Modulation of epidermal growth factor receptors by human alpha interferon.

Treatment of Madin-Darby bovine kidney (MDBK) cells with human interferon (IFN)-alpha 2 at 37 degrees C results in a dose-dependent inhibition of cell growth and a reduction of the subsequent binding of 125I-labeled epidermal growth factor (EGF) at 4 degrees C. Human IFN-beta and -gamma, which exhibit little antiviral and antiproliferative activities on MDBK cells, have little effect on cell growth or the binding of 125I-labeled EGF to these cells. The binding of EGF is decreased after exposure to IFN-alpha for greater than 8 hr. Scatchard analyses of the EGF binding data indicate that a 20-hr exposure period results in a decrease in the apparent number of cell-surface EGF receptors and a reduction in the affinity of EGF for its receptor. The rate of internalization of EGF by MDBK cells does not appear to be affected by IFN treatment.     

Decreased expression of hepatic epidermal growth factor receptor gene in diabetic mice

The expression of hepatic epidermal growth factor (EGF) receptor gene was studied in genetically diabetic (C57BL/KsJ db/db) mice and streptozotocin-induced diabetic mice. The binding of 125I-labelled EGF to hepatic membrane preparations of genetically diabetic mice was only 35% of that of non-diabetic mice. Levels of EGF receptor messenger RNAs (10 and 6 kb) in the liver of the diabetic animals were also reduced by about 75%. In streptozotocin-induced diabetic mice, levels of hepatic EGF binding and messenger RNAs for EGF receptor were decreased to 27 and 30% of control levels respectively, at 5 weeks after injection of the drug. There were, however, no significant differences in levels of messenger RNAs for the structural protein beta-actin in the liver. In addition, levels of EGF receptor messenger RNAs in the kidney were similar between control and the two kinds of diabetic mice. Daily administration of insulin to the streptozotocin-induced diabetic mice increased the hepatic levels of EGF receptor messenger RNAs to almost normal levels. These results indicate that EGF binding to its receptor decreases in the liver of diabetic mice, involving alterations in the level of EGF receptor messenger RNAs, and that insulin is important for the regulation of EGF receptor gene expression in the liver but not in the kidney.     

Gonadal and adrenal effects on hepatic epidermal growth factor receptor expression in a murine model

We hypothesize that the actions of epidermal growth factor (EGF) may be modulated by changes in cell surface EGF receptor (EGF-R) expression under endocrine influences. Mouse liver cell membrane preparations were used in a RRA. During ontogenesis, both sexes showed a significant increase (P less than 0.005) in hepatic EGF-R numbers at puberty; however, males demonstrated significantly higher levels than females (P less than 0.005). Gonadectomy of adult males and females resulted in a significant (P less than 0.05) decrease and increase, respectively, in hepatic EGF-R expression. Prepubertal gonadectomy in both sexes resulted in EGF-R levels similar to those observed in adult females. Adrenalectomy of adult animals of both sexes had no effect on hepatic EGF-R numbers, but gonadectomy plus adrenalectomy virtually obliterated EGF-R expression. Short term treatment with testosterone of adult females or gonadectomized female and male mice significantly increased EGF-R numbers (P less than 0.05) to adult male levels. 17 beta-Estradiol given short term to adult males or gonadectomized male and female mice did not significantly alter EGF-R levels. EGF-R expression in androgen-insensitive male mice was significantly reduced (P less than 0.005) to female levels. We conclude that 1) hepatic EGF-R numbers increase post-pubertally in both sexes; 2) hepatic EGF-R expression is significantly stimulated by testosterone, and this effect depends on a functional androgen receptor; 3) the ovary has an inhibitory effect on adult hepatic EGF-R numbers; however, this effect does not appear to be mediated by estrogens; and 4) the adrenal gland has a stimulatory effect on adult hepatic EGF-R expression.     

Over expression of transforming growth factor-alpha and epidermal growth factor receptor in human hepatic cirrhosis tissues

BACKGROUND/AIMS: Transforming growth factor-alpha has 30% amino acid homology to epidermal growth factor and binds with the same membrane-bound receptor, epidermal growth factor receptor. The purpose of this study was to investigate the expression of transforming growth factor-alpha and epidermal growth factor receptor in human hepatic cirrhosis tissues. METHODOLOGY: Expression of transforming growth factor-alpha and epidermal growth factor receptor was evaluated by immunohistochemistry stain in sixty-three hepatic cirrhosis specimens and five normal liver specimens. RESULTS: The transforming growth factor-alpha and epidermal growth factor receptor expression rates were 84.1% (53/63) and 52.4% (33/63), respectively. These positive granules were brown and most common in cytoplasm or cell membrane of hepatocytes. There was prominently positive correlation between transforming growth factor-alpha and epidermal growth factor receptor (P<0.05, gamma=0.32). In five normal liver tissues, transforming growth factor-alpha and epidermal growth factor receptor were not detectable in hepatocytes and bile ducts. CONCLUSIONS: Hepatic cirrhosis might be under the autocrine regulation of transforming growth factor-alpha and its receptor, epidermal growth factor receptor. Increasing expression of transforming growth factor-alpha and epidermal growth factor receptor might be one of the important events in hepatic cirrhosis pathogenesis. Furthermore, transforming growth factor-alpha might play a role in morphogenesis and regeneration of intrahepatic bile ducts.     

Correlation of human epidermal growth factor receptor protein expression and colorectal cancer

AIM: To investigate the correlation between human epidermal growth factor receptor (HER-2) protein expression and colorectal cancer (CRC) using a case-control study and meta-analysis.METHODS: Tumor tissue specimens from 162 CRC patients were selected for the case group. Fifty cases were randomly selected, and normal CRC tissue at least 10 cm away from the tumor margins of these cases was used to generate the control group. The expression of the HER-2 protein in the 162 CRC tissue samples and the 50 adjacent normal mucosa tissue samples was detected via immunohistochemistry. The experimental data were analyzed using SPSS 18.0 software, and R software version 3.1.0 was utilized for further verification.RESULTS: The expression of HER-2 protein in the 162 CRC tissue samples was significantly higher than in the normal tissue specimens. The data showed that the expression of HER-2 in CRC was related to the Dukes’ stage, the depth of invasion and lymph node metastasis. The HER-2-positive patients had lower 3- and 5-year OS rates than the HER-2-negative patients, but there was no significant difference. However, there was a statistically significant difference in the 3- and 5-year disease-free survival (DFS) rates of HER-2-positive and HER-2-negative patients. The results of the meta-analysis showed that the expression of HER-2 in CRC patients was statistically significantly increased over that of healthy people. The 3-year DFS rate in HER-2-positive patients was markedly lower than that in HER-2-negative patients.CONCLUSION: Down-regulation of HER-2 expression might be a dependable strategy for CRC therapy.     

Characterization of epidermal growth factor receptor gene expression in malignant and normal human cell lines.

To investigate the possibility that the epidermal growth factor (EGF) receptor functions as an oncogene product, we have determined the levels of EGF receptor protein and RNA in a variety of malignant and normal human cells, using a specific polyclonal antibody to the EGF receptor and a cDNA clone (plasmid pE7) that encodes the EGF receptor, respectively. Besides A431 epidermoid carcinoma cells, which are known to make large amounts of EGF receptor, cell lines from two ovarian cancers, two cervical cancers, and one kidney cancer were found to contain substantial amounts of receptor protein (11-22% of A431). Normal human fibroblasts (Detroit 551), a human lymphocyte line (IM-9), and a leukemic lymphocyte line (CEM) contained low or undetectable levels of EGF receptor. RNA blot analysis showed that among the human cell lines examined the levels of a 10- and a 5.6-kilobase species of pE7-specific RNA generally correlated with the amount of the EGF receptor protein. Genomic DNA blot analysis revealed that except for A431 none of these cell lines expressing high levels of EGF receptor protein possessed amplified receptor gene sequences. A431 cells are known to secrete a truncated form of the EGF receptor. An abundant 2.9-kilobase RNA is found only in A431 cells; it could encode the truncated form of the EGF receptor.     

In vivo transfer and expression of a human epidermal growth factor gene accelerates wound repair.

This report details the transfer of a human epidermal growth factor (hEGF) expression plasmid to porcine partial-thickness wound keratinocytes by particle-mediated DNA transfer (Accell). After gene transfer an external sealed fluid-filled wound chamber was used to protect the wound, provide containment of the exogenous DNA and expressed peptide, and permit sampling of the wound fluid. Analysis of wound fluid for hEGF and total protein, an indicator of reformation of the epithelial barrier, showed that wounds bombarded with the hEGF plasmid exhibited a 190-fold increase in EGF concentration and healed 20% (2.1 days) earlier than the controls. EGF concentrations in wound fluid persisted over the entire 10-day monitored period, decreasing from 200 pg/ml to 25 pg/ml over the first 5 days. Polymerase chain reaction results showed that plasmid DNA was present in the wound for at least 30 days. These findings demonstrate the possible utility of in vivo gene transfer to enhance epidermal repair.     

Chemical synthesis of a gene for human epidermal growth factor urogastrone and its expression in yeast.

We have chemically synthesized and expressed in yeast a gene coding for human epidermal growth factor (urogastrone), a 53-amino-acid polypeptide that has been shown to promote epithelial cell proliferation and to inhibit gastric acid secretion. The synthetic gene, consisting of 170 base pairs, was designed with yeast-preferred codons and assembled by enzymatic ligation of synthetic fragments produced by phosphoramidite chemistry. The DNA synthesis protocol used allows for facile synthesis of oligonucleotides larger than 50 bases. Yeast cells were transformed with plasmids containing the synthetic gene under control of a yeast glyceraldehyde-3-phosphate dehydrogenase gene promoter and were shown to synthesize a biologically active human epidermal growth factor.     

Phosphorylation of human growth hormone by the epidermal growth factor-stimulated tyrosine kinase.

In the present study, we have demonstrated that human growth hormone (hGH) can be phosphorylated by the epidermal growth factor (EGF)-stimulated tyrosine kinase of A431 cell membranes. Phosphotyrosine was the predominant phosphoamino acid released from phosphorylated hGH on partial acid hydrolysis. All five tyrosine-containing tryptic peptides of hGH are also phosphorylated by the EGF-stimulated tyrosine kinase. The highest phosphate incorporation was found for peptide T4 (residues 20-38), which is distinguished by a high frequency of acidic amino acids. The phosphorylated peptides have been characterized by HPLC and two-dimensional mapping on paper. Comparison with the labeled peptides obtained on tryptic digestion of phosphorylated hGH suggests that tyrosine phosphorylation is restricted to two tryptic peptides, T4 (tyrosine-28 or -35) and T6 (tyrosine-42). It is suggested that the absence of early insulin-like activity in the naturally occurring Mr 20,000 variant of hGH, which has an internal deletion spanning residues 32-46, may be a consequence of the loss of the tyrosine phosphorylation sites at residues 35 and 42.     

Structural analysis of p185c-neu and epidermal growth factor receptor tyrosine kinases: oligomerization of kinase domains.

The epidermal growth factor receptor (EGFR) and p185c-neu proteins associate as dimers to create an efficient signaling assembly. Overexpression of these receptors together enhances their intrinsic kinase activity and concomitantly results in oncogenic cellular transformation. The ectodomain is able to stabilize the dimer, whereas the kinase domain mediates biological activity. Here we analyze potential interactions of the cytoplasmic kinase domains of the EGFR and p185c-neu tyrosine kinases by homology molecular modeling. This analysis indicates that kinase domains can associate as dimers and, based on intermolecular interaction calculations, that heterodimer formation is favored over homodimers. The study also predicts that the self-autophosphorylation sites located within the kinase domains are not likely to interfere with tyrosine kinase activity, but may regulate the selection of substrates, thereby modulating signal transduction. In addition, the models suggest that the kinase domains of EGFR and p185c-neu can undergo higher order aggregation such as the formation of tetramers. Formation of tetrameric complexes may explain some of the experimentally observed features of their ligand affinity and hetero-receptor internalization.     

胃癌组织人类表皮生长因子受体2基因扩增与蛋白表达的一致性

目的 检测胃癌组织中人类表皮生长因子受体2(HER2)基因扩增与蛋白表达结果的一致性.方法 收集2010年至2012年120例胃癌患者的胃癌组织标本,其中100份为手术标本,20份为胃镜活组织检查标本.应用免疫组织化学(IHC)方法检测120份标本的HER2蛋白表达情况.根据IHC检测结果,选取IHC切片中HER2阳性部位制作成组织芯片进行荧光原位杂交(FISH),检测HER2的基因扩增情况.对IHC检测灶性(+++)(≤10％的肿瘤细胞强染色)的标本进行不同着色强度部位对比FISH检测.统计学处理采用Kappa检验.结果 120份胃癌组织中,IHC检测显示77份为不同强度阳性染色,其中16份为(+++),6份为灶性(+++),37份为(++),18份为(+),IHC检测HER2蛋白表达阳性率为18.3％(22/120).FISH检测显示41份阳性,HER2基因扩增率为34.2％,其中21份IHC检测为(++),15份为(+++),5份为灶性(+++).IHC与FISH联合检测HER2阳性率为35.0％(42/120).IHC与FISH检测的符合率为91.6％(76/83).Kappa系数为0.960(P＜0.01).对5份FISH阳性且IHC检测为灶性(+++)标本的不同着色强度部位进行对比FISH检测,发现全部扩增.结论 组织芯片技术与IHC技术在胃癌组织中检出HER2具有一致性,并可提高HER2的检出率.     

Nuclear localization of epidermal growth factor and epidermal growth factor receptors in human thyroid tissues.

Epidermal growth factor (EGF) has widespread growth effects, and in some tissues proliferation is associated with the nuclear localization of EGF and epidermal growth factor receptor (EGFR). In the thyroid, EGF promotes growth but differs from thyrotropin (TSH) in inhibiting rather than stimulating functional parameters. We have therefore studied the occurrence and cellular distribution of EGF and EGFR in normal thyroid, in Graves' disease, where growth is mediated through the thyrotropin receptor (TSHR), and in a variety of human thyroid tumors. In the normal gland the staining was variable, but largely cytoplasmic, for both EGF and EGFR. In Graves' disease there was strong cytoplasmic staining for both EGF and EGFR, with frequent positive nuclei. Nuclear positivity for EGF and particularly for EGFR was also a feature of both follicular adenomas and follicular carcinomas. Interestingly, nuclear staining was almost absent in papillary carcinomas. These findings document for the first time the presence of nuclear EGF and EGFR in thyroid. Their predominant occurrence in tissues with increased growth (Graves' disease, follicular adenoma, and carcinoma) may indicate that nuclear EGF and EGFR play a role in growth regulation in these conditions. The absence of nuclear EGF and EGFR in papillary carcinomas would suggest that the role played by EGF in growth control differs between papillary carcinoma and follicular adenomas/carcinomas of the thyroid.