OBJECTIVE: To review the role of matrix metalloproteinase-1 (MMP-1) in the course of healing in wounded skin. METHODS: The recent literatures on MMP-1 in skin wound repair were reviewed, which gave the insight into the local effect of MMP-1 during re-epithelialization. RESULTS: Following injury, basal keratinocytes, moving from the wound edge and interact with dermal matrix proteins in the wound bed, were induced to express MMP-1 in a specific space-time pattern. MMP-1 cleaved the collagen, thereby altering its structure and affinity by which the keratinocytes binded it. MMP-1 served a beneficial role in wound healing by facilitating the proliferation and movement of keratinocytes over the collagen-rich wound bed during re-epithelialization. CONCLUSION: MMP-1 expression of migrating keratinocytes directly influences the re-epithelialization during the course of healing of the wounded skin.
Objective To investigate the effect of glucocorticoid on the expression levels of osteoprotegerin (OPG)/receptor activator of nuclear factor kappa B ligand (RANKL)-matrix metalloproteinases (MMP)/tissue inhibitor of matrix metalloproteinase (TIMP) system in bone tissues of femoral head of rats, and to discuss its interrelated action mechanism in glucocorticoid-induced avascular necrosis of femoral head (ANFH). Methods Forty adult Sprague Dawley rats, weighing 250-300 g, half males and half females, were randomly divided into 4 groups: high dose glucocorticoid group (HD, n=10), medium dose glucocorticoid group (MD, n=10), low dose glucocorticoid group (LD, n=10), and control group (n=10). The rats in HD group, MD group, and LD group were intramuscularly injected with 25.0, 12.5, and 7.0 mg/kg of prednisolone respectively, and the rats in the control group were injected with physiological saline. After 4 weeks intervention, the osteonecrosis of left femoral heads was observed by HE staining, total RNA was extracted from the right femoral head bone tissue and the mRNA expression levels of OPG, RANKL, MMP-2, MMP-9, TIMP-1, and TIMP-2 were detected by RT-PCR. Results After injection of prednisolone, 4 rats of HD group and 1 rat of MD group died of systemic failure caused by the decreased food and weight culminating in cachexia. HE staining showed that the integrity of bone trabecula and osteon was destroyed at different levels, discontinuous bone chips formed, and osteocytes were replaced by granulation tissue in some lacunae in HD, MD, and LD groups; the integrated osteon was observed, the lamellar structure formed concentric circles around the blood vessel and osteocytes were seen in the lacunae in the control group. The necrosis rates of femoral head were 83.3% (5/6), 66.7% (6/9), 30.0% (3/10), and 0 (0/10) in HD, MD, LD, and control groups. The results of RT-PCR showed: the mRNA expression levels of the OPG, TIMP-1, TIMP-2 in HD, MD, and LD groups were lower than those in the control group, showing significant differences (P lt; 0.05) and there was negative correlation with the hormone dosage. The difference in OPG expression was significant between the hormone groups (P lt; 0.05); the differences in the TIMP-1 and TIMP-2 expressions were not significant between the LD group and MD group (P gt; 0.05), but there were significant differences when compared with HD group (P lt; 0.05). The RANKL, MMP-2, and MMP-9 mRNA expression levels in HD, MD, and LD groups were higher than those in the control group and there was a positive correlation with the hormone dosage, showing significant differences when compared MD and HD groups with control group (P lt; 0.05); there was no significant difference in RANKL expression between HD group and MD group (P gt; 0.05), but there was significant difference when compared HD and MD groups with LD group (P gt; 0.05); no significant difference was observed in the MMP-2 and MMP-9 expression between MD group and LD group (P gt; 0.05), but the differences were significant when compared with HD group (P lt; 0.05). Conclusion Glucocorticoid-induced ANFH may be related to the expression levels of OPG/RANKL-MMP/TIMP mRNA regulated by glucocorticoid.
Objective To examine the effects of newly designed LY52 on the expression of matrix metalloproteinases and invasive ability of hepatocellular carcinoma HepG2 cells. Methods The effects of LY52 on the proliferations of HepG2 cells were detected by MTT assay. Gelatin zymography and Western blot were used to detect the effects of LY52 on matrix metalloproteinase-2 expression in the cell line. Transwell chamber assay was used to detect the effects of LY52 on the invasion of the cells. Results No obvious inhibitory or cytotoxicity effects of LY52 was found in lower concentrations (lt;200 μg/ml) of LY52. Gelatin zymography and Western blot showed that matrix metalloproteinase-2 expression were inhibited by LY52 in a dose-dependent manner in HepG2 cells. Furthermore, transwell chamber assay showed that LY52 could significantly inhibit the invasion of the cell line in a dose-dependent manner.Conclusion The results suggest that LY52 may inhibit the invasion of hepatocellular carcinoma cells by suppressing the matrix metalloproteinase-2 activity.
Objective To explore the expressions of E-cadherin， matrix metalloproteinases-2 （MMP-2） protein，and matrix metalloproteinases-9 （MMP-9） protein in gastric cancer tissues， and to analyze the possible statistical relati-onship between the expressions of E-cadherin， MMP-2 protein， and MMP-9 protein， and clinicopathological features ofgastric cancer. Methods The ABC immunohistochemical staining was adopted to examine the expressions of E-cadherin，MMP-2 protein， and MMP-9 protein in 40 paraffin slices of gastric cancer （gastric cancer group）， with the adjacent tissue as the control group （adjacent tissue group）. The positive rates of 3 kinds of protein were compared between the2 groups， in addition， the statistical relationship between the expressions of the 3 kinds of protein and clinicopathological features of gastric cancer was examined respectively by SPSS 19.0 software. Results The expressions of E-cadherin， MMP-2 protein， and MMP-9 protein were all found in gastric cancer tissues and adjacent tissues. In gastric cancer tissue group， the expression of E-cadherin downregulated while the expressions of MMP-2 protein and MMP-9 protein upregulated in comparison to adjacent tissue group （P＜0.05）. The significant association was found between the expre-ssion of E-cadherin and the gastric cancer tissues of T3＋T4 stage， N1-N3 stage， and Ⅲ＋Ⅳ stage， which had lower positive expression rate （P＜0.05）. The expression of MMP-2 protein in gastric cancer tissues of M1 stage and Ⅲ＋Ⅳstage upregulated （P＜0.05）， and the expression of MMP-9 protein upregulated in gastric cancer tissues of T3＋T4 stage，Ⅲ＋Ⅳ stage， or lowlydifferentiated＋undifferentiated （P＜0.05）. No significant relationship was found in other clinical-pathological features and 3 kinds of protein except aforementioned significant relationship （P＞0.05）. Conclusions In the development progress of gastric cancer， the E-cadherin may get involved in the mechanism of tumor invasion and lymph node metastasis， MMP-2 protein may get involved in the mechanism of distant metastasis， and MMP-9 protein may get involved in the mechanism of differentiation and tumor invasion. The examination of those 3 kinds of markers may play an role in the judgment of tumor stage and estimation of prognosis in gastric cancer clinically.
Objective To investigate the role of transforming growth factor β(TGF-β)in the regulation of the gene expression of matrix metalloproteinase 13(MMP-13)in the human hyaline chondrocytes. Methods The human hyaline chondrocytes harvested enzymatically and cultured in DMEM supplemented with 20% fetus calf serum were divided into 7 groups. Group 1 was used as a contol, and 1 ng/ml TGF-β(group 2), 10 ng/ml TGF-β(group 3), 100 ng/ml TGF-β(group 4), 1 ng/ml TGF-β＋10 ng/ml IL-1β(group 5), 10 ng/ml TGF-β＋10 ng/ml IL-1β(group 6),and 100 ng/ml TGF-β＋10 ng/ml IL1β(group 7) were given for 12-hour coculture. The MMP-13 mRNA levels of passaged human hyaline chondrocytes were assessed by reverse transcriptionpolymerase chain reaction(RT-PCR) and real-time fluorescent quantitative PCR. Results TGF-β can increase the MMP-13 mRNA level respectively in the passagedhyaline chondrocytes. In the multifactor treated groups, TGF-β can decrease the MMP-13 mRNA level respectively and there was significant difference between groups (Plt;0.05).The level of MMP-13 mRNA expression had significant coherence withthe dosage of TGF-β. Conclusion The above results show that human chondrocytes express MMP-13 mRNA. TGF-β could cause a dosedependent stimulation on MMP-13 gene expression in human chondrocytes and have a potent effect of antagonizing IL-1β in osteoarthritis. TGF-β may play a crucial role in the occurrence anddevelopment of osteoarthritis through regulating MMP-13.
【Abstract】Objective To construct a recombinant adenoviral vector carrying antisense matrix metalloproteinase2 (MMP2) for use in the gene therapy to inhibit the invasiveness and migratory capacity of hepatocellular carcinoma (HCC) cell line HepG2 in vitro and in vivo models. Methods Total RNA was extracted from HCC, and then a 500 bp fragment at the 5′ end of human MMP2 cDNA was synthesized by polymerase chain reaction (PCR) and was reversely inserted into the multiclone site (MCS) of the shuttle plasmid pAdTrack-CMV,with the resultant plasmid and the backbone plasmid pAdEasy-1,the homologous recombination took place in the E.coli BJ5183 and the recombinant adenoviral plasmid carrying the antisense MMP2 gene was constructed and generated. The adenoviruses(Ad-MMP2AS) were packaged and amplified in the HEK 293 cells.Then the viral titer was checked by GFP. Results The recombinant adenovirus vector carrying antisense MMP2 was constructed successfully, the b green fluorescence was observed in HEK 293 cells under a fluorescence microscopy. The viral titer was 1×108/ml. Conclusion The recombinant adenovirus Ad-MMP2AS constructed by us could introduce the antisense MMP2 into HepG2 effectively,which would provide experimental basis for reversing the overexpression of MMP2 in HCC and for inhibiting the invasiveness and migratory capacity of HepG2 in vitro and in vivo models.
Objective To observe the proapoptotic effect ofthe homogenate of different parts of pig’s full thickness dermal wounds on cultured fibroblasts. Methods The tissues were dissected from the wound center and subneoepithelium separately 15 days after homogenization and sterilization, the specimens stored at -70℃. The forth passage of the fibroblasts were cultured for 16 hours in different culture solutions and were grouped into 7 groups: DMEM containing 5% fetal bovine serum as Group Ⅰ, DMEM containing 5% homogenate of tissue from wound center as GroupⅡ, DMEM containing 5% homogenate of tissue from subneoepithelium as Group Ⅲ, the culture solution of Group Ⅱmixed with 10 μg/ml GM6001 in Group Ⅳ, with the culturing medium of Group Ⅲplus 10 μg/ml GM6001 as Group Ⅴ, the culture solution of Group Ⅱ mixed with 10 ng/ml aFGF as Group Ⅵ, and the culture solution of Group Ⅲ mixed with 10 ng/ml aFGF as Group Ⅶ. In all groups except Group Ⅰ, the fibroblasts of the 6 pigs were treated with the homogenate derived from the same animal respectively. After being incubated in Annexin Ⅴ-FITC and PI, cells were analyzed by Flow Cytometry and the rate of apoptotic cells was acquired. The data were analyzed by SPSS 11.0 using Leastsignificant Difference test(LSD). Results The apoptotic rate of the 7 groups were as follows:4.39%±0.41% in Group Ⅰ，10.98%±1.42% in Group Ⅱ，13.47%±1.44% in Group Ⅲ，7.2%±0.46% in Group Ⅳ，12.1%±0.85% in Group Ⅴ，3.9%±0.63% in Group Ⅵ，9.8%±0.50% in Group Ⅶ； there were significant differences between every two groups except Group Ⅰand Group Ⅵ. Conclusion Homogenate of the tissue derived from the subneoepithelium has greater proapoptotic effect than that from the wound center; the proapoptotic effect of homogenate of the tissue both under neoepithelium and in wound center can be significantly alleviated by acid fibroblast growth factor, partly because of MMPs.
Abstract: Objective To study the pathophysiological mechanism of the morphological change of immature pulmonary vessels in the piglet model of congenital heart defect with decreased pulmonary blood flow established with balloon atrial septostomy and pulmonary artery banding. Methods Twenty piglets at an age of one to two months were divided into three groups with random number table. For the control group (group C,n=6), small incisions were carried out on the right chest to produce a transient reduction in the pulmonary blood; for the lowmedium pulmonary artery stenosis group (group T1, n=7), the balloon dilator was delivered through the surface of the right atrium and septostomy and pulmonary artery banding were performed, and the systolic transpulmonary artery banding pressure (Trans-PABP) was controlled to be 20.30 mm Hg; For the severe pulmonary artery stenosis group (group T2, n=7), the same surgical procedures with group T1 were performed while TransPABP was controlled to be more [CM(159mm]than 3050 mm Hg.At 2 months after surgery respectively,a lung tissue of 1.0 cm×0.8 cm×0.8 cm from the lateral segment of the right middle lobe was taken out to be observed under optic microscope. The morphological change of the distal arterioles was detected. Furthermore, the content of vascular endothelial growth factor (VEGF) and matrix metalloproteinase2( MMP2) were also examined by the method of enzymelinked immunosorbent assay (ELISA). Results The model was successfully established in all the survival piglets of the group T1 and group T2. Two months after operation, the inner diameter of the pulmonary arterioles in group T1 was significantly higher than that in group C (82.89±10.72 μm vs.74.12±9.28 μm;t=－5.892, Plt;0.05), so as group T2 (85.47±5.25 μm vs.74.12±9.28 μm;t=-6.325, Plt;0.05); the number of arterioles per square centimeter (NAPSC) of group T1 was significantly lower than that of the group C (229.70±88.00 entries/cm 2 vs. 431.50±40.60 entries/cm2; t=39.526, Plt;0.05), so as group T2 (210.00±40.30 entries/cm2 vs. 431.50±40.60 entries/cm2; t=67.858, Plt;0.05). Two months after operation, the lung expression of MMP -2 and VEGF in group T1 was significantly lower than that in group C (58.30±19.60 ng/ml vs. 81.20±16.70 ng/ml, t=14.261, Plt;0.05; 17.80±3.00 pg/ml vs. 21.40±3.80 pg/ml, t=8.482, P＜0.05), so does group T2 (42.10±15.20 ng/ml vs. 81.20±16.70 ng/ml, t=27.318, P＜0.05; 12.30±3.20 pg/ml vs. 21.40±3.80 pg/ml, t=15.139, P＜0.05). Conclusion Structural remodeling of pulmonary extracellular matrix is an important feature of the piglet model of congenital heart defect with decreased pulmonary blood flow. The arterioles show significant hypoplasia or degradation. Change in the structural proteins and cytokines during the reduction of blood in the lung is the key to structural remodeling.
Abstract: Objective To observe the expression of integrinlinked kinase (ILK) and matrix metalloproteinases9 (MMP9) in human nonsmall cell lung cancer (NSCLC) and investigate the correlation of ILK and MMP9 expression with the prognosis of NSCLC. Methods The expression of ILK and MMP9 in 75 specimens of NSCLC resected from January 2002 to January 2004 were detected by immunohistochemistry. According to the median of integral optical density (IOD), all patients were divided into the high or low ILK expression group and the high or low MMP-9 expression group. The relativity of ILK and MMP9 was determined, and the relationship of survival time with clinical features including expression of ILK and MMP-9 was compared by Logrank test. Results Both ILK and MMP-9 were expressed in NSCLC specimens. The expression between ILK and MMP-9 was positively correlated in 75 patients of our group (r=0.79, Plt;0.05). Patients with lower expression of ILK and MMP9 had a significantly longer survival time than those with higher expression of ILK and MMP-9 in the postoperative followup (χ2=15.067,14301,Plt;0.05). The survival time was not correlated with sex,age,smoking history or pathological type(χ2=0450,0078, 1.460, 1.623，Pgt;0.05), while tumor diameter, lymph node metastasis, TNM stage, the expression of ILK and MMP-9 significantly influenced the survival time (χ2=3.963, 15.169,20.529, 15.067，14.301，Plt;0.05). Conclusion The expression of ILK and MMP9 affects the prognosis of NSCLC. MMP-9 may advance infiltration and metastasis of tumor cells through ILK pathway. In summary, the expression of ILK and MMP9 may play an important role in the evaluation of prognosis for patients with NSCLC.
ObjectiveTo explore the value of matrix metalloproteinase 9 (MMP-9) in predicting the occurrence of heterotopic ossification by observing the expression of MMP-9 in heterotopic ossification of the early trauma rat model. MethodsA total of 132 male Sprague Dawley rats, aged 4-5 weeks, weighing (135.0±6.5) g, were randomly divided into experimental group and control group (n=66). In experimental group, the Achilles tendon was cut off and clamped to prepare heterotopic ossification model; in control group, only Achilles tendon was exposed by making a incision. The general condition of the rats was observed after operation; at 2, 3, 4, 5, 6, 7, and 8 days after operation, the Achilles tendon tissue was harvested for gross observation, histological observation, and immunohistochemical staining observation; the serum and Achilles tendon tissue were harvested to detect the expressions of MMP-9 protein and mRNA by ELISA and RT-PCR. The X-ray films at 5 and 10 weeks and histological examination at 10 weeks after operation were used to observe heterotopic ossification. ResultsAll rats survived to the end of the experiment. The Achilles tendon had no significant change in control group at each time point, showing normal tendon structure. In experimental group, the hardness of Achilles tendon tissue gradually increased with the time; there were a large number of irregular connective tissue and cartilage cells; and immunohistochemical staining for MMP-9 was positive results. The MMP-9 protein and mRNA expression levels of experimental group were significantly higher than those of the control group at each time point (P < 0.05). MMP-9 protein and mRNA expression levels of experimental group showed an increasing tendency (P < 0.05). According to the results of X-ray films and histological observation, heterotopic ossification occurred at 10 weeks after operation in experimental group, but no heterotopic ossification was observed in control group. ConclusionIn early heterotopic ossification of rat Achilles tendon, the expression of MMP-9 increases significantly, indicating that it has reference significance in predicting heterotopic ossification.