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AIM:To investigate the role of miR-125b in regulating the sensitivity of CD133+ colorectal cancer cells to cisplatin.METHODS:The expression of miR-125b was detected by RT-qPCR in the routine SW480 cells and CD133+ SW480 cells.Flow cytometry analysis was performed to measure the percentage of CD133+ cell population in the SW480 cell line treated with miR-125b and cisplatin.MTT assay was performed to evaluate the effect of miR-125b on the cisplatin-induced cell death in the CD133+ SW480 cells.Bioinformatics and Western blot were performed to determine whether the expression of HAX-1 was regulated by miR-125b.JC-1 staining, Annexin V staining and Western blot analysis were used to study the pathway of apoptosis in the CD133+ SW480 cells co-treated with miR-125b and cisplatin.RESULTS:The expression of miR-125b was significantly lower in the CD133+ SW480 cells than that in the routine SW480 cells and normal colonic epithelial FHC cells.Treatment with cisplatin alone increased the percentage of CD133+ SW480 cell population.However, miR-125b significantly inhibited the enrichment of CD133+ cell population induced by cisplatin.In addition, the results of MTT assay showed that the anti-tumor effect of cisplatin was significantly enhanced when the miR-125b was transfected into the CD133+ SW480 cells.The results of Western blot indicated that the HAX-1 gene was the target of miR-125b.Furthermore, the apoptosis induced by the combination of miR-125b and cisplatin was dependent on the dysfunction of mitochondrial membrane, leading to the release of cytochrome C into the cytoplasm and the subsequently activation of apoptosis in the CD133+ SW480 cells.CONCLUSION:miR-125b increased the sensitivity of CD133+ colo-rectal cancer cells to cisplatin by down-regulating the expression of HAX-1.
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Objective To evaluate the accuracy of the new and old electrocardiographic algorithms for differentiating the origins of outflow tract ventricular arrhythmias. Methods The clinical data of 202 patients treated between 2010 and 2013 were retrospectively reviewed for the investigations of the four algorithms including the transitional zone index, the V2 transition ratio, V2 R-wave duration and R/S-wave amplitude indices and the Sv2/Rv3 index. Results Regardless of rotation, the V2 transition ratio had the highest sensitivity (93.5%), while the Sv2/Rv3 index had the highest specificity (93.8). The maximal area under the ROC curve of four was more than 0.8, while the transitional zone index had the minimal area (0.804) with statistical significance (P 0.05). Conclusion Regardless of rotation, the Sv2/Rv3 index has the highest specificity and equal diagnostic value, with equal diagnostic value of the V2 transition ratio and V2 R-wave duration and R/S-wave amplitude indices. Compared with other algorithms, the Sv2/Rv3 index is simple and can be a complement as well for the direction of ablation therapy.
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<p><b>OBJECTIVES</b>To detect the level of matrix metalloproteinase-2 (MMP-2) mRNA and the tissue inhibitor of metalloproteinase-2 (TIMP-2) mRNA in bladder transitional cell carcinoma (BTCC), and to estimate the prognosis for bladder tumor based on the quality and quantity of MMP-2 and TIMP-2 mRNA.</p><p><b>METHODS</b>Thirty-five samples of human BTCC and 15 normal fresh bladder tissues were studied by RT-PCR analysis followed by computer-assisted image analysis.</p><p><b>RESULTS</b>The level of the MMP-2 mRNA in BTCC was significantly increased compared with that in normal bladder epithelium. The positive rates of MMP-2 and TIMP-2 mRNA were 71.4% and 65.7% in BTCC, and 66.7% and 60.0% in the normal bladder wall. The expression intensity of the MMP-2 mRNA by image analysis tended to increase with tumor grading and staging, which showed statistical significance. Similarly, the MMP-2 to TIMP-2 ratio also showed statistically significant difference between normal bladder tissue and bladder carcinoma (P < 0.01).</p><p><b>CONCLUSIONS</b>A high level of the MMP-2 mRNA exists in BTCC, which may function to damage collagen IV inside the basement membrane and the extracellular basement of the bladder. The level of the MMP-2 mRNA is proportional to BTCC grading and staging, which may have prognostic value. The MMP-2 to TIMP-2 ratio may play a more significant role in the determination of the aggressiveness and prognosis of bladder tumor.</p>
Subject(s)
Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Gene Expression Regulation, Neoplastic , Matrix Metalloproteinase 2 , Genetics , Neoplasm Staging , Predictive Value of Tests , Prognosis , RNA, Messenger , Genetics , Metabolism , Tissue Inhibitor of Metalloproteinase-2 , Genetics , Urinary Bladder , Metabolism , Pathology , Urinary Bladder Neoplasms , Genetics , PathologyABSTRACT
Objective To study the effects of transforming growth factor-β1 (TGF-β1) and transforming growth factor-α (TGF-α) on the growth and invasive potentials in human bladder tumor cells. Methods The effects of TGF-β1 and TGF-α on the growth of EJ cell line and the effects on MMPs and TIMP-2 were studied by means of MTT,Western Blot and RT-PCR methods. Results (1)TGF-β1 and TGF-α tended to inhibit the growth of EJ cells but statistically nonsigficant.(2)Higher levles of MMP-9 mRNA but lower levels of MMP-2,TIMP-2,MT1-MMP mRNA were found in EJ cells following the treatment with TGF-β1.The same was true for the expression of MMP-2,TIMP-2 mRNA in TGF-α groups.However,MMP-9 mRNA was not found in both TGF-α groups and the control groups. (3)TGF-β1 (0.1,1.0 ng/ml) enhanced MMP-2 protein but not the TIMP-2 protein,while TGF-β1 (5.0,10.0 ng/ml)decreased TIMP-2 protein but not on MMP-2.In TGF-α groups,when the concentration was 1.0,5.0,10.0ng/ml,TIMP-2 protein expression was decreased but MMP-2 did not.When the concentration reached 100.0 ng/ml,it increased MMP-2 protein level,not the TIMP-2 protein. Conclusions TGF-β1 and TGFα do not inhibit the proliferation of EJ cells whereas the enhanced-invasiveness and metastasis may be associated with regulating the expression of MMPs and TIMP-2.
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Objective To detect both the protein and gene expression of matrix metalloproteinase (MMP 2,MMP 9) and its tissue inhibitor (TIMP 2) in tissues of bladder transitional cell carcinoma (BTCC), and to study the correlation with the grading and staging of the neoplasm and with the prognosis. Methods 41 human BTCC ,15 normal bladder tissue were studied by Western Blot and RT PCR analysis followed by computer assisted image analysis in order to detect the A values of their expression. Results In BTCC, the A value of MMP 2, MMP 9 were increased significantly as compared to normal bladder epithelium. There was no statistical significant difference of A value of TIMP 2 between normal bladder tissue and bladder cancer tissue. The ratio of MMP 2/TIMP 2 in bladder cancer showed statistical significantly difference as compared with normal bladder tissue. Conclusions This study demonstrated there was a high expression of MMP 2 and MMP 9 in BTCC,by which collagen Ⅳ inside basement membrane of bladder was damaged.However, the ratio of MMP 2/TIMP 2 has more prognostic value than the expression of MMP 2 and MMP 9 alone.
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Objective To study the effects of transforming growth factor ?1 (TGF ?1) and transforming growth factor ? (TGF ?) on the growth and invasive potentials in human bladder tumor cells. Methods The effects of TGF ?1 and TGF ? on the growth of EJ cell line and the effects on MMPs and TIMP 2 were studied by means of MTT,Western Blot and RT PCR methods. Results (1)TGF ?1 and TGF ? tended to inhibit the growth of EJ cells but statistically nonsigficant.(2)Higher levles of MMP 9 mRNA but lower levels of MMP 2,TIMP 2,MT1 MMP mRNA were found in EJ cells following the treatment with TGF ?1.The same was true for the expression of MMP 2,TIMP 2 mRNA in TGF ? groups.However,MMP 9 mRNA was not found in both TGF ? groups and the control groups. (3)TGF ?1 ( 0.1 , 1.0 ng/ml) enhanced MMP 2 protein but not the TIMP 2 protein,while TGF ?1 ( 5.0 , 10.0 ng/ml)decreased TIMP 2 protein but not on MMP 2.In TGF ? groups,when the concentration was 1.0 , 5.0 , 10.0 ng/ml,TIMP 2 protein expression was decreased but MMP 2 did not.When the concentration reached 100.0 ng/ml,it increased MMP 2 protein level,not the TIMP 2 protein. Conclusions TGF ?1 and TGF? do not inhibit the proliferation of EJ cells whereas the enhanced invasiveness and metastasis may be associated with regulating the expression of MMPs and TIMP 2.
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Objective To evaluate the inhibitory effect of endostatin on bladder tumor and to investigate the possible mechanism of the inhibition. Methods (1)By means of MTT method,the effects of endostatin on ECV304 and EJ cells proliferation were studied.(2)Ten nude mice were subcutaneously implanted with EJ bladder tumor cells (1?10 7/ml). After the tumor volume exceeded 100～200 mm 3,the mice were randomized into two groups.One group received recombinant human endostatin (2 mg?kg -1 ?d -1 ) whereas the other group received PBS as control.All the treatment lasted 21 days.(3) After that,the mice were sacrificed and then MMPs and TIMPs mRNA expression were measured in implanted tumor by RT PCR and Western blot method. Results (1)Endostatin inhibits ECV304 proliferation stimulated with bFGF (5 ng/ml).In addition,we report for the first time that endostatin also inhibits EJ cells proliferation.(2)The mean tumor weight of endostatin treated group (0.70?0.16 g) was significantly lower than that of control group (1.14?0.21)g, P