【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 evaluate the effect of smooth muscle cell transplantation on myocardial interstitial reconstruction shortly after myocardial infarction. Methods A total of 48 female Wister rats were randomly divided into two groups with the random number table, the control group (n=24) and the smooth muscle cell transplantation group (n=24). The left coronary artery was ligated to set up the myocardial infarction animal model. An amount of 05 ml phosphate buffered saline(PBS) containing 1×106 smooth muscle cells or 0.5 ml PBS without cells was injected into the injured myocardium immediately. By immunoblot and reverse transcriptionolymerase china reaction (RT-PCR), we observed the amount of protein and mRNA of matrix metalloproteinase2(MMP-2), matrix metalloproteinase-9(MMP-9) and tissue inhibitor of metalloprotease-3 (TIMP-3) in the myocardium of the rats. Results The transplanted smooth muscle cells survived well. Compared with the control group, myocardial TIMP3 mRNA (1.06±0.22 vs. 0.81±0.19, t=－2.358, P=0.033) and protein content (3.33±0.53 vs. 1.63±0.47, t=－6.802, Plt;0.001) were significantly increased in the transplantation group. Myocardial MMP-2, MMP-9 mRNA (0.49±0.12 vs. 1.16±0.18, t=8.453, Plt;0.001; 0.45±0.12 vs. 0.80±0.11, t=5.884, Plt;0.001) and protein content (3.98±1.08 vs. 6.05±0.91, t=4.139, P=0.001; 0.39±0.14 vs. 0.57±0.17, t=2.409, P=0.031) [CM(1585mm]were significantly reduced in the transplantation group compared with the control group. Conclusion transplanted smooth muscle cells can survive well in the infarction myocardium and can increase the amount of myocardial TIMP-3 mRNA and protein content and reduce myocardial MMP-2, MMP-9 mRNA and protein content, which is an effective way to prevent harmful cardiac remodeling.
Objective To explore the expression of matrix metalloproteinase （MMP）-2 in patients with papillary thyroid cancer （PTC） recurrence or residuum and its value of clinical application. Methods The serums from 68 patients with thyroid disease underwent operation and 15 health examinations under stomachs empty from March 2009 to December2009 in this hospital were gathered. Sixty-eight patients with thyroid disease were divided into PTC with lymph node metastasis （LNM） group （n=19），PTC recurrence or residuum with LNM group （n=17），PTC without LNM group （n=10），and benign thyroid disease group （n=22） according to the postoperative pathological findings，15 health examinations as control group. The expression of MMP-2 of serum sample was detected by ELISA method. The difference of the expression of MMP-2 in each group was analyzed. Results The expressions of MMP-2 in the PTC recurrence or residuum with LNM group，PTC with LNM group，PTC without LNM group， benign thyroid disease group，and the control group were （1 724.00±762.24） ng/ml，（1 329.16±776.59） ng/ml，（1 489.61±546.53） ng/ml，（1 264.87±817.27） ng/ml，and （608.43±88.63） ng/ml，respectively. The expressions of MMP-2 in the PTC with LNM group and PTC recurrence or residuum with LNM group were significantly higher than those in the benign thyroid disease group （P＜0.05） and the control group （P＜0.05），respectively，which in the PTC without LNM group was significantly higher than that in the control group （P＜0.05）. There was no significant difference of MMP-2 expression of serum between the benign thyroid disease group and the control group （P＞0.05），which were no significant difference among the other threemalignant disease groups （P＞0.05）. The MMP-2 positive expression rates were 79%，76%，80%，41%，and 20% in the PTC with LNM，PTC recurrence or residue with LNM group，PTC without LNM group，benign thyroid disease group，and control group，respectively. The MMP-2 positive expressions rates of serums were not significantly different among three malignant disease groups （P＞0.05），but which were significantly higher than those of the benign thyroid disease group （P＜0.01） and control group （P＜0.01），respectively. Conclusions The MMP-2 level of serum can be used as an index to judge preoperative thyroid nodules，which can not be use to determine whether PTC metastasis or not.
ObjectiveTo observe the dynamic changes of the concentrations of serum matrix metalloproteinase (MMP)-2 and MMP-9, and to discuss its clinical significance. MethodsFrom January to May 2014, 50 cases of clinically diagnosed cerebral infarction patients were included in the study as the cerebral infarction group, and we randomly selected 30 healthy volunteers at the same time in the same age group as the control group. The serum MMP-2 and MMP-9 of patients with acute cerebral infarction were detected in the onset of 24 hours, 7th day and 14th day respectively, which were compared with the control group accordingly. The patients with cerebral infarction were divided into small infarction group (1.5-3.0 cm), middle infarction group (3.1-5.0 cm) and large infarction group (>5.0 cm) according to the infraction volume. According to neurological functional deficit score they were divided into mild (0-15 points), moderate (16-30 points) and severe group (31-45 points). Changes of the level of MMP-9 and MMP-2 were compared in patients with different cerebral infarction volume and different impairment degree. ResultsFor the cerebral infarction group, the serum MMP-2 and MMP-9 levels were significantly higher in the onset of 24 hours, 7th day and 14th day[MMP-2:(2.36±0.76), (2.86±0.87), and (2.20±0.79) ng/mL; MMP-9:(238.8±99.6), (360.4±141.8), and (152.2±80.4) ng/mL] than the control group[MMP-2:(1.20±0.27) ng/mL; MMP-9:(124.8±28.2) ng/mL] (P<0.05). The larger the infarction volume was in the patients with acute ischemic stroke, the higher the levels of serum MMP-9 and MMP-2. The severer the neurologic impairment degree was in the patients with acute ischemic stroke, the higher the levels of serum MMP-9 and MMP-2 were. ConclusionFor patients with acute cerebral infarction, the levels of serum MMP-2 and MMP-9 are closely related to time of onset, infarct volume and neurological deficits, which can be used as an important basis to estimate the condition and assess the prognosis.
ObjectiveTo investigate expressions and biological function of membrane type matrix metallopro-teinase-1 (MT1-MMP) and matrix metalloproteinase-2 (MMP-2) in papillary thyroid carcinoma. MethodsThe expre-ssions of MT1-MMP and MMP-2 in 164 cases of papillary thyroid carcinoma and paracancerous tissues were detected by immunohistochemistry.The association between the expressions of MT1-MMP and MMP-2 and clinicopathological characteristics of papillary thyroid carcinoma was analyzed. ResultsIn paracancerous tissues, the positive expression rate of MT1-MMP was 11.0% (18/164), and the positive expression rate of MMP-2 was 14.0% (23/164).In papillary thyroid carcinoma tissues, the positive expression rates of MT1-MMP and MMP-2 was 61.6% (101/164) and 67.7% (111/164), respectively.The expressions of MT1-MMP and MMP-2 in carcinoma tissues and para carcinoma tissues were statistically significant differences (P < 0.05).The expressions of MT1-MMP and MMP-2 in papillary thyroid carcinoma tissues correlated with the lymph node metastasis (P < 0.05).In addition, the expression of MMP-2 in papillary thyroid carcinoma tissues correlated with capsule invasion (P < 0.05).The positive correlation was found between the expressions of MT1-MMP and MMP-2 in papillary thyroid carcinoma tissues (r=0.256, P < 0.05). ConclusionsMT1-MMP and MMP-2 may be involved the thyroid capsule invasion and lymph node metastasis of papillary thyroid carcinoma.MT1-MMP and MMP-2 may be involved in the progression of papillary thyroid carcinoma.
ObjectiveTo clarify the relationship between the G/C polymorphism of inflammatory gene matrix metalloproteinase-2 (MMP2) and warfarin therapy after cardiac valve replacement (CVR). MethodsWe finally identified 96 patients who received additional warfarin therapy after CVR as a trial group and 78 patients without the warfarin therapy as a control group. Gene sequencing techniques were adopted to determine single nucleotide polymorphism allele. We analyzed genotype and clinical features of the two groups and explored the relationship between the different MMP2 geno-types and warfarin therapy after CVR. Logistic regression was used to analyze the correlation between genotypes and risk factors after CVR and Kaplan-Meier survival curves were performed to analyze the survival time and efficacy of patients carrying MMP2 GC and GG genotypes. ResultsThe distribution of MMP2 genotype in patients receiving warfarin therapy after surgery was different from that in patients without warfarin therapy. The results of multivariate logistic regression analysis showed that GC and GG genotypes were risk factors of complications of CVR. The proportion of GG genotype was higher in the patients with postoperative complications compared with those without. The survival time of patients carrying genotype MMP2 GG was shorter than those carrying GC genotype (P < 0.05), which reveals that the level of MMP2 GG genotype was associated with the prognosis. ConclusionG allele of MMP2 is a risk factor of complications following CVR. GG genotype is relevant to CVR and prognosis, which can be regarded as a risk factor post CVR.