Objective Cytotoxin-associated protein （CagA） of H. pylori has been confirmed to be closely associated with gastric inflammation and tumorigenesis, but the mechanism behind it is little understood. In this study, we...Objective Cytotoxin-associated protein （CagA） of H. pylori has been confirmed to be closely associated with gastric inflammation and tumorigenesis, but the mechanism behind it is little understood. In this study, we try to determine roles of CagA＋ strain in activating PI3K/Akt1 signaling pathway, and affecting expression of p21WAF1/CIP1 and p27KIP1, and also in releasing IL-8 in host cells. Methods Akt1 phosphorylation and IL-8 levels of CagA＋ and CagAˉ strain infected AGS cells were detected by ELISAs. Two quantitative RT-PCRs were established to measure p21WAF1/CIP1 and p27KIP1 mRNA levels in the CagA＋ and CagAˉ strain infected cells. LY294002, an inhibitor of PI3K/Akt pathway, was used to define effect of the pathway in IL-8 release. Results CagA＋ strain could induce an obvious elevation of Akt1 phosphorylation in the infected AGS cells while CagAˉ strain failed to do so. The CagA＋ H. pylori strain infected AGS cells showed significant drops both in p21WAF1/CIP1 and p27KIP1 mRNA levels, whereas the CagAˉ H. pylori strain caused a remarkable increase in p21WAF1/CIP1 mRNA without affecting p27KIP1 gene transcription in the AGS cells. Both the CagA＋ and CagAˉ H. pylori strains enabled AGS cells to produce close elevated levels of IL-8, and the LY294002 block resulted in unexpected elevations of IL-8 levels. Conclusion CagA can activate PI3K/Akt1 pathway that plays an inhibitory role in IL-8 release in H. pylori infected AGS cells. Activation of PI3K/Akt1 pathway and subsequent negative regulation of p21^WAF1/CIP1 and p27^KIP1 expression might be involved in CagA-associated carcinogenesis.展开更多
Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, w...Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, we found that EGF treatment could activate Ras-related C3 botulinum toxin substrate 1 （Racl）, PI3K/Akt and p21- actived kinase （PAK1） along with cell migration. Ectopic expression of PAK1 K299R, a dominant negative PAK1 mutant, could largely abolish EGF-induced cell migration. Blocking PI3K/Akt signalling with LY294002 or Akt siRNA remarkably inhibited both EGF-induced PAK1 activation and cell migration. Furthermore, expression of dominant-negative Racl （T17N） could largely block EGF-induced PI3K/Akt-PAK1 activation and cell migration. Interestingly, EGF could induce a significant production of ROS, and N-acetyl-L-cysteine, a scavenger of ROS which abolished the EGF-induced ROS generation, cell migration, as well as activation of PI3K/Akt and PAK, but not Racl. Our study demonstrated that EGF-induced cell migration involves a cascade of signalling events, including activation of Racl, generation of ROS and subsequent activation of PI3K/Akt and PAK1.展开更多
基金supported by a grant (2008ZZ06) from the National Key Laboratory for Diagnosis and Treatment of Infectious Diseases of China
文摘Objective Cytotoxin-associated protein （CagA） of H. pylori has been confirmed to be closely associated with gastric inflammation and tumorigenesis, but the mechanism behind it is little understood. In this study, we try to determine roles of CagA＋ strain in activating PI3K/Akt1 signaling pathway, and affecting expression of p21WAF1/CIP1 and p27KIP1, and also in releasing IL-8 in host cells. Methods Akt1 phosphorylation and IL-8 levels of CagA＋ and CagAˉ strain infected AGS cells were detected by ELISAs. Two quantitative RT-PCRs were established to measure p21WAF1/CIP1 and p27KIP1 mRNA levels in the CagA＋ and CagAˉ strain infected cells. LY294002, an inhibitor of PI3K/Akt pathway, was used to define effect of the pathway in IL-8 release. Results CagA＋ strain could induce an obvious elevation of Akt1 phosphorylation in the infected AGS cells while CagAˉ strain failed to do so. The CagA＋ H. pylori strain infected AGS cells showed significant drops both in p21WAF1/CIP1 and p27KIP1 mRNA levels, whereas the CagAˉ H. pylori strain caused a remarkable increase in p21WAF1/CIP1 mRNA without affecting p27KIP1 gene transcription in the AGS cells. Both the CagA＋ and CagAˉ H. pylori strains enabled AGS cells to produce close elevated levels of IL-8, and the LY294002 block resulted in unexpected elevations of IL-8 levels. Conclusion CagA can activate PI3K/Akt1 pathway that plays an inhibitory role in IL-8 release in H. pylori infected AGS cells. Activation of PI3K/Akt1 pathway and subsequent negative regulation of p21^WAF1/CIP1 and p27^KIP1 expression might be involved in CagA-associated carcinogenesis.
基金supported by grants from the National Natural Science Foundation of China (No. 30872926)the Program for AdvancedTalents within Six Industries of Jiangsu Province (08-D) to Dr. Luo Gu+1 种基金the Science Development Foundation of Nanjing Medical University (No. 2010NJMUZ35)the Research Program funded by Schoolof Basic Medical Science, Nanjing Medical University to Dr. Jun Du
文摘Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, we found that EGF treatment could activate Ras-related C3 botulinum toxin substrate 1 （Racl）, PI3K/Akt and p21- actived kinase （PAK1） along with cell migration. Ectopic expression of PAK1 K299R, a dominant negative PAK1 mutant, could largely abolish EGF-induced cell migration. Blocking PI3K/Akt signalling with LY294002 or Akt siRNA remarkably inhibited both EGF-induced PAK1 activation and cell migration. Furthermore, expression of dominant-negative Racl （T17N） could largely block EGF-induced PI3K/Akt-PAK1 activation and cell migration. Interestingly, EGF could induce a significant production of ROS, and N-acetyl-L-cysteine, a scavenger of ROS which abolished the EGF-induced ROS generation, cell migration, as well as activation of PI3K/Akt and PAK, but not Racl. Our study demonstrated that EGF-induced cell migration involves a cascade of signalling events, including activation of Racl, generation of ROS and subsequent activation of PI3K/Akt and PAK1.
