T signals to the nucleus as well as signals that L optical density of 561024 at 600 nm (OD600 = 561024), and the suspension was regulate cell-matrix connections. Cadherins comprise a large family of cell ell adhesion molecules that include the classical, desmosomal, and D in most HAT patients, since miR-144* has been reported to atypical cadherins. E-cadherin, which is expressed primarily in epithelial cells, is an adhesion protein that is encoded by the CDH1 gene and functions in multiple processes, including development, tissue integrity, cell migration, morphology, and polarity [17,18,19]. Ecadherin is also a tumor suppressor whose expression is frequently reduced or silenced, and its re-expression can induce morphologic reversion [20,21]. The EGF-dependent activation of the EGFR has been reported to be inhibited in an E-cadherin adhesiondependent manner, which inhibits the ligand-dependent activation of diverse receptor tyrosine kinases. N-cadherin, as an invasion promoter, is frequently upregulated. The expression of N-cadherin in epithelial cells induces changes in morphology to a fibroblastic phenotype, rendering the cells more motile and invasive. Recent studies indicate that cancer cells have up-regulated N-cadherin in addition to the loss of E-cadherin. This change in cadherin expression is called the “cadherin switch”. We observed a down-regulation of E-cadherin mRNA and increased phosphorylation, which induces the endocytosis of Ecadherin, in PKM2-depleted cells. We also found that the Ncadherin protein expression level was increased in the BGC823 cell line when PKM2 was depleted. The knockdown of PKMPKM2 Enhanced the Activities of the EGF/EGFR Downstream Signaling Pathways in AGS Cells and was Correlated with ERK Activity in Gastric Cancer SpecimensTo analyze whether the EGFR may be involved in the migration and invasion of AGS cells, these cells were treated with EGF, which binds to the EGFR and activates the downstream signaling pathways. EGF treatment resulted in the phosphorylation of the EGFR and the subsequent activation of the downstream EGFR pathways, including the PLCc1 and ERK1/ 2 pathways (Fig. 4A). We found that the activities of PLCc1 and ERK1/2 were greater in cells where PKM2 was not depleted than in the PKM2-depleted cells after either a short or long (24 h) incubation with EGF. This result is the opposite of what was observed with the BGC823 and SGC7901 cells; in AGS cells, PKM2 came into play as a stimulus and promoted cell migration and invasion. We next investigated MMP7 expression using RTPCR in AGS-sipk cells and the control cells. Treatment with EGF enhanced MMP7 expression at the level of transcription in AGSpu6 cells but not in AGS-sipk cells (Fig. 4B). The activity of ERK1/2 was obviously higher in AGS-pu6 cells compared with AGS-sipk cells after 0 h and 24 h treatment with EGF (Fig. 4A). We next performed immunohistochemical (IHC) analyses to examine E-cadherin expression, PKM2 localization and ERK1/2 phosphorylation in serial sections of 15 human gastric cancer specimens using antibodies with validated specificities. Figure 4C shows that the levels of E-cadherin expression, ERK1/2 phosphorylation, and cytoplasmic PKM2 expression were correPkM2 Regulates the EGF/EGFR SignalFigure 3. Depletion of PKM2 attenuated the motility of AGS cells and the functional changes after rescuing PKM2 in gastric cancer cell lines. (A) E-cadherin expression levels were detected by immunoblot analysis in BGC823, SGC7901 and AGS cells. (B) A cross-shaped wound was created in the monolayer, and the AGS stable cells were cultured for an additional 24 h wi.T signals to the nucleus as well as signals that regulate cell-matrix connections. Cadherins comprise a large family of cell ell adhesion molecules that include the classical, desmosomal, and atypical cadherins. E-cadherin, which is expressed primarily in epithelial cells, is an adhesion protein that is encoded by the CDH1 gene and functions in multiple processes, including development, tissue integrity, cell migration, morphology, and polarity [17,18,19]. Ecadherin is also a tumor suppressor whose expression is frequently reduced or silenced, and its re-expression can induce morphologic reversion [20,21]. The EGF-dependent activation of the EGFR has been reported to be inhibited in an E-cadherin adhesiondependent manner, which inhibits the ligand-dependent activation of diverse receptor tyrosine kinases. N-cadherin, as an invasion promoter, is frequently upregulated. The expression of N-cadherin in epithelial cells induces changes in morphology to a fibroblastic phenotype, rendering the cells more motile and invasive. Recent studies indicate that cancer cells have up-regulated N-cadherin in addition to the loss of E-cadherin. This change in cadherin expression is called the “cadherin switch”. We observed a down-regulation of E-cadherin mRNA and increased phosphorylation, which induces the endocytosis of Ecadherin, in PKM2-depleted cells. We also found that the Ncadherin protein expression level was increased in the BGC823 cell line when PKM2 was depleted. The knockdown of PKMPKM2 Enhanced the Activities of the EGF/EGFR Downstream Signaling Pathways in AGS Cells and was Correlated with ERK Activity in Gastric Cancer SpecimensTo analyze whether the EGFR may be involved in the migration and invasion of AGS cells, these cells were treated with EGF, which binds to the EGFR and activates the downstream signaling pathways. EGF treatment resulted in the phosphorylation of the EGFR and the subsequent activation of the downstream EGFR pathways, including the PLCc1 and ERK1/ 2 pathways (Fig. 4A). We found that the activities of PLCc1 and ERK1/2 were greater in cells where PKM2 was not depleted than in the PKM2-depleted cells after either a short or long (24 h) incubation with EGF. This result is the opposite of what was observed with the BGC823 and SGC7901 cells; in AGS cells, PKM2 came into play as a stimulus and promoted cell migration and invasion. We next investigated MMP7 expression using RTPCR in AGS-sipk cells and the control cells. Treatment with EGF enhanced MMP7 expression at the level of transcription in AGSpu6 cells but not in AGS-sipk cells (Fig. 4B). The activity of ERK1/2 was obviously higher in AGS-pu6 cells compared with AGS-sipk cells after 0 h and 24 h treatment with EGF (Fig. 4A). We next performed immunohistochemical (IHC) analyses to examine E-cadherin expression, PKM2 localization and ERK1/2 phosphorylation in serial sections of 15 human gastric cancer specimens using antibodies with validated specificities. Figure 4C shows that the levels of E-cadherin expression, ERK1/2 phosphorylation, and cytoplasmic PKM2 expression were correPkM2 Regulates the EGF/EGFR SignalFigure 3. Depletion of PKM2 attenuated the motility of AGS cells and the functional changes after rescuing PKM2 in gastric cancer cell lines. (A) E-cadherin expression levels were detected by immunoblot analysis in BGC823, SGC7901 and AGS cells. (B) A cross-shaped wound was created in the monolayer, and the AGS stable cells were cultured for an additional 24 h wi.
