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Objective Analysis of the effect and the mechanism of adenovirus with down regulation of matrix metalloproteinase inhibitor 1 (TIMP-1) achieved targeting by ultrasound microbubbles combined with ultrasound irradiation for liver fibrosis in rats. Methods Recombinant adenovirus-mediated with down regulation of TIMP-1 gene was constructed and a mixture of recombinant adenovirus and ultrasound contrast agent was prepared.The rat liver fibrosis model was established and divided into 5 groups : model group; adenovirus group (recombinant adenovirus); adenovirus microbubble group (mixture of recombinant adenovirus and ultrasound contrast agent); experimental group (ultrasound irradiation + mixture of recombinant adenovirus and ultrasound contrast agent); ultrasound adenovirus group (ultrasound irradiation + recombinant adenovirus). The rats were sacrificed after 24 hours and liver sections were prepared. The expression of EGFP in each group was observed and the transfection rate was analyzed. The liver slices were stained by Masson to judge the stage of liver fibrosis. ANOVA analysis was used to compare the levels of alanine aminotransferase (ALT) , aspartate aminotransferase (AST) , bydroxyproline (Hyp) , hyaluronic acid (HA) , type IV collagen (CIV) and laminin (LN) in each group. The relative expression levels of TIMP-1 and matrix metalloproteinase 13 (MMP-13) in each group were detected by Western blot. Results The transfection rate of the experimental group was higher than that of the adenovirus group (q = 3.418) , the adenovirus microbubble group (q = 3.756) and the ultrasonic adenovirus group (q = 5.502) , and the difference was statistically significant (P < 0.05); Pathological examination showed that the degree of fibrosis in the experimental group and the grade of liver fibrosis were lower than the other groups (P < 0.01). The activities of ALT, AST, HA, LN, CIV and Hyp in the experimental group were lower than those in the other 4 groups.Western blot showed that the level of TIMP-1 protein expression was highest while the level of MMP-13 protein expression was lowest in the experimental group than those in the other groups. Conclusion Adenovirus with down regulation of TIMP-1 achieved targeting by ultrasound microbubbles combined with ultrasound irradiation can inhibit the activity of TIMP-1 and improve the degree of liver fibrosis. Gene therapy is an potential therapeutic method in the application of treating liver fibrosis.
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Objective: To investigate the effects of artesunate (ART) on diabetic retinopathy of rats by detecting the expression variation of matrixmetalloproteinase-9 (MMP-9) and inhibitory factor matrix metalloproteinase inhibitor-1 (TIMP-1) in retina. Methods: A total of 24 healthy male SD rats were randomly divided into negative control, model control and ART treated groups. The model control group and ART treated group were given injection of streptozotocin (STZ) to establish the type 1 diabetic rat model. After being raised for three months, the ART treated rats' abdomina were given ART injection for 10 d. All retinas of three groups were isolated. The expression of MMP-9 and TIMP-1 mRNA was detected by Real-time PCR (RT-qPCR) and the expression of MMP-9 protein was detected by immunohistochemical staining. Results: The expression of MMP-9 in ART treated group was significantly decreased compared with model control group (P < 0.05), whereas TIMP-1 in ART treated group was significantly increased (P < 0.05). The immunohistochemical staining showed that the positive expression of MMP-9 protein in ART treated group was significantly less than that in model control group. Conclusion: ART has inhibitory effects on the expression of MMP-9 in the diabetic retinopathy of rats.
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Objective; To investigate the expression levels of matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9) in dentin of healthy adults under different adhesion conditions, and to provide the basis for improving the binding effect of matrix metalloproteinases (MMPs) inhibitors in clinic. Methods; A total of 20 adult freshlyextracted molars were collected and ground into dentin powder under liquid nitrogen cooling. The oral condition was simulated. The dentin was treated with self-etching bonding (Single Bond Universal) and totaletching bonding (35% phosphate gel +Adpter Single Bond 2). The dentin without any treatment was regarded as negative control group, the dentin treated with 10% phosphoric acid (self-etching bonding) or 10% phosphoric acid+ 35% phosphate gel (total-etching bonding) was regarded as positive control group, the dentin treated with Single Bond Universal (self-etching bonding) or 35% phosphate gel+Adper Single Bond 2 (total-etching bonding) was regarded as blank control group; the dentin pretreated with the MMPs inhibitors chlorhexidine (CHX) and minocycline (MI) during the bonding process respectively was regarded as CHX group and MI group, and the dentin treated with 10 % sodium hypochlorite was regarded as aging group; on the basis of aging group, the dentin treated with CHX and MI was regarded as CHX aging group and MI aging group. Gelatin zymography was used to perform the polyacrylamide gel electrophoresis; after incubating, staining and decoloring, the bands were analyzed by gel image analysis system, and the expression levels of MMP-2 and MMP-9 in dentin were calculated. Results: Under the condition of self-etching bonding, compared with blank control group, the expression levels of MMP-2 and MMP-9 in dentin in CHX group were decreased (P<0. 05), and the expression levels of MMP-2 and MMP-9 in the dentin powder in MI group were decreased (P<0. 05); compared with aging blank control group, the expression levels of MMP-2 and MMP-9 in dentin in CHX aging group were decreased (P<0. 05), and the expression levels of MMP-2 and MMP-9 in dentin in MI aging group were decreased (P<0. 05). Under the condition of total-etching bonding, compared with blank control group, the expression levels of MMP-2 in dentin in CHX group and MI group were decreased (P<0. 05); compared with aging blank control group, the expression levels of MMP-2 in dentin in CHX aging group and MI aging group were decreased (P<0. 05). Conclusion; In the process of dentin bonding, CHX and MI could slow down the enzymatic reaction and improve the bonding strength by decreasing the expression levels of MMP-2 and MMP-9.
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Objective:To investigate the curative efficacy of combined use of dexamethasone palmitate,vitamin B12 and lidocaine in the treatment of knee osteoarthritis and its effects on the seum levels of interleukin-1 (IL-1),matrix metalloproteinase-3 (MMP-3) and matrix metalloproteinase-1 (TIMP-1).Methods:84 patients with osteoarthritis of knee were selected from August 2014 to July 2015 in Sixth Affiliated Hospital of Xinjiang Medical University and divided into the observation group and the control group according to the order of admission,42 cases in each group.The observation group was given intra-articular injection of dexamethasone palmitate,vitamin B12 and lidocaine,and the control group was treated with traditional Chinese medicine.The clinical efficacy was evaluated after treatment.The changes of serum IL-1,MMP-3,TIMP-1 levels and visual analogue (VAS) score were compared before and at 1 months after treatment.Results:After treatment,the total effective rate of observation group was significantly higher than that of the control group (P＜0.05),the serum levels ofIL-1,MMP-3 levels and VAS score were significantly lower than those before treatment,serum TIMP-1 level was obviously higher than that before treatment(P ＜0.05),the serum levels of IL-1,MMP-3 levels were significantly lower in the observation group than those of the control group,while the serum TIMP-1 level was obviously higher than(P＜0.05) Conclusion:Intra-articular injection with dexamethasone palmitate,vitamin B12 and lidocaine could effectively decrease the levels of serum IL-1 and MMP-3,increase the level ofTIMP-1,and relieve the pain in the patients with knee osteoarthritis.
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Objective:To investigate the effect of the extract of Schisandra chinensis on the matrix metalloproteinases(MMPs) system in kidney tissue of the diabetic rats,and to explore its protective effect on the kidney tissue from the matrix degradation perspective.Methods:STZ was used to establish rat models of diabetes mellitus.A total of 45 diabetic rats were randomly divided into model group,extract of Schisandra chinensis group and Benazepril group,and there were 15 rats in each group.Another 15 rats were selected and used as normal control group.12 weeks after administration,the routine blood and urine biochemical indexes,the histological changes,blood glucose (BG),blood urea nitrogen(BUN),serum creatinine(Scr),high density lipoprotein cholesterol(HDL-C),low density lipoprotein cholesterin(LDL-C),total cholesterol(T-CHO),and triglyceride(TG) levels,excretion rates of albuminuria and proteinuria of the rats in various groups were detected;the expression amounts of fibronectin (FN),type Ⅳ collagen (Col Ⅳ),and matrix metalloproteinase inhibitor metalloproteinase-2 (TIMP-2) in kidney tissue of the rats were detected by immunohistochemical method;the activity of matrix metalloproteinase-2 (MMP-2) was detected by zymography.Results:Compared with model group,the glomeruli matrix accumulation of the rats in extract of Schisandra chinensis group was significantly improved,the excretion rate of albuminuria,LDL-C level and serum MDA level were decreased(P<0.05),the activities of CAT(P<0.01)and SOD(P<0.05)in kidney tissue were increased,and the level of MDA in kidney tissue was decreased(P<0.05).The immunohistochemistry results showed that compared with model group,the expression amounts of FN,Col Ⅳ,and TIMP-2 in kidney tissue of the rats in extract of Schisandra chinensis group were significantly decreased.The zymography results showed that compared with model group,the activity of MMP-2 in kidney tissue of the rats in extract of Schisandra chinensis group was significantly increased(P<0.05).Conclusion:Extract of Schisandra chinensis has protective effect on the kidney tissue of the diabetic rats induced by STZ,and the mechanism may be related to the inhibition of oxidative stress and the improvement of MMP-2 activity as well as the inhibition of TIMP-2 expression which could improve the matrix degradation.
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We evaluated the safety of matrix metalloproteinase (MMP) inhibitor in experimental glaucoma filtration surgery in an animal model. Fifteen New Zealand white rabbits underwent an experimental trabeculectomy and were randomly allocated into 3 groups according to the adjuvant agent: no treatment group (n = 5), 0.02% mitomycin C (MMC) soaking group (n = 5), and MMP inhibitor (ilomastat) subconjunctival injection group (n = 5). Slit lamp examination with Seidel testing, pachymetry, and specular microscopy was performed preoperatively and postoperatively. The conjunctiva and ciliary body toxicity were evaluated with scores according to the pathologic grading systems. Electron microscopy was used to examine the structural changes in cornea, conjunctiva, and ciliary body. In the ilomastat-treated group, there was no statistically significant change in central corneal thickness preoperatively and at 28 days postoperatively (P = 0.655). There were also no significant changes in specular microscopy findings over the duration of the study in the ilomastat-treated group. The conjunctival toxicity score was 1 in the control group, 1.5 in the ilomastat-treated group, and 2 in the MMC-treated group. When assessing ciliary body toxicity scores, the ilomastat-treated group score was 0.5 and the MMC-treated group score was 1.5. Transmission electron microscopy did not show structural changes in the cornea and ciliary body whereas the structural changes were noticed in MMC group. A single subconjunctival injection of MMP inhibitor during the experimental trabeculectomy showed a less toxic affect in the rabbit cornea, conjunctiva, and ciliary body compared to MMC.
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Background The primary pathologic mechanism of posterior capsular opacification (PCO) is proliferation,migration and epithelial-mesenchymal transformation of residuary lens epithelial cells (LECs) following cataract surgery.Matrix metalloproteinases (MMPs) play a role during the migration of LECs.Researches showed that GM6001,a broad inhibitor of MMPs,can arrest the migration of LECs,but as specific inhibitors of MMPs,the efficacy and safety of MMP-2/9 inhibitor Ⅰ and Ⅱ on LECs migration remain unclear.Objective This study was to determine and compare the inhibitory efficacy among GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ on human LECs and search the clinical medication to prevent PCO.Methods Human LECs were cultured and passaged in vitro,and the cells of 3-4 generation were incubated in 6-well plates.Then the cells of 70％ confluent monolayer were cultured in DMEM without fetal bovine serum for 12 hours.GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ at different concentrations (0.25,0.50,1.00,2.00,4.00,8.00,16.00,32.00,64.00,128.00 μmol/L) were added into the culture medium for 24 hours separately,and regularly cultured cells served as the control group.A bare area was made by a 200 μl sterile spear on the cell layer,and the migrated distance and inhibitory rate were calculated.The second or third generation of cells were incubated in 96-well plates at a density of 5×105/ml (200 μl/well).GM6001 (128.00 μmol/L),MMP-2/9 inhibitor Ⅰ (64.00 μmol/L) and Ⅱ (32.00 μmol/L) were added into the culture medium for 24 hours,and the cell viability was assayed by using MTT assay.Results Cultured cells grew well with irregular arrangement and presented the polygon in shape.The migrated distance was gradually reduced as the increase of concentrations of GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ,showing significant differences among the various concentration groups (GM6001:F=248.647,P＜0.05;MMP-2/9 inhibitor Ⅰ:F=357.125,P＜0.05;MP2/9 inhibitor Ⅱ:F=396.374,P＜ 0.05).The cell migrated distance in the control group was set to 1,the relative migrated distances were 0.478 ± 0.091,0.294±0.088 and 0.191 ±0.081 in the GM6001 group,MMP-2/9 inhibitor Ⅰ group and MMP-2/9 inhibitor Ⅱ group at the concentrations of 32.00 μmol/L,respectively,showing a significant difference among the groups (F =116.031,P＜0.01),and cell migrated distance was obviously shorter in the MMP-2/9 inhibitor Ⅱ group than that in the GM6001 group or MMP-2/9 inhibitor Ⅰ group (all at P＜0.01).The A values were 0.607±0.016,0.567±0.015,0.583±0.010 and 0.595 ±0.0138 in the control group,GM6001 group (128.00 μmol/L),MMP-2/9 inhibitor Ⅰ group (64.00 μmol/L) and MMP-2/9 inhibitor Ⅱ group (32.00 μmol/L),respectively,without significant difference among the groups (F=1.403,P＞0.05).Conclusions GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ reduce the mobility of human LECs effectively but do not affect the viability of the cells in vitro.MMP-2/9 inhibitor Ⅱ appears to be most dominant in inhibiting migration of human LECs.
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The aim of this study was to investigate whether matrix metalloproteinase (MMP) inhibitors attenuate neuroinflammation in an ischemic brain following photothrombotic cortical ischemia in mice. Male C57BL/6 mice were anesthetized, and Rose Bengal was systemically administered. Permanent focal ischemia was induced in the medial frontal and somatosensory cortices by irradiating the skull with cold white light. MMP inhibitors, such as doxycycline, minocycline, and batimastat, significantly reduced the cerebral infarct size, and the expressions of monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor-alpha (TNF-alpha), and indoleamine 2,3-dioxygenase (IDO). However, they had no effect on the expressions of heme oxygenase-1 and neuroglobin in the ischemic cortex. These results suggest that MMP inhibitors attenuate ischemic brain injury by decreasing the expression levels of MCP-1, TNF-alpha, and IDO, thereby providing a therapeutic benefit against cerebral ischemia.
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Animals , Humans , Male , Mice , Brain , Brain Injuries , Brain Ischemia , Chemokine CCL2 , Cold Temperature , Doxycycline , Globins , Heme Oxygenase-1 , Indoleamine-Pyrrole 2,3,-Dioxygenase , Ischemia , Light , Matrix Metalloproteinase Inhibitors , Minocycline , Nerve Tissue Proteins , Phenylalanine , Rose Bengal , Skull , Thiophenes , Tumor Necrosis Factor-alphaABSTRACT
BACKGROUND: Many inflammatory mediators and collagenases are involved in the pathogenesis of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). The increase of matrix metalloproteinase-9 (MMP-9, gelatinase-B) produced mainly by inflammatory cells was reported in many ALI models and ARDS patients. Cyclic mechanical stress also can induce MMP-9 production from alveolar macrophages and connective tissue cells. In this study, the expression of MMP-9 in ventilator-induced lung injury (VILI) model and the effects of matrix metalloproteinase inhibitor (MMPI) on VILI were investigated. METHODS: Eighteen Sprague-Dawley rats were divided into three groups: low tidal volume (LVT, 7mL/Kg tidal volume, 3 cmH2O PEEP, 40/min.), high tidal volume (HVT, 30mL/Kg tidal volume, no PEEP, 40/min) and high tidal volume with MMPI (HVT+MMPI) groups. Mechanical ventilation was performed in room air for 2 hours. The 20 mg/Kg of CMT-3 (chemically modified tetracycline-3, 6-demethyl 6-deoxy 4-dedimethylamino tetracycline) was gavaged as MMPI from three days before mechanical ventilation. The degree of lung injury was measured with wet-to-dry weight ratio and acute lung injury score. Expression of MMP-9 was studied by immunohistochemical stain with a mouse monoclonal anti-rat MMP-9 IgG1. RESULTS: In the LVT, HVT and HVT + MMPI groups, the wet-to-dry weight ratio was 4.70+/-0.14, 6.82+/-1.28 and 4.92+/-0.98, respectively. In the HVT group, the ratio was significantly higher than other groups (p<0.05). Acute lung injury score measured by five-point scale was 3.25+/-1.17, 12.83+/-1.17 and 4.67+/-0.52, respectively. The HVT group was significantly damaged by VILI and MMPI protects injuries by mechanical ventilation (p<0.05). Expression of MMP-9 measured by four-point scale was 3.33+/-2.07, 12.17+/-2.79 and 3.60+/-1.95, respectively, which were significantly higher in the HVT group (p<0.05). CONCLUSION: VILI increases significantly the expression of MMP-9 and MMPI prevents lung injury induced by mechanical ventilation through the inhibition of MMP-9.
Subject(s)
Animals , Humans , Mice , Rats , Acute Lung Injury , Collagenases , Connective Tissue Cells , Immunoglobulin G , Lung Injury , Macrophages, Alveolar , Matrix Metalloproteinase 9 , MMPI , Rats, Sprague-Dawley , Respiration, Artificial , Respiratory Distress Syndrome , Stress, Mechanical , Tidal Volume , Ventilator-Induced Lung InjuryABSTRACT
Matrix metalloproteinase (MMP) are a family of zinc-dependent endoproteinases whose enzymatic activity is directed against components of the extracelluar matrix(ECM). Their activities are inhibited by the tissue inhibitor of metalloproteinase (TIMP). MMP exerted an important role in invasion and metastasis of gastrointestinal cancer through degrading the ECM. With further research on inhibition of MMP, synthetic MMP inhibitor will have good prospects in the treatment of tumor invasion and metastasis.
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Objective By using estradiol(E2) as positive control to observe the effects of genistein(GST) on the matrix metalloproteinase inhibitor 1 mRNA levels of HSC-T6 cell in vitro.Methods HSC-T6 cells were exposed to different concentrations of E2 0.1?mol/L and GST 0.5-50?mol/L for 36 hours.The TIMP-1 mRNA levels were measured by the quantitative reverse-transcription polymerase chain reaction(RT-PCR).Results The TIMP-1 mRNA levels of HSC-T6 cells at different concentrations of E2 and GST were lower than those of the normal control(P