AIM: To demonstrate that (-)-Epigallocatechin-3-gallate (EGCG) inhibits vascular endothelial growth factor (VEGF) expression and angiogenesis induced by interleukin-6 (IL-6) via suppressing signal transducer and activ...AIM: To demonstrate that (-)-Epigallocatechin-3-gallate (EGCG) inhibits vascular endothelial growth factor (VEGF) expression and angiogenesis induced by interleukin-6 (IL-6) via suppressing signal transducer and activator of transcription 3 (Stat3) activity in gastric cancer. METHODS: Human gastric cancer (AGS) cells were treated with IL-6 (50 ng/mL) and EGCG at different concentrations. VEGF, total Stat3 and activated Stat3 protein levels in the cell lyses were examined by Western blotting, VEGF protein level in the conditionedmedium was measured by enzyme-linked immunosorbent assay, and the level of VEGF mRNA was evaluated by reverse transcription polymerase chain reaction (RTPCR). Stat3 nuclear translocation was determined by Western blotting with nuclear extract, and Stat3-DNA binding activity was examined with Chromatin immunoprecipitation (ChIP) assay. IL-6 induced endothelial cell proliferation was measured with 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazoliumbromide assay, in vitro angiogenesis was determined with endothelial cell tube formation assay in Matrigel, and IL-6-induced angiogenesis in vitro was measured with Matrigel plug assay. RESULTS: There was a basal expression and secretion of VEGF in AGS cells. After stimulation with IL-6, VEGF expression was apparently up-regulated and a 2.4-fold increase was observed. VEGF secretion in the conditioned medium was also increased by 2.8 folds. When treated with EGCG, VEGF expression and secretion were dose-dependently decreased. IL-6 also increased VEGF mRNA expression by 3.1 folds. EGCG treatment suppressed VEGF mRNA expression in a dose-dependent manner. EGCG dose-dependently inhibited Stat3 activation induced by IL-6, but did not change the total Stat3 expression. When treated with EGCG or AG490, VEGF expressions were reduced to the level or an even lower level in the tumor cells not stimulated with IL-6. However, PD98059 and LY294002 did not change VEGF expression induced by IL-6. EGCG inhibited Stat3 nucleus translocation, and Stat3-DNA binding activity was also markedly decreased by EGCG. Furthermore, EGCG inhibited IL-6 induced vascular endothelial cell proliferation and tube formation in vitro and angiogenesis in vitro . CONCLUSION: EGCG inhibits IL-6-induced VEGF expression and angiogenesis via suppressing Stat3 activity in gastric cancer, which has provided a novel mechanistic insight into the anti-angiogenic activity of EGCG.展开更多
We aimed to investigate the impact of X-rays on miR-130a and miR-25 expressions of A549 cell lines and to understand the mechanism of miR-130a and miR-25 on the regulation of invasion and metastasis of A549 cell lines...We aimed to investigate the impact of X-rays on miR-130a and miR-25 expressions of A549 cell lines and to understand the mechanism of miR-130a and miR-25 on the regulation of invasion and metastasis of A549 cell lines. Human adenocarcinoma cells of the line A549 were cultured and radiated by X-rays at the absorbed doses of 2 and 4, respectively by linear accelerator. Transwell invasion and migration assay were employed to exam the metastasis ability of A549 cells post X-rays irradiation. Both the miRNA and mRNA were extracted from A549 cells at different time points post radiation. The expressions of miR-130a and miR-25, as well as the expressions of VEGF and CCR-7 mRNAs, were detected in A549 cells with 2 and 4 Gy X-rays radiation, respectively by real time PCR. Results were statistically analyzed by SAS 8.2 software, which showed that the invasiveness of A549 cells post 2 and 4 Gy X-rays increased significantly compared with that of untreated A549 cells (the migration cell numbers are 20, 48 and 62 in Group 0, 2 and 4 Gy X-rays, respectively). Furthermore, the expressions of miR-130a and miR-25 also increased significantly at the time-points of 1, 2, 4, and 8 h post radiation, and began to decrease to the control level at 12 h post radiation. VEGF and CCR-7 mRNAs were detected to be up-regulated 18 h post radiation and remained at a high level till 72 h post radiation. The expression of VEGF mRNA has a close correlation with that of CCR-7 mRNA, and the expression of miR-130a also has a correlation with that of VEGF and CCR-7mRNAs. It is concluded that the metastasis of A549 cells in vitro could be promoted by X-rays, and miR-130a might play a role in the metastasis of A549 cells via regulating the expressions of VEGF and CCR-7 mRNAs.展开更多
基金Supported by National Natural Science Foundation of China, Grant, No. 30571833Natural Science Foundation of Guangdong Province, 05001785China Postdoctoral Science Foundation 20100470963
文摘AIM: To demonstrate that (-)-Epigallocatechin-3-gallate (EGCG) inhibits vascular endothelial growth factor (VEGF) expression and angiogenesis induced by interleukin-6 (IL-6) via suppressing signal transducer and activator of transcription 3 (Stat3) activity in gastric cancer. METHODS: Human gastric cancer (AGS) cells were treated with IL-6 (50 ng/mL) and EGCG at different concentrations. VEGF, total Stat3 and activated Stat3 protein levels in the cell lyses were examined by Western blotting, VEGF protein level in the conditionedmedium was measured by enzyme-linked immunosorbent assay, and the level of VEGF mRNA was evaluated by reverse transcription polymerase chain reaction (RTPCR). Stat3 nuclear translocation was determined by Western blotting with nuclear extract, and Stat3-DNA binding activity was examined with Chromatin immunoprecipitation (ChIP) assay. IL-6 induced endothelial cell proliferation was measured with 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazoliumbromide assay, in vitro angiogenesis was determined with endothelial cell tube formation assay in Matrigel, and IL-6-induced angiogenesis in vitro was measured with Matrigel plug assay. RESULTS: There was a basal expression and secretion of VEGF in AGS cells. After stimulation with IL-6, VEGF expression was apparently up-regulated and a 2.4-fold increase was observed. VEGF secretion in the conditioned medium was also increased by 2.8 folds. When treated with EGCG, VEGF expression and secretion were dose-dependently decreased. IL-6 also increased VEGF mRNA expression by 3.1 folds. EGCG treatment suppressed VEGF mRNA expression in a dose-dependent manner. EGCG dose-dependently inhibited Stat3 activation induced by IL-6, but did not change the total Stat3 expression. When treated with EGCG or AG490, VEGF expressions were reduced to the level or an even lower level in the tumor cells not stimulated with IL-6. However, PD98059 and LY294002 did not change VEGF expression induced by IL-6. EGCG inhibited Stat3 nucleus translocation, and Stat3-DNA binding activity was also markedly decreased by EGCG. Furthermore, EGCG inhibited IL-6 induced vascular endothelial cell proliferation and tube formation in vitro and angiogenesis in vitro . CONCLUSION: EGCG inhibits IL-6-induced VEGF expression and angiogenesis via suppressing Stat3 activity in gastric cancer, which has provided a novel mechanistic insight into the anti-angiogenic activity of EGCG.
基金supported by the National Natural Science Foundation of China (Grant No. 8117230)
文摘We aimed to investigate the impact of X-rays on miR-130a and miR-25 expressions of A549 cell lines and to understand the mechanism of miR-130a and miR-25 on the regulation of invasion and metastasis of A549 cell lines. Human adenocarcinoma cells of the line A549 were cultured and radiated by X-rays at the absorbed doses of 2 and 4, respectively by linear accelerator. Transwell invasion and migration assay were employed to exam the metastasis ability of A549 cells post X-rays irradiation. Both the miRNA and mRNA were extracted from A549 cells at different time points post radiation. The expressions of miR-130a and miR-25, as well as the expressions of VEGF and CCR-7 mRNAs, were detected in A549 cells with 2 and 4 Gy X-rays radiation, respectively by real time PCR. Results were statistically analyzed by SAS 8.2 software, which showed that the invasiveness of A549 cells post 2 and 4 Gy X-rays increased significantly compared with that of untreated A549 cells (the migration cell numbers are 20, 48 and 62 in Group 0, 2 and 4 Gy X-rays, respectively). Furthermore, the expressions of miR-130a and miR-25 also increased significantly at the time-points of 1, 2, 4, and 8 h post radiation, and began to decrease to the control level at 12 h post radiation. VEGF and CCR-7 mRNAs were detected to be up-regulated 18 h post radiation and remained at a high level till 72 h post radiation. The expression of VEGF mRNA has a close correlation with that of CCR-7 mRNA, and the expression of miR-130a also has a correlation with that of VEGF and CCR-7mRNAs. It is concluded that the metastasis of A549 cells in vitro could be promoted by X-rays, and miR-130a might play a role in the metastasis of A549 cells via regulating the expressions of VEGF and CCR-7 mRNAs.
