metalloproteinase 2 ; genetics ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Tissue Inhibitor of Metalloproteinases ; genetics ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo study the relationship between atrial structural remodeling and the expressions of matrix metalloproteinase (MMPs) and their tissue inhibitors (TIMPs) in atrial fibrillation (AF).

METHODSBiopsy samples of the right atrial appendages were collected from 20 patients with sinus rhythm and 30 with AF undergoing heart valve replacement surgery for rheumatic heart diseases. All the patients received echocardiographic examination preoperatively. MMP-1, -3, -7, -9 and TIMP-1, -2, -3, -4 protein expressions were detected by immunohistochemistry and RT-PCR.

RESULTSCompared with those in patients with sinus rhythm, the AF patients had significantly increased left and right atrial diameters and mRNA levels of MMP-3, -7, -9 and TIMP-1, -2, -3, -4 (P<0.01). MMP-1 expression also showed an increase in AF patients, but the difference was no statistically significant from that in patients with sinus rhythm.

CONCLUSIONThe expressions of MMP-1, -3, -7, -9 and TIMP-1, -2, -3, -4 increase in fibrillating atrial tissue, which may contribute to atrial structural remodeling and atrial dilatation in AF patients.

Adult ; Aged ; Atrial Fibrillation ; enzymology ; pathology ; physiopathology ; Female ; Humans ; Male ; Matrix Metalloproteinases ; genetics ; metabolism ; Middle Aged ; Tissue Inhibitor of Metalloproteinases ; genetics ; metabolism

Hepatic fibrogenesis, a complex process that involves a marked accumulation of extracellular matrix components, activation of cells capable of producing matrix materials, cytokine release, and tissue remodeling, is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). The MMP-TIMP balance can regulate liver fibrogenesis. The aim of this study was to evaluate the expression patterns of MMPs and TIMPs during thioacetamide (TAA)-induced liver fibrogenesis. Chronic liver injury was induced with TAA (200 mg/kg i.p.) for 4 or 7 weeks in male Sprague-Dawley rats. Hepatic injury and fibrosis were assessed by hematoxylin-eosin (H&E) staining, and collagen deposition was confirmed by Sirius Red staining. The level of hepatic injury was quantified by serological analysis. The transcriptional and translational levels of alpha-smooth muscle actin (alpha-SMA), MMPs, and TIMPs in the liver were measured by Western blotting, RT-PCR, and immunohistochemistry. MMP, TIMP, and alpha-SMA were observed along fibrotic septa and portal spaces around the lobules. TAA treatment increased transcription of both MMPs and TIMPs, but only TIMPs showed increased translation. The dominant expression of TIMPs may regulate the function of MMPs to maintain liver fibrosis induced by TAA.
Animals ; Collagen/metabolism ; Extracellular Matrix/chemistry/metabolism ; *Liver Cirrhosis/chemically induced/metabolism/pathology ; Male ; Matrix Metalloproteinases/genetics/*metabolism ; Rats ; Rats, Sprague-Dawley ; Thioacetamide/*toxicity ; Tissue Inhibitor of Metalloproteinases/genetics/*metabolism

OBJECTIVEThe effect of triptolide on the DNA methylation level of MMP-9 gene and the mRNA expression of tissue inhibitors of met-alloproteinases (TIMPs) were examined in human fibrosarcoma HT-1080 cells to explore the molecular mechanisms involved in the anticancer activity of triptolide.

METHODHT-1080 cells were cultured in MEM containing 10% newborn calf serum and 1% penicillin-streptomycin. Triptolide was dissolved in dimethyl sulfoxide (DMSO) at a concentration of 1 goL-1 and stored at -20 degrees C. Triptolide was freshly diluted with culture medium perior to use and directly added to cell cultures at the indicated concentration, and incubated for 72 hours at 37 degrees C in a humidified atmosphere with 5% CO2, with changes of reagents every 24 hours. Methylation specific PCR (MSP)was applied to assess the methylation status of MMP-9 gene promoter, and semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) was employed to measure the mRNA expression of tissue inhibitors of metalloproteinases (TIMPs) in human fibrosarcoma HT-1080 cells after 72 hours of treatment with 6 nmol x L(-1), 12 nmol x L(-1) or 18 nmol x L(-1) triptolide, respectively.

RESULTSThe methylation index of MMP-9 gene promoter was statistically elevated in HT-1080 cells after 72 hours of treatment with 18 nmol L(-1) triptolide, compared with those in controls (0.61 +/- 0.10 vs 0.39 +/- 0.10, P < 0.05), while no significant difference was noted between 6 nmol x L(-1) or 12 nmol x L(-1) triptolide treated HT-1080 cells and controls (0.40 +/- 0.15 vs 0.39 +/- 0.10, 0.46 +/- 0.20 vs 0.39 +/- 0.10, respectively, both P > 0.05). The mRNA expression of TIMP-1, -2, -3 or -4 was not significantly changed in HT-1080 cells after 72 hours of treatment with the indicated concentrations of triptolide, respectively compared with those in controls (all P > 0.05).

CONCLUSIONThe results demonstrated that triptolide upregulates the methylation level of MMP-9 gene in HT-1080 cells in vitro.

Antineoplastic Agents, Alkylating ; pharmacology ; Cell Line, Tumor ; DNA Methylation ; drug effects ; Diterpenes ; pharmacology ; Epoxy Compounds ; pharmacology ; Fibrosarcoma ; drug therapy ; genetics ; metabolism ; Gene Expression Regulation, Enzymologic ; drug effects ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Phenanthrenes ; pharmacology ; Tissue Inhibitor of Metalloproteinases ; genetics ; metabolism

AIMThe changes of tissue inhibitor of metalloproteinase-4 (TIMP-4) expression in mouse ovary during pregnant and postpartum period were studied to investigate the role of TIMP-4 in corpus luteum (CL).

METHODSRT-PCR was used to deter mine the change of TIMP-4 mRNA and indirect immunofluorescence was used to observe the change of TIMP-4 protein. The expression of TIMP-4 mRNA was observed in various periods throughout the stage of pregnancy and postpartum day 1.

RESULTSThe expression of TIMP-4 was gradually enhanced from day 1 to day 8, reached a maximal expression at day 8, while decreased at day 11 and to the lowest level at postpartum day 1. Indirect immunofluorescence results further indicated that TIMP-4 protein was localized to CL and theca-intera cells in various periods throughout the pregnancy and postpartum day 1. In addition, the change pattern of TIMP-4 protein agreed with that of the TIMP-4 mRNA in pregnancy CL.

CONCLUSIONThe expression of TIMP-4 in mouse ovary during pregnancy and postpartum is in spatio-temporal pattern and it may be involved in the formation and function maintain of CL during pregnancy in mice.

Animals ; Female ; Mice ; Mice, Inbred Strains ; Ovary ; metabolism ; Postpartum Period ; Pregnancy ; RNA, Messenger ; genetics ; Tissue Inhibitor of Metalloproteinases ; metabolism

OBJECTIVETo investigate the effect of asiaticoside on the expressions of transforming growth factor (TGF)-beta mRNA, matrix metalloproteinases (MMPS) and tissue inhibitors of metalloproteinases (TIMPs) in postburn hypertrophic scars.

METHODSNine specimens of postburn (5-8 months) hypertrophic scars with asiaticoside treatment and 9 without asiaticoside treatment were collected for testing the expressions of MMPS, TIMPs, type I and III collagen and TGF-beta mRNA by immunohistochemistry and in situ hybridization methods, followed by image analysis of the results.

RESULTSThe expressions of TGF-beta mRNA and MMPS/TIMPS were all detected in the fibroblast cytoplasm. The expression of TGF-beta(1) mRNA in asiaticoside-treated scars was significantly lower than that in scars without asiaticoside treatment (P<0.01). In contrast, the expression of TGF-beta(3) mRNA was significantly higher in asiaticoside group (P<0.05). The expression of TIMP1 in asiaticoside group was significantly lower than that in non-asiaticoside group (P<0.01), and the expression of type I collagen in asiaticoside-treated scars was lower than that in non-asiaticoside-treated specimens (P<0.05), but the expression of MMP(1), MMP(2) and TIMP(2) and type III collagen exhibited no significant differences between the two groups (P>0.05).

CONCLUSIONAsiaticoside can down-regulate TGF-beta(1) mRNA and TIMP(1) expressions and up-regulate TGF-beta(3) mRNA expression in postburn hypertrophic scars, and is also capable of decomposing the products of type I collagen, contributing to the reduction of hypertrophic scar formation.

Anti-Infective Agents ; therapeutic use ; Burns ; complications ; Cicatrix, Hypertrophic ; etiology ; metabolism ; Collagen Type I ; biosynthesis ; genetics ; Female ; Humans ; Male ; Matrix Metalloproteinases ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Skin ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; biosynthesis ; genetics ; Transforming Growth Factor beta ; biosynthesis ; genetics ; Triterpenes ; therapeutic use

OBJECTIVETo study the relationship between activation of pro-MMP-2 and expression of matrix metalloproteinases (MMP)-2, MT1-MMP and tissue inhibitor of metalloproteinases (TIMP)-2 mRNA in thymoma and thymic carcinoma; and to study the molecular mechanism of invasion and metastasis of thymic epithelial tumors.

METHODSFresh tissue specimens of thymoma, thymic carcinoma and normal thymus were included. The mRNA expression of MMP-2, MT1-MMP and TIMP-2 were analyzed by real-time reverse transcription polymerase chain reaction. The pro-MMP-2 activation ratio and its localization were determined by gelatin zymography and film in-situ gelatin-Zymography, respectively. Correlation of mRNA expression of MMP-2, MT1-MMP and TIMP-2 was investigated in tumors with different histological subtypes and clinical stages.

RESULTSThere were no significant differences in the expressions of MMP-2, MT1-MMP and TIMP-2 mRNA between I and II stage or III and IV stage thymomas (P > 0.05). However, significant differences of the expressions were observed between three tumor groups: I-II stage, III-IV stage and thymic carcinomas (P < 0.005), and between three histological subtypes: AB-B1 (lymphocyte-rich and mixed types), B2-B3 (cortical and predominantly polygonal cells types) and thymic carcinomas (P < 0.05). Expression levels of MT1-MMP and TIMP-2 mRNA were correlated with pro-MMP-2 activation ratio (Spearman rank correlation: r = 0.7235, r = 0.7647, P < 0.005). The expression of MMP-9 did not show significant differences between thymomas and thymic carcinomas.

CONCLUSIONSMMP-2, MT1-MMP and TIMP-2 mRNA expression levels are correlated with the histologic subtypes and clinical stages of thymoma. The mRNA expressions of MT1-MMP and TIMP-2 are correlated with the activation ratio of pro-MMP-2. It is speculated that upregulation of MT1-MMP gene expression may induce an activation of pro-MMP-2 through TIMP-2.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Squamous Cell ; enzymology ; metabolism ; pathology ; Enzyme Activation ; Female ; Humans ; Male ; Matrix Metalloproteinase 2 ; biosynthesis ; genetics ; Matrix Metalloproteinases ; biosynthesis ; genetics ; Matrix Metalloproteinases, Membrane-Associated ; Middle Aged ; Neoplasm Staging ; RNA, Messenger ; biosynthesis ; genetics ; Thymoma ; classification ; enzymology ; metabolism ; pathology ; Thymus Gland ; enzymology ; metabolism ; Thymus Neoplasms ; classification ; enzymology ; metabolism ; pathology ; Tissue Inhibitor of Metalloproteinase-2 ; biosynthesis ; genetics

We investigated the expression of membrane type-1 (MT1)-MMP, MMP2, MMP9 and TIMP2 mRNAs and their roles in ductal carcinoma in situ (DCIS) and T1 and T2 invasive ductal carcinoma of the breast. We further compared these two types of carcinomas for differences in microvessel density, and expression of angiogenic factors and CD44std. MT1-MMP, MMP2, MMP9 and TIMP2 mRNA were expressed in both DCIS and invasive ductal carcinomas. Expression rates of MT1-MMP, MMP2, MMP9 and TIMP2 mRNAs were not statistically different between DCIS and invasive ductal carcinomas, nor did they differ statistically when grouped by tumor size, histologic grade or nuclear grade of invasive ductal carcinoma. Microvessel density and expression of VEGF and TGF-beta were not statistically different between DCIS and invasive ductal carcinoma. CD44std expression was significantly increased in DCIS compared to invasive ductal carcinoma (p < 0.05) and it was also significantly increased in lower clinical stage, histologic grade and nuclear grade of invasive ductal carcinoma (p < 0.05). Axillary node metastasis was significantly correlated with MT1-MMP mRNA, VEGF and TGF-beta expression (p < 0.05) and MT1-MMP mRNA was positively correlated with VEGF expression and TIMP2 mRNA (p < 0.05). In summary, patterns of MMP mRNA expression in DCIS and invasive ductal carcinoma suggest that the invasive potential of breast carcinoma is already achieved before morphologically overt invasive growth is observed. As MT1-MMP mRNA expression is significantly correlated with axillary nodal metastasis, it may be useful as a prognostic indicator of invasive ductal carcinoma. Considering the positive correlation of MT1-MMP mRNA and TIMP2mRNA expression, our finding supports a role for TIMP2 in tumor growth, as well as the utility of CD44std as a prognostic indicator of breast cancer.
Tissue Inhibitor of Metalloproteinase-2/genetics ; RNA, Messenger/metabolism ; Matrix Metalloproteinases, Membrane-Associated ; Matrix Metalloproteinases/*genetics ; Matrix Metalloproteinase 9/genetics ; Matrix Metalloproteinase 2/genetics ; Matrix Metalloproteinase 1/genetics ; Humans ; Gene Expression Regulation, Neoplastic ; Gene Expression Regulation, Enzymologic ; Female ; Carcinoma, Ductal, Breast/genetics/*physiopathology ; Carcinoma in Situ/genetics/*physiopathology ; Breast Neoplasms/genetics/*physiopathology

We investigated the expression of membrane type-1 (MT1)-MMP, MMP2, MMP9 and TIMP2 mRNAs and their roles in ductal carcinoma in situ (DCIS) and T1 and T2 invasive ductal carcinoma of the breast. We further compared these two types of carcinomas for differences in microvessel density, and expression of angiogenic factors and CD44std. MT1-MMP, MMP2, MMP9 and TIMP2 mRNA were expressed in both DCIS and invasive ductal carcinomas. Expression rates of MT1-MMP, MMP2, MMP9 and TIMP2 mRNAs were not statistically different between DCIS and invasive ductal carcinomas, nor did they differ statistically when grouped by tumor size, histologic grade or nuclear grade of invasive ductal carcinoma. Microvessel density and expression of VEGF and TGF-beta were not statistically different between DCIS and invasive ductal carcinoma. CD44std expression was significantly increased in DCIS compared to invasive ductal carcinoma (p < 0.05) and it was also significantly increased in lower clinical stage, histologic grade and nuclear grade of invasive ductal carcinoma (p < 0.05). Axillary node metastasis was significantly correlated with MT1-MMP mRNA, VEGF and TGF-beta expression (p < 0.05) and MT1-MMP mRNA was positively correlated with VEGF expression and TIMP2 mRNA (p < 0.05). In summary, patterns of MMP mRNA expression in DCIS and invasive ductal carcinoma suggest that the invasive potential of breast carcinoma is already achieved before morphologically overt invasive growth is observed. As MT1-MMP mRNA expression is significantly correlated with axillary nodal metastasis, it may be useful as a prognostic indicator of invasive ductal carcinoma. Considering the positive correlation of MT1-MMP mRNA and TIMP2mRNA expression, our finding supports a role for TIMP2 in tumor growth, as well as the utility of CD44std as a prognostic indicator of breast cancer.
Tissue Inhibitor of Metalloproteinase-2/genetics ; RNA, Messenger/metabolism ; Matrix Metalloproteinases, Membrane-Associated ; Matrix Metalloproteinases/*genetics ; Matrix Metalloproteinase 9/genetics ; Matrix Metalloproteinase 2/genetics ; Matrix Metalloproteinase 1/genetics ; Humans ; Gene Expression Regulation, Neoplastic ; Gene Expression Regulation, Enzymologic ; Female ; Carcinoma, Ductal, Breast/genetics/*physiopathology ; Carcinoma in Situ/genetics/*physiopathology ; Breast Neoplasms/genetics/*physiopathology