AIM: To investigate the anti-tumor effects of nuclear factor-κB (NF-κB) inhibitor SN50 and related mechanisms of SGC7901 human gastric carcinoma cells. METHODS: MTT assay was used to determine the cytotoxic effects ...AIM: To investigate the anti-tumor effects of nuclear factor-κB (NF-κB) inhibitor SN50 and related mechanisms of SGC7901 human gastric carcinoma cells. METHODS: MTT assay was used to determine the cytotoxic effects of SN50 in gastric cancer cell line SGC7901. Hoechst 33258 staining was used to detect apoptosis morphological changes after SN50 treatment. Activation of autophagy was monitored with monodansylcadaverine (MDC) staining after SN50 treatment.Immunofluorescence staining was used to detect the expression of light chain 3 (LC3). Mitochondrial membrane potential was measured using the fluorescent probe JC-1. Western blotting analysis were used to determine the expression of proteins involved in apoptosis and autophagy including p53, p53 upregulated modulator of apoptosis (PUMA), damage-regulated autophagy modulator (DRAM), LC3 and Beclin 1. We detected the effects of p53-mediated autophagy activation on the apoptosis of SGC7901 cells with the p53 inhibitor pifithrin-α. RESULTS: The viability of SGC7901 cells was inhibited after SN50 treatment. Inductions in the expression of apoptotic protein p53 and PUMA as well as autophagic protein DRAM, LC3 and Beclin 1 were detected with Western blotting analysis. SN50-treated cells exhibited punctuate microtubule-associated protein 1 LC3 in immunoreactivity and MDC-labeled vesicles increased after treatment of SN50 by MDC staining. Collapse of mitochondrial membrane potential Δψ were detected for 6 to 24 h after SN50 treatment. SN50-induced increases in PUMA, DRAM, LC3 and Beclin 1 and cell death were blocked by the p53 specific inhibitor pifithrin-α. CONCLUSION: The anti-tumor activity of NF-κB inhibitors is associated with p53-mediated activation of autophagy.展开更多
Objective： The aim of this study was to test the protective effect of mesenchymal stem cells （MSCs） on cardiomyocytes in vitro and to investigate the anti-apoptotic signaling pathway. Methods： MSCs from Sprague-Da...Objective： The aim of this study was to test the protective effect of mesenchymal stem cells （MSCs） on cardiomyocytes in vitro and to investigate the anti-apoptotic signaling pathway. Methods： MSCs from Sprague-Dawley （SD） rats were separated and cultured. MSC medium was collected from MSCs （DMEM） under hypoxia. Cultured cardiomyocytes from neonatal cultured in serum-free Dulbecco＇s modified eagle medium SD rats were exposed to hypoxia/reoxygenation （H/R） and treated with MSC medium. The apoptotic cardiomyocytes were stained with Annexin-V-fluorescein isothiocyanate （FITC）, Hoechst 33342 and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL）. The mitochondrial transmembrane potential of cardiomyocytes was assessed using a fluorescence microscope. The expression of Bcl-2, Bax, cyto- chrome C, apoptosis-induced factor （AIF）, and caspase-3 was tested by Western blot analysis. Results： Our data demonstrated that MSC medium reduced H/R-induced cardiomyocyte apoptosis, increased the Bcl-2/Bax ratio, and reduced the release of cyto- chrome C and AIF from mitochondria into the cytosol. Conclusion： MSCs protected the cardiomyocytes from H/R-induced apoptosis through a mitochondrial pathway in a paracrine manner.展开更多
基金Supported by Health Foundation of Jiangsu Province (H20 0719)the Higher Education Foundation of Jiangsu Province (08KJB320014)+2 种基金the Natural Science Foundation of Jiangsu Province (BK2008168)Suzhou High-Level Talents Project (2008-11)the Science, Education and Health Foundation of Soochow City (SWKQ00814)
文摘AIM: To investigate the anti-tumor effects of nuclear factor-κB (NF-κB) inhibitor SN50 and related mechanisms of SGC7901 human gastric carcinoma cells. METHODS: MTT assay was used to determine the cytotoxic effects of SN50 in gastric cancer cell line SGC7901. Hoechst 33258 staining was used to detect apoptosis morphological changes after SN50 treatment. Activation of autophagy was monitored with monodansylcadaverine (MDC) staining after SN50 treatment.Immunofluorescence staining was used to detect the expression of light chain 3 (LC3). Mitochondrial membrane potential was measured using the fluorescent probe JC-1. Western blotting analysis were used to determine the expression of proteins involved in apoptosis and autophagy including p53, p53 upregulated modulator of apoptosis (PUMA), damage-regulated autophagy modulator (DRAM), LC3 and Beclin 1. We detected the effects of p53-mediated autophagy activation on the apoptosis of SGC7901 cells with the p53 inhibitor pifithrin-α. RESULTS: The viability of SGC7901 cells was inhibited after SN50 treatment. Inductions in the expression of apoptotic protein p53 and PUMA as well as autophagic protein DRAM, LC3 and Beclin 1 were detected with Western blotting analysis. SN50-treated cells exhibited punctuate microtubule-associated protein 1 LC3 in immunoreactivity and MDC-labeled vesicles increased after treatment of SN50 by MDC staining. Collapse of mitochondrial membrane potential Δψ were detected for 6 to 24 h after SN50 treatment. SN50-induced increases in PUMA, DRAM, LC3 and Beclin 1 and cell death were blocked by the p53 specific inhibitor pifithrin-α. CONCLUSION: The anti-tumor activity of NF-κB inhibitors is associated with p53-mediated activation of autophagy.
基金supported by the National Natural Science Foundation of China (No.30670868)the Natural Science Foundation of Zheji-ang Province,China (No.R206007)
文摘Objective： The aim of this study was to test the protective effect of mesenchymal stem cells （MSCs） on cardiomyocytes in vitro and to investigate the anti-apoptotic signaling pathway. Methods： MSCs from Sprague-Dawley （SD） rats were separated and cultured. MSC medium was collected from MSCs （DMEM） under hypoxia. Cultured cardiomyocytes from neonatal cultured in serum-free Dulbecco＇s modified eagle medium SD rats were exposed to hypoxia/reoxygenation （H/R） and treated with MSC medium. The apoptotic cardiomyocytes were stained with Annexin-V-fluorescein isothiocyanate （FITC）, Hoechst 33342 and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL）. The mitochondrial transmembrane potential of cardiomyocytes was assessed using a fluorescence microscope. The expression of Bcl-2, Bax, cyto- chrome C, apoptosis-induced factor （AIF）, and caspase-3 was tested by Western blot analysis. Results： Our data demonstrated that MSC medium reduced H/R-induced cardiomyocyte apoptosis, increased the Bcl-2/Bax ratio, and reduced the release of cyto- chrome C and AIF from mitochondria into the cytosol. Conclusion： MSCs protected the cardiomyocytes from H/R-induced apoptosis through a mitochondrial pathway in a paracrine manner.
