Objective To explore the effect of overwork (OW) on extracellular matrix of arterial vessel wall in rats. Methods Random number grouping method was employed to assign 18 Sprague-Dawley rats into three groups(n=6):the control group(no special treatment),group OW(forced swimming twice a day for 15 days),and sleep deficiency(SD)+OW group(in addition to forced swimming twice a day,the rats were put on the platforms in water to limit sleep for 15 days).On the 16th day,the abdominal aorta and common carotid artery were collected after blood sampling from heart under deep anesthesia.A part of the abdominal aorta sample was taken for Masson staining of collagen fiber,and Verhoeff-Van Gieson staining was carried out for the elastic fiber of common carotid artery.Image J was employed for the quantitative analysis of collagen fiber and elastic fiber content.The expression of collagen 1(Col-1) protein was quantified by immunohistochemistry and the ultrastructure of vascular matrix was examined by transmission electron microscopy.The other part of the abdominal aorta sample was used to determine the mRNA levels of matrix metalloproteinase(MMP)-1,MMP-2,MMP-9,tissue inhibitor of metalloproteinases-1(TIMP-1),and Col-1 by quantitative real-time polymerase chain reaction. Results Compared with that in control group,the content of collagen fiber in groups OW and SD+OW had no significant change(all P>0.05);the content of elastic fiber in groups OW and SD+OW decreased(all P<0.001) and had no significant difference between each other(P>0.05).The vascular vessel wall of group OW showed slight fiber breakage,while that of group SD+OW presented wormhole-like or spongy fiber fragmentation.The mRNA levels of MMP-1 and MMP-2 in groups OW and SD+OW had no significant difference between each other(P>0.05) but were higher than that in control group(all P<0.001).The mRNA levels of MMP-9 and TIMP-1 had no significant difference among the three groups(all P>0.05).Groups OW and SD+OW had lower mRNA level(all P<0.001) and protein level(all P<0.001) of Col-1 than control group,while the mRNA and protein levels of Col-1 had no significant difference between groups OW and SD+OW(P>0.05). Conclusion OW can reduce the content of Col-1 and elastic fibers in the extracellular matrix of arterial vessels,destroy the elastic lamina of vascular wall,up-regulate the expression of MMP-1 and MMP-2,thereby injuring arterial vessels.
Animals ; Collagen Type I ; Extracellular Matrix/metabolism* ; Matrix Metalloproteinase 1/metabolism* ; Matrix Metalloproteinase 2/metabolism* ; Matrix Metalloproteinase 9/metabolism* ; RNA, Messenger/genetics* ; Rats ; Rats, Sprague-Dawley ; Tissue Inhibitor of Metalloproteinase-1/metabolism*

Salvianolic acid A (SAA) is a water-soluble component from the root of Salvia Miltiorrhiza Bge, a traditional Chinese medicine, which has been used for the treatment of cerebrovascular diseases for centuries. The present study aimed to determine the brain protective effects of SAA against cerebral ischemia reperfusion injury in rats, and to figure out whether SAA could protect the blood brain barrier (BBB) through matrix metallopeptidase 9 (MMP-9) inhibition. A focal cerebral ischemia reperfusion model was induced by middle cerebral artery occlusion (MCAO) for 1.5-h followed by 24-h reperfusion. SAA was administered intravenously at doses of 5, 10, and 20 mg·kg. SAA significantly reduced the infarct volumes and neurological deficit scores. Immunohistochemical analyses showed that SAA treatments could also improve the morphology of neurons in hippocampus CA1 and CA3 regions and increase the number of neurons. Western blotting analyses showed that SAA downregulated the levels of MMP-9 and upregulated the levels of tissue inhibitor of metalloproteinase 1 (TIMP-1) to attenuate BBB injury. SAA treatment significantly prevented MMP-9-induced degradation of ZO-1, claudin-5 and occludin proteins. SAA also prevented cerebral NF-κB p65 activation and reduced inflammation response. Our results suggested that SAA could be a promising agent to attenuate cerebral ischemia reperfusion injury through MMP-9 inhibition and anti-inflammation activities.
Animals ; Anti-Inflammatory Agents ; administration & dosage ; Blood-Brain Barrier ; drug effects ; enzymology ; immunology ; Brain ; Brain Ischemia ; drug therapy ; enzymology ; genetics ; Caffeic Acids ; administration & dosage ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Lactates ; administration & dosage ; Male ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; enzymology ; genetics ; immunology ; prevention & control ; Salvia miltiorrhiza ; chemistry ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Transcription Factor RelA ; genetics ; immunology

BACKGROUND: Macrophage colony-stimulating factor (M-CSF), matrix metalloproteinase-9 (MMP-9), and its specific tissue inhibitor - tissue inhibitor of metalloproteinases-1 (TIMP-1) may play an important role in the pathogenesis and spread of cancer. We investigated the plasma levels of M-CSF, MMP-9, and TIMP-1 in comparison with a commonly accepted tumor marker CA 15-3 in breast cancer patients and in control groups. METHODS: The cohort included 110 breast cancer patients in groups at stages I-IV. The control group consisted of 50 healthy volunteers and 50 benign tumor patients. Plasma levels of M-CSF, MMP-9, and TIMP-1 were determined by using ELISA, while CA 15-3 concentrations were determined by using chemiluminescent microparticle immunoassay (CMIA). RESULTS: The results showed significant differences in concentrations of the analyzed parameters and in levels of CA 15-3 between the groups of breast cancer patients and the two control groups. Diagnosis using these markers was equal to that using CA 15-3 in terms of sensitivity, predictive values of positive and negativetest results (PPV, NPV) and area under the ROC curve (AUC) in the studied groups. The diagnostic specificities of MMP-9, TIMP-1, M-CSF, and CA 15-3 showed equally high values (95%). The combined use of all tested parameters with CA 15-3 resulted in increased sensitivity, NPV, and AUC, especially in the combination of M-CSF with tumor markers (76%, 64%, and 0.8653). CONCLUSIONS: These findings suggest the tested parameters are useful in the diagnosis of breast cancer patients (except stage I), when combined with CA 15-3.
Adult ; Aged ; Area Under Curve ; Biomarkers, Tumor/*blood ; Breast Neoplasms/*diagnosis/genetics/pathology ; Case-Control Studies ; Female ; Humans ; Macrophage Colony-Stimulating Factor/*blood ; Matrix Metalloproteinase 9/*blood ; Middle Aged ; Mucin-1/blood ; Neoplasm Staging ; Poland ; ROC Curve ; Sensitivity and Specificity ; Tissue Inhibitor of Metalloproteinase-1/*blood

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 investigate the effect of RbAp48 knockdown on the migration and invasion of human cervical cancer cells and explore the mechanism.

METHODSA small interference RNA (siRNA) was used to knock down the expression of RbAp48 in MS751 cells. The changes in cell migration and invasion were evaluated using wound healing assay and Transwell assay, respectively, and the expressions of RbAp48, vimentin, N-cadherin, E-cadherin, Snail, Twist, MMP-2 and TIMP-2 were determined with Western blotting.

RESULTSAfter siRNA-mediated RbAp48 knockdown, MS751 cells showed a significantly reduced expression of RbAp48 with significantly suppressed cell migration and invasion (P<0.01). RbAp48 knockdown induced obvious down-regulation of the expressions of interstitial cell phenotype proteins vimentin, N-cadherin, and MMP-2 and up-regulation of epithelial cell phenotype proteins E-cadherin and TIMP-2, suggesting the inhibition of epithelial- mesenchymal transition of the cells. The expressions of Snail and Twist were significantly down-regulated in the cells following RbAp48 knockdown.

CONCLUSIONKnockdown of RbAp48 can significantly inhibit epithelial-mesenchymal transition and suppress the migration and invasion of cervical cancer cell line MS751, the mechanism of which may involve the down-regulation of Snail and Twist expressions.

Antigens, CD ; metabolism ; Cadherins ; metabolism ; Cell Line, Tumor ; Cell Movement ; Down-Regulation ; Epithelial-Mesenchymal Transition ; Female ; Gene Knockdown Techniques ; Humans ; Matrix Metalloproteinase 2 ; metabolism ; Neoplasm Invasiveness ; Nuclear Proteins ; metabolism ; RNA, Small Interfering ; Retinoblastoma-Binding Protein 4 ; genetics ; Snail Family Transcription Factors ; Tissue Inhibitor of Metalloproteinase-2 ; metabolism ; Transcription Factors ; metabolism ; Twist-Related Protein 1 ; metabolism ; Up-Regulation ; Uterine Cervical Neoplasms ; pathology ; Vimentin ; metabolism

OBJECTIVETo investigate the effects of Biejia Ruangan Tablet ([symbol in text], BRT)-containing serum on the expression of matrix metalloproteinase (MMP-9) and tissue inhibitor of metalloproteinase (TIMP-1) in cultured renal interstitial fibroblasts.

METHODSDifferent BRT-containing sera were prepared by gastric gavages to rats with the high-dose (7 g/kg), mid-dose (3.5 g/kg), and low-dose (1.75 g/kg) BRT respectively. The expression of extracellular matrix in NRK-49F cells was induced by treatment with human transforming growth factor-β1 (recombined human TGF-β1), and BRT-containing serum. Western blotting and Northern blotting were used to measure type I and III procollagen, MMP-9, and TIMP-1.

RESULTSThe high dose BRT-containing serum could decrease the type I and III procollagen gene expression which boosted by TGF-β1, at the same time cut down TIMP-1 protein and gene expression which increased by TGF-β1 (P <0.05). Treatment of cells with recombined human TGF-β1 had no significant effect on MMP-9 expression and BRT-containing serum also had no effect on MMP-9 expression.

CONCLUSIONSHigh dose BRT has anti-fibrosis effects in NRK-49F cells, as indicated by its inhibition of type I and III procollagen and TIMP-1 expression.

Animals ; Cells, Cultured ; Collagen Type I ; genetics ; metabolism ; Collagen Type III ; genetics ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Fibroblasts ; drug effects ; metabolism ; Gene Expression Regulation ; drug effects ; Humans ; Kidney ; cytology ; Male ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats, Sprague-Dawley ; Serum ; metabolism ; Tablets ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Transforming Growth Factor beta1 ; pharmacology

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 effects of Fengbaisan (, FBS) on the expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in lung tissue of rats with chronic obstructive pulmonary disease (COPD) and to investigate the preventive and therapeutic mechanisms of FBS.

METHODSThe COPD rat model was established by cigarette smoke exposure and lipopolysaccharide (LPS) intra-tracheal dripping. The histopathological changes of lung tissue was observed via hematoxylin/eosin staining. The expression of MMP-9 and TIMP-1 in lung tissue was measured by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry.

RESULTSThe typical histopathological changes of COPD were displayed in the model group, Ambroxol Hydrochloride group and FBS group, and the pathological lesions in the FBS group were less than those in the model group. The expression of MMP-9 and TIMP-1 in the model group increased significantly compared with those in the normal group (P<0.05). After treatment for successive 28 days, the expression of MMP-9 and TIMP-1 in the FBS group decreased remarkably as compared with the model group (P<0.05).

CONCLUSIONSFBS can regulate MMP-9/TIMP-1 imbalance to prevent airway and lung parenchyma remodeling process via reducing the expression of MMP-9 and TIMP-1 in the lung tissue of COPD rats, and this may be a possible therapeutic mechanism of FBS on COPD.

Animals ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Gene Expression Regulation, Enzymologic ; drug effects ; Immunohistochemistry ; Lung ; drug effects ; enzymology ; pathology ; Male ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Pulmonary Disease, Chronic Obstructive ; drug therapy ; enzymology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism

BACKGROUNDExposure of adult mice to more than 95% O2 produces a lethal injury by 72 hours. Nuclear factor kappa B (NF-κB) is a transcriptional factor that plays a key role in the modulation of cytokine networks during hyperoxia-induced acute lung injury (ALI). Osteopontin (OPN) is a phosphorylated glycoprotein produced principally by macrophages. Studies have reported that exogenous OPN can maintain the integrity of the cerebral microvascular basement membrane and reduce brain damage through inhibiting NF-κB activities in the brain after subarachnoid hemorrhage. However, it is not clear whether OPN can reduce lung injury during ALI by inhibiting transcriptional signal pathways of NF-κB and consequent inhibition of inflammatory cytokines. Thus we examined the effects and mechanisms of recombinant OPN (r-OPN) on ALI.

METHODSNinety-six mice were randomly divided into phosphate buffered saline (PBS) and r-OPN groups. Mice were put in an oxygen chamber (>95% O2) and assessed for lung injury at 24, 48, and 72 hours. Expressions of NF-κB, matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9), and tissue inhibitors of MMP-2 and MMP-9 (TIMP-1, TIMP-2) mRNA in lungs were examined with RT-PCR. Expression and distribution of NF-κB protein in lungs were measured with immunohistochemistry.

RESULTSExposure to hyperoxia for 72 hours induced more severe lung injury in the PBS group compared with the r-OPN group. Expression of NF-κB mRNA in the PBS group exposed to hyperoxia for 48 and 72 hours was significantly higher than the r-OPN group (P < 0.05). With 72-hour exposure, expression of TIMP-1 mRNA in the r-OPN group was significantly higher than that of the PBS group (P < 0.05). Expression of TIMP-2 mRNA in the r-OPN group at 48 and 72 hours was significantly higher than those in the PBS group (P < 0.05). After 72-hour exposure, expression of NF-κB protein in airway epithelium in the PBS group was significantly higher than that in the r-OPN group (P < 0.05).

CONCLUSIONr-OPN can inhibit the release and activation of MMPs through inhibition of the expression of NF-κB and promotion of the expression of TIMPs, and alleviate hyperoxia-induced ALI.

Acute Lung Injury ; genetics ; metabolism ; Animals ; Hyperoxia ; metabolism ; physiopathology ; Matrix Metalloproteinase 2 ; genetics ; metabolism ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Mice ; NF-kappa B ; genetics ; metabolism ; Osteopontin ; genetics ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Tissue Inhibitor of Metalloproteinase-2 ; genetics ; metabolism

OBJECTIVETo investigate the effects of Herbal Compound 861 (Cpd 861) on collagen synthesis and degradation in rat mesangial cells exposed to high glucose.

METHODSThe third to fifth passage of rat mesangial cells were exposed to high glucose and Cpd 861 at a concentration of 0.25-4.00 g/L for 24, 48 and 72 h, respectively. Benazepril (10(-7)-10(-3) mmol/L) was selected as positive control. The methyl thiazolyl tetrazolium colorimetric assay was used to evaluate the effect of Cpd 861 on cell proliferation. After incubation with Cpd 861 at a concentration of 2.00 g/L for 48 h, the protein secretions of collagen type IV, matrix metallopeptidase 9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), transforming growth factor beta 1 (TGF-β1), and hepatocyte growth factor (HGF) were detected by enzyme-linked immunosorbent assay method. And rat mesangial cells were harvested to determine MMP-9, TIMP-1, TGF-β1 and HGF mRNA expression by reverse transcription polymerase chain reaction.

RESULTSCpd 861 inhibited cell proliferation induced by high glucose in a dose- and time-dependent manner. Compared with high glucose, collagen type IV production was decreased significantly by Cpd 861 (P<0.01). Cpd 861 increased the protein secretions and mRNA expressions of MMP-9 and HGF, whereas the protein secretions and mRNA expressions of TIMP-1 and TGF-β1 were reduced markedly (P<0.05). The ratio of MMP-9 to TIMP-1 was enhanced by Cpd 861 significantly. There was no significant difference in all above-mentioned effects between Cpd 861 (2.00 g/L) and benazepril (10(-5) mmol/L).

CONCLUSIONThe anti-glomerulosclerosis mechanisms of Cpd 861 were partly attributed to its effects of inhibiting mesangial cell proliferation, decreasing collagen synthesis and enhancing collagen degradation.

Animals ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen Type IV ; biosynthesis ; secretion ; Drugs, Chinese Herbal ; pharmacology ; Fibrosis ; Glucose ; toxicity ; Hepatocyte Growth Factor ; secretion ; Matrix Metalloproteinase 9 ; metabolism ; Mesangial Cells ; cytology ; drug effects ; enzymology ; metabolism ; Polymerase Chain Reaction ; Proteolysis ; drug effects ; RNA, Messenger ; genetics ; metabolism ; Rats ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism ; Transforming Growth Factor beta1 ; secretion