The cytosolic liver-specific growth factor-hepatic stimulator substance (HSS) has been shown to be able to amplify the rat hepatocyte proliferation responded to EGF. In order to get more insight into the mechanism, th...The cytosolic liver-specific growth factor-hepatic stimulator substance (HSS) has been shown to be able to amplify the rat hepatocyte proliferation responded to EGF. In order to get more insight into the mechanism, the regulatory effect of HSS on EGF-receptor(EGF-R) and the receptor phosphorylation at molecular level was studied. HSS partially purified from weanling rat liver was given to cultured hepatocytes and its influence on EGF-R specific binding and internalization as well as mRNA expression were investigated. The results showed that preincubation of hepatocytes with HSS could lead to an increase in [125I]-EGF binding to its receptors and inhibit EGFinduced receptor down-regulation. Furthermore, the overexpression of EGF-R mRNA stimulated by HSS was seen during 2-12 h after the incubation. Additionally, it was demonstrated with human hepatoma sMMC-7721 cells in Western blot that the EGF-R expression and the receptor autophosphorylation were increased with dose/timedependency after HSS treatment. These results strongly suggest that the mechanism of HSS action on hepatocyte growth might be related to its modulation on EGF-R and receptor-mediated signaling transduction.展开更多
Since the epidermal growth factor receptor (EGFR) is a key regulator in cell signaling pathways of cancer cell. To investigate the mechanism between cancer cells survival and its EGFR expression, drug selection of can...Since the epidermal growth factor receptor (EGFR) is a key regulator in cell signaling pathways of cancer cell. To investigate the mechanism between cancer cells survival and its EGFR expression, drug selection of cancer cells target therapy, we generated a cell line, 9L-EGFR, which stably expressed human EGFR; the parental rat glioma cell line, 9L, does not contain endogenous EGFR message or protein. Our results show that 9L-EGFR cells had high levels of EGFR on their cell surface by using RT-PCR, Western analysis and Flow cytometry analysis. The EGFR transfected into 9L cells was capable of being activated by EGF, in which either phosphorylated (p-EGFR) or total (EGFR) was showed by Western blot. This investigation may contribute to the further studies of cancer cells bearing EGFR.展开更多
Epidermal growth factor （EGF） induced intracellular free calcium ([Ca2+]i) response was studied in fura-2- or fluo-3-loaded human hepatoma cells of BEL-7404 cell line. Single cell [Ca2+]i analysis and [Ca2+], measur...Epidermal growth factor （EGF） induced intracellular free calcium ([Ca2+]i) response was studied in fura-2- or fluo-3-loaded human hepatoma cells of BEL-7404 cell line. Single cell [Ca2+]i analysis and [Ca2+], measurement in cell populations revealed that EGF triggered a rapid [Ca2+]iincrease in the dose-dependent and time- dependent manner. Pretreatment of cells with an endoplasmic reticulum （ER） Ca2+-ATPase inhibitor, thapsigargin （TG） at 100 nM concentration for 20 min, completely abolished EGF-induced [Ca2+]i increase, and chelating extracellular calcium by excess EGTA partially inhibited the increase.Furthermore, the expression of antisense EGF receptor sequence in BEL-7404 cells suppressed the [Ca2+]i response to EGF. The results suggest that EGF receptor-mediated [Ca2+]i increase in the human hepatoma cells is essentially dependent on the Ca2+ storage in ER.展开更多
Three anti-EGF receptor MoAbs were used in these studies. Administration of MoAbs 3 and 176 inhibited tumor formation in nude mice by CNE-2, a poorly differentiated nasopharyngeal carcinoma cell line and A431, an epid...Three anti-EGF receptor MoAbs were used in these studies. Administration of MoAbs 3 and 176 inhibited tumor formation in nude mice by CNE-2, a poorly differentiated nasopharyngeal carcinoma cell line and A431, an epidermoid carcinoma cell line. When the same MoAbs were used in treatment against HeLa, a cervical carcinoma, tumor growth was not affected. The number of EGF receptors and apparent dissociation constants for <sup>125</sup>I-EGF on CNE-2 and A431 was 1.3×10<sup>?</sup>/cell (Kd 7.7×10<sup>-8</sup>mol/L) and 1.4×10<sup>6</sup>/cell(Kd 2.4×10<sup>-9</sup>mol/L), respectively. Both MoAbs 3 and 176, capable of competing with EGF for receptor binding, showed significant tumor growth inhibition. MoAb 101 was incapable of blocking the binding of EGF to its receptor, and not as effective as MoAbs 3 and 176 in tumor growth inhibition. Our observation is that the MoAb anti-EGF receptor is cytostatic rather than cytocidal, in vitro against CNE-2 and A431.展开更多
A non-radioisotopic method was developed for the assay of epidermal growth factor receptor (EGFR). A peptide with twenty amino acid residues around Tyr 1173, the major phosphorylation site of EGFR, was cloned as a GST...A non-radioisotopic method was developed for the assay of epidermal growth factor receptor (EGFR). A peptide with twenty amino acid residues around Tyr 1173, the major phosphorylation site of EGFR, was cloned as a GST fusion protein and used as substrate. Anti-phosphoty-rosine monoclonal antibody PY99 was used for the determination of the extent of phosphorylation. Both the specificity and the sensitivity were substantially higher than that of the existing method. Km value of the fusion protein is much lower (10 μmol/L) than that of the synthetic peptide (110 μmol/L). The method can be applied to the measurement of the tyrosine kinase activity of c-erb B2 (Neu/HER2).展开更多
Epidermal growth factor (EGF) is in close relationship with gastric mucosa. Theremarkable effects of EGF on the gastric mucosa are the stimulation of DNA synthe-sis and inhibition of gastric acid secretion. EGF recept...Epidermal growth factor (EGF) is in close relationship with gastric mucosa. Theremarkable effects of EGF on the gastric mucosa are the stimulation of DNA synthe-sis and inhibition of gastric acid secretion. EGF receptors have been found in gas-tric glands isolated from some animals, which suggests that the way in which EGFinteracts with gastric mucosal cells is via binding to EGF receptors located on theirplasma membrane. The present work first proves the expression of EGF展开更多
The relationship between antiproliferative effect of human IFN γ EGF 3 fusion protein and the influence of EGF receptor binding activity has been studied on A431 cell line. Antiproliferative activity of human IF...The relationship between antiproliferative effect of human IFN γ EGF 3 fusion protein and the influence of EGF receptor binding activity has been studied on A431 cell line. Antiproliferative activity of human IFN γ EGF 3 was higher than that of its parent IFN γ. In the 125 I EGF receptor competition experiment, the inhibition of EGF receptor binding capacity on the target cells was observed in the treatments of human IFN γ or IFN γ EGF 3, but the later was more significant. Our data suggests that the antiproliferative effects by IFN γ and its fusion protein are closely related to their EGF receptor competitions.展开更多
文摘The cytosolic liver-specific growth factor-hepatic stimulator substance (HSS) has been shown to be able to amplify the rat hepatocyte proliferation responded to EGF. In order to get more insight into the mechanism, the regulatory effect of HSS on EGF-receptor(EGF-R) and the receptor phosphorylation at molecular level was studied. HSS partially purified from weanling rat liver was given to cultured hepatocytes and its influence on EGF-R specific binding and internalization as well as mRNA expression were investigated. The results showed that preincubation of hepatocytes with HSS could lead to an increase in [125I]-EGF binding to its receptors and inhibit EGFinduced receptor down-regulation. Furthermore, the overexpression of EGF-R mRNA stimulated by HSS was seen during 2-12 h after the incubation. Additionally, it was demonstrated with human hepatoma sMMC-7721 cells in Western blot that the EGF-R expression and the receptor autophosphorylation were increased with dose/timedependency after HSS treatment. These results strongly suggest that the mechanism of HSS action on hepatocyte growth might be related to its modulation on EGF-R and receptor-mediated signaling transduction.
文摘Since the epidermal growth factor receptor (EGFR) is a key regulator in cell signaling pathways of cancer cell. To investigate the mechanism between cancer cells survival and its EGFR expression, drug selection of cancer cells target therapy, we generated a cell line, 9L-EGFR, which stably expressed human EGFR; the parental rat glioma cell line, 9L, does not contain endogenous EGFR message or protein. Our results show that 9L-EGFR cells had high levels of EGFR on their cell surface by using RT-PCR, Western analysis and Flow cytometry analysis. The EGFR transfected into 9L cells was capable of being activated by EGF, in which either phosphorylated (p-EGFR) or total (EGFR) was showed by Western blot. This investigation may contribute to the further studies of cancer cells bearing EGFR.
文摘Epidermal growth factor （EGF） induced intracellular free calcium ([Ca2+]i) response was studied in fura-2- or fluo-3-loaded human hepatoma cells of BEL-7404 cell line. Single cell [Ca2+]i analysis and [Ca2+], measurement in cell populations revealed that EGF triggered a rapid [Ca2+]iincrease in the dose-dependent and time- dependent manner. Pretreatment of cells with an endoplasmic reticulum （ER） Ca2+-ATPase inhibitor, thapsigargin （TG） at 100 nM concentration for 20 min, completely abolished EGF-induced [Ca2+]i increase, and chelating extracellular calcium by excess EGTA partially inhibited the increase.Furthermore, the expression of antisense EGF receptor sequence in BEL-7404 cells suppressed the [Ca2+]i response to EGF. The results suggest that EGF receptor-mediated [Ca2+]i increase in the human hepatoma cells is essentially dependent on the Ca2+ storage in ER.
文摘Three anti-EGF receptor MoAbs were used in these studies. Administration of MoAbs 3 and 176 inhibited tumor formation in nude mice by CNE-2, a poorly differentiated nasopharyngeal carcinoma cell line and A431, an epidermoid carcinoma cell line. When the same MoAbs were used in treatment against HeLa, a cervical carcinoma, tumor growth was not affected. The number of EGF receptors and apparent dissociation constants for <sup>125</sup>I-EGF on CNE-2 and A431 was 1.3×10<sup>?</sup>/cell (Kd 7.7×10<sup>-8</sup>mol/L) and 1.4×10<sup>6</sup>/cell(Kd 2.4×10<sup>-9</sup>mol/L), respectively. Both MoAbs 3 and 176, capable of competing with EGF for receptor binding, showed significant tumor growth inhibition. MoAb 101 was incapable of blocking the binding of EGF to its receptor, and not as effective as MoAbs 3 and 176 in tumor growth inhibition. Our observation is that the MoAb anti-EGF receptor is cytostatic rather than cytocidal, in vitro against CNE-2 and A431.
基金the fund of Chinese Academy of Sciences (Grant No. KJ952-S1-15).
文摘A non-radioisotopic method was developed for the assay of epidermal growth factor receptor (EGFR). A peptide with twenty amino acid residues around Tyr 1173, the major phosphorylation site of EGFR, was cloned as a GST fusion protein and used as substrate. Anti-phosphoty-rosine monoclonal antibody PY99 was used for the determination of the extent of phosphorylation. Both the specificity and the sensitivity were substantially higher than that of the existing method. Km value of the fusion protein is much lower (10 μmol/L) than that of the synthetic peptide (110 μmol/L). The method can be applied to the measurement of the tyrosine kinase activity of c-erb B2 (Neu/HER2).
基金Project supported by the National Natural Science Foundation of China.
文摘Epidermal growth factor (EGF) is in close relationship with gastric mucosa. Theremarkable effects of EGF on the gastric mucosa are the stimulation of DNA synthe-sis and inhibition of gastric acid secretion. EGF receptors have been found in gas-tric glands isolated from some animals, which suggests that the way in which EGFinteracts with gastric mucosal cells is via binding to EGF receptors located on theirplasma membrane. The present work first proves the expression of EGF
文摘The relationship between antiproliferative effect of human IFN γ EGF 3 fusion protein and the influence of EGF receptor binding activity has been studied on A431 cell line. Antiproliferative activity of human IFN γ EGF 3 was higher than that of its parent IFN γ. In the 125 I EGF receptor competition experiment, the inhibition of EGF receptor binding capacity on the target cells was observed in the treatments of human IFN γ or IFN γ EGF 3, but the later was more significant. Our data suggests that the antiproliferative effects by IFN γ and its fusion protein are closely related to their EGF receptor competitions.
