AIM: To explore the mechanism by which H pylori causes activation of gastric epithelial cells. METHODS: A VacA (+) and CagA (+) standard H pylori line NCTC 11637 and a human gastric adenocarcinoma derived gastric epit...AIM: To explore the mechanism by which H pylori causes activation of gastric epithelial cells. METHODS: A VacA (+) and CagA (+) standard H pylori line NCTC 11637 and a human gastric adenocarcinoma derived gastric epithelial cell line BGC-823 were applied in the study. MTT assay and 3H-TdR incorporation test were used to detect the proliferation of BGC-823 cells and Western blotting was used to detect the activity and existence of related proteins. RESULTS: Incubation with H pylori extract increased the proliferation of gastric epithelial cells, reflected by both live cell number and DNA synthesis rate. The activity of extracellular signal-regulated protein kinase (ERK) signal transduction cascade increased within 20 min after in- cubation with H pylori extract and appeared to be a sus- tained event. MAPK/ERK kinase (MEK) inhibitor PD98059 abolished the action of H pylori extract on both ERK activity and cell proliferation. Incubation with H pylori extract increased c-Fos expression and SRE-dependent gene expression. H pylori extract caused phosphorylation of several proteins including a protein with molecular size of 97.4 kDa and tyrosine kinase inhibitor genistein inhibited the activation of ERK and the proliferation of cells caused by H pylori extract. CONCLUSION: Biologically active elements in H pylori extract cause proliferation of gastric epithelial cells through activating tyrosine kinase and ERK signal trans- duction cascade.展开更多
AIM： To elucidate the localization of RhoA in gastric SGC-7901 cancer cells and its translocation by lysophosphatidic acid （LPA） and/or 8-chlorophenylthio- cAMP （CPT-cAMP）.METHODS： Immunofluorescence microscopy ...AIM： To elucidate the localization of RhoA in gastric SGC-7901 cancer cells and its translocation by lysophosphatidic acid （LPA） and/or 8-chlorophenylthio- cAMP （CPT-cAMP）.METHODS： Immunofluorescence microscopy was used to determine the localization of RhoA. Western blotting was used to detect both endogenous and exogenous RhoA in different cellular compartments （membrane, cytosol, nucleus） and the translocation of RhoA following treatment with LPA, CPT-cAMP, or CPT-cAMP ＋ LPA.RESULTS： Immunofluorescence staining revealed endogenous RhoA to be localized in the membrane, the cytosol, and the nucleus, and its precise localization within the nucleus to be the nucleolus. Western blotting identified both endogenous and exogenous RhoA within different cellular compartments （membrane, oltosol, nucleus, nucleolus）. After stimulation with LPA, the amount of RhoA within membrane and nuclear extracts increased, while it decreased in the cytosol fractions. After treatment with CPT-cAMP the amount of RhoA within the membrane and the nuclear extracts decreased, while it increased within the cytosol fraction. Treatment with a combination of both substances led to a decrease in RhoA in the membrane and the nucleus but to an increase in the cytosol.CONCLUSION： In SGC-7901 cells RhoA was found to be localized within the membrane, the cytosol, and the nucleus. Within the nucleus its precise localization could be demonstrated to be the nucleolus. Stimulation with LPA caused a translocation of RhoA from the cytosol towards the membrane and the nucleus; treatment with CPT-cAMP caused the opposite effect. Furthermore, pre-treatment with CPT-cAMP was found to block the effect of LPA.展开更多
AIM： To elucidate the localization of vasodilator stimulated phosphoprotein （VASP）, a cytoskeletal organizing protein and a substrate of protein kinases A and G in mitotic gastric cancer cells. METHODS： Immunofluo...AIM： To elucidate the localization of vasodilator stimulated phosphoprotein （VASP）, a cytoskeletal organizing protein and a substrate of protein kinases A and G in mitotic gastric cancer cells. METHODS： Immunofluorescence microscopy was used to observe the localization of α-tubulin, VASP and Ser157 phosphorylated VASP （p-VASP） in interphase of mitotic gastric cancer of the cell line SGC-7901. RESULTS： Immunofluorescence staining showed that p-VASP but not VASP was co-localized with α-tubulin on spindle poles and fibers in prophase, metaphase and anaphase of the mitotic process of the gastric cancer cell line SGC-7901. H89, an inhibitor of protein kinases A and G, had no effect on the localization of p-VASP on the spindles. CONCLUSION： VASP may play a role in assembling and stabilizing the mitotic spindle of cells, and phosphorylation of the protein is the precondition for it to exert this function.展开更多
基金Supported by the National Natural Science Foundation of China, No. 30340036 and No. 30470891 Grant from Jiangsu University and Zhenjiang Key Institute of Clinical Laboratory Medicine (SH2006066)
文摘AIM: To explore the mechanism by which H pylori causes activation of gastric epithelial cells. METHODS: A VacA (+) and CagA (+) standard H pylori line NCTC 11637 and a human gastric adenocarcinoma derived gastric epithelial cell line BGC-823 were applied in the study. MTT assay and 3H-TdR incorporation test were used to detect the proliferation of BGC-823 cells and Western blotting was used to detect the activity and existence of related proteins. RESULTS: Incubation with H pylori extract increased the proliferation of gastric epithelial cells, reflected by both live cell number and DNA synthesis rate. The activity of extracellular signal-regulated protein kinase (ERK) signal transduction cascade increased within 20 min after in- cubation with H pylori extract and appeared to be a sus- tained event. MAPK/ERK kinase (MEK) inhibitor PD98059 abolished the action of H pylori extract on both ERK activity and cell proliferation. Incubation with H pylori extract increased c-Fos expression and SRE-dependent gene expression. H pylori extract caused phosphorylation of several proteins including a protein with molecular size of 97.4 kDa and tyrosine kinase inhibitor genistein inhibited the activation of ERK and the proliferation of cells caused by H pylori extract. CONCLUSION: Biologically active elements in H pylori extract cause proliferation of gastric epithelial cells through activating tyrosine kinase and ERK signal trans- duction cascade.
基金The National Natural Science Foundation of China,No.30340036and No.30470891Natural Science Foundation of Jiangsu Province,No.BK2007091Grant from Jiangsu University and Zhenjiang Key Institute of Clinical Laboratory Medicine
文摘AIM： To elucidate the localization of RhoA in gastric SGC-7901 cancer cells and its translocation by lysophosphatidic acid （LPA） and/or 8-chlorophenylthio- cAMP （CPT-cAMP）.METHODS： Immunofluorescence microscopy was used to determine the localization of RhoA. Western blotting was used to detect both endogenous and exogenous RhoA in different cellular compartments （membrane, cytosol, nucleus） and the translocation of RhoA following treatment with LPA, CPT-cAMP, or CPT-cAMP ＋ LPA.RESULTS： Immunofluorescence staining revealed endogenous RhoA to be localized in the membrane, the cytosol, and the nucleus, and its precise localization within the nucleus to be the nucleolus. Western blotting identified both endogenous and exogenous RhoA within different cellular compartments （membrane, oltosol, nucleus, nucleolus）. After stimulation with LPA, the amount of RhoA within membrane and nuclear extracts increased, while it decreased in the cytosol fractions. After treatment with CPT-cAMP the amount of RhoA within the membrane and the nuclear extracts decreased, while it increased within the cytosol fraction. Treatment with a combination of both substances led to a decrease in RhoA in the membrane and the nucleus but to an increase in the cytosol.CONCLUSION： In SGC-7901 cells RhoA was found to be localized within the membrane, the cytosol, and the nucleus. Within the nucleus its precise localization could be demonstrated to be the nucleolus. Stimulation with LPA caused a translocation of RhoA from the cytosol towards the membrane and the nucleus; treatment with CPT-cAMP caused the opposite effect. Furthermore, pre-treatment with CPT-cAMP was found to block the effect of LPA.
基金Supported by National Natural Science Foundation of China, No. 30340036 and 30470891 Startup Grant from Jiangsu University, and Grant of Zhenjiang Key Institute of Clinical Laboratory Medicine (SH2006066)
文摘AIM： To elucidate the localization of vasodilator stimulated phosphoprotein （VASP）, a cytoskeletal organizing protein and a substrate of protein kinases A and G in mitotic gastric cancer cells. METHODS： Immunofluorescence microscopy was used to observe the localization of α-tubulin, VASP and Ser157 phosphorylated VASP （p-VASP） in interphase of mitotic gastric cancer of the cell line SGC-7901. RESULTS： Immunofluorescence staining showed that p-VASP but not VASP was co-localized with α-tubulin on spindle poles and fibers in prophase, metaphase and anaphase of the mitotic process of the gastric cancer cell line SGC-7901. H89, an inhibitor of protein kinases A and G, had no effect on the localization of p-VASP on the spindles. CONCLUSION： VASP may play a role in assembling and stabilizing the mitotic spindle of cells, and phosphorylation of the protein is the precondition for it to exert this function.
