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Biomedical subjects

G Carpenter

Publications and source records attributed to G Carpenter.

At least 163 records · Page 9Linked to original sources
PubMed1983

Characteristics of antibodies to the epidermal growth factor receptor-kinase.

Polyclonal antibodies to different antigenic forms of the epidermal growth factor (EGF) receptor-kinase from human A-431 cells have been produced, and their properties have been characterized and compared. Biochemically active receptor-kinase purified by affinity chromatography was employed as one type of antigen. Denatured receptor-kinase prepared by sodium dodecyl sulfate-gel electrophoresis of the affinity-purified receptor was used as the second type of antigen. Animals immunized with either type of antigen produced antibody capable of immunoprecipitating the receptor-kinase molecule. Antibodies produced in response to the biochemically active antigenic form of the receptor-kinase are capable of blocking 125I-EGF binding to the receptor and inhibited EGF-stimulated biological responses. These antisera are not species specific in their ability to inhibit growth-factor binding to the EGF receptor of various mammalian cells. However, these rabbit antisera were unable to inhibit 125I-EGF binding to rabbit cells. Although antisera produced in response to the denatured receptor-kinase molecule are not able to block 125I-EGF binding or EGF-stimulated biological responses, they are particularly efficient for the immunoprecipitation of solubilized 125I-EGF:receptor complexes. None of the antisera contain antibodies capable of interfering with basal receptor-kinase phosphorylation activity. Although each of the antisera immunoprecipitated this kinase activity, none of the antisera contained antibody which served as a phosphorylation substrate for the EGF receptor-kinase in contrast to the immunoglobulins present antisera to the src gene product of the Rous sarcoma virus.

Animals↗
PubMed1983

The biochemistry and physiology of the receptor-kinase for epidermal growth factor.

Epidermal growth factor (EGF) is a polypeptide mitogen that stimulates the proliferation of epidermal and epithelial cells in intact animals and a variety of cell types in cell culture. The biological effects of EGF are mediated by a 170000 dalton receptor located in the plasma membrane. Studies with cultured cells and more recently with subcellular membrane preparations have revealed significant information about the mechanism of action of EGF. In particular, events which occur immediately upon the formation of hormone-receptor complexes have been described. The biochemical signaling that must occur between the membrane-localized EGF-receptor complexes and the cell nucleus to initiate replicative DNA synthesis appears to involve an initial phosphorylation step. The EGF receptor molecule contains a protein kinase active site and this kinase activity is enhanced by EGF binding. Since the kinase phosphorylates tyrosyl residues, this normal growth-regulating system bears a similarity to the mechanism employed by certain oncogenic viruses to transform normal cellular growth behavior.

Animals↗
PubMed1983

Antibodies to the epidermal growth factor receptor block the biological activities of sarcoma growth factor.

The role of the epidermal growth factor (EGF) receptor system in mediating the biological activities of sarcoma growth factor (SGF) has been assessed by using specific anti-EGF receptor antibodies. There are two classes of anti-EGF receptor antibodies, those that block binding of 125I-labeled EGF (125I-EGF) and those that do not block binding but do interact with a portion of the EGF receptor on the surface of intact cells. Antisera of both types have been assayed for their capacity to affect the biological activities of SGF. The antisera that block 125I-EGF binding to its receptor block the induction of DNA synthesis in human fibroblasts by either EGF or SGF but not by other polypeptide mitogens. Titration of the anti-EGF receptor antiserum indicates the presence of one population of antibody that blocks the site of both EGF and SGF action. Antisera to the EGF receptor that block 125I-EGF binding also inhibited the SGF-dependent anchorage-independent growth of normal cells in soft agar. The antisera to the EGF receptor that does not block 125I-EGF binding or EGF activity did not inhibit any of the biological activities of SGF. The results suggest that occupation of the EGF receptor is required for both the mitogenic and colony-forming activity of SGF.

Animals↗
PubMed1983

Enteral and parenteral feeding influences mortality after hemoglobin-E. coli peritonitis in normal rats.

Enteral feeding with 25% dextrose-4.25% Freamine II (TPN) improves the survival of malnourished animals to normal levels after hemoglobin-E. coli adjuvant peritonitis, whereas intravenous feeding does not. To determine whether intravenous feeding maintained a high survival rate in previously well-nourished animals, 81 rats received TPN via gastrostomy or intravenous infusion for 12 days. They were then fasted for 24 hours and given a septic challenge. Gastrostomy-fed animals survived the challenge significantly better than intravenously fed animals. Enteral feeding appears to be important in producing a high survival rate after hemoglobin-E. coli adjuvant peritonitis.

Animals↗
PubMed1983

Developments in the mechanism of growth factor action: activation of protein kinase by epidermal growth factor.

The interaction between epidermal growth factor (EGF) and its target cells has been used as a model for studying the regulation of cell proliferation. Many of the details of binding and subsequent internalization and degradation of this growth factor have been elucidated by following the fate of [125I]EGF in the presence of responsive cells. To investigate the membrane-localized biochemical consequences of EGF-receptor complex formation, a subcellular membrane system has been developed. In this system, EGF enhances phosphorylation of its receptor as well as other endogenous proteins. This EGF-stimulable protein kinase activity is not separated from the EGF receptor activity either by detergent solubilization or by affinity purification of the solubilized membranes. The data suggest that the EGF-binding activity and EGF-sensitive protein kinase activity reside in a single membrane protein.

Animals↗
PubMed1982

Effects of enteral and parenteral feeding of malnourished rats on body composition.

Many studies have reported similar weight gains and nitrogen balance in groups of well-nourished animals fed either orally or intravenously, but none have investigated the effects on malnourished animals. After protein depleting for 14 days, rats were refed with a D25-4.25% amino acid solution orally ad lib, by gastrostomy, or intravenously. IV animals gained more weight with greater body fat formation than either enteral group but had lower intestinal mass and nitrogen. Route of administration influenced substrate utilization in malnourished animals fed identical diets. We conclude that weight gain cannot be used as a comparative indicator of nitrogen retention between malnourished animals fed enterally and those fed intravenously.

Animals↗
PubMed1981

Epidermal growth factor-receptor-protein kinase interactions.

Membranes prepared from A-431 human epidermoid carcinoma cells retained the ability to bind 125I-labeled epidermal growth factor (EGF) in a specific manner. In the presence of [gamma-32]ATP and Mn2+ or Mg2+, this membrane preparation was capable of phosphorylating specific endogenous membrane proteins, as well as exogenously added histone. The binding of EGF to membranes in vitro resulted in a several-fold stimulation of the phosphorylation reaction. The phosphorylation reaction was not dependent on cyclic AMP or cyclic GMP. These findings suggest that one of the biochemical consequences of the binding of EGF to membranes is a rapid activation of a cyclic AMP-independent phosphorylating system. The activation of the membrane-associated protein kinase by EGF appears to be a reversible phenomenon. The membrane preparation could be solubilized by a number of non-ionic detergents with the retention of both 125I-labeled EGF binding activity and EGF-enhanced phosphorylation of specific membrane proteins. The solubilized membrane preparation was purified by affinity chromatography. The purified preparation retained both EGF-binding activity and EGF-enhanced phosphorylation activity. Analysis of the affinity-purified preparation by SDS gel electrophoresis indicated the presence of one major protein band of molecular weight 150,000 and several trace bands. The evidence suggests that the major 150,000 protein band is the receptor for EGF and is a substrate of the phosphorylation reaction. The co-purification of EGF-binding activity and EGF-stimulated phosphorylation activity suggests an inherent close relationship. The EGF-stimulated phosphorylation reaction appears to be specific for tyrosine residues and in this regard resembles the src protein kinase.

Carcinoma, Squamous Cell↗
PubMed1981

Malnutrition and immunocompetence: increased mortality following an infectious challenge during hyperalimentation.

Malnutrition is associated with an increased susceptibility to infection by altering host defense mechanisms. A number of investigators have demonstrated restoration of in vitro immunocompetence following nutritional repletion with intravenous hyperalimentation. This study was designed to assess in vivo host defense mechanisms following protein depletion and repletion using a septic challenge. Female Fischer rats (150 gm) were fed a regular diet or a 2% agar protein depletion (DEP) diet for 14 days. The rats were then administered an infectious challenge with intraperitoneal injections of 7 ml/kg of a solution of 10(9) organisms/ml of E. coli with 4 gm% hemoglobin as red cells. Two additional organisms were repleted by regular diet (DEP-Oral) or by an intravenous solution (DEP-TPN) of dextrose-amino acid-lipid (D25 3.75% Aminosyn, 10% Intralipid) for an additional 2-week period, and given the experimental peritonitis challenge. Normal rats challenged with E. coli-hemoglobin adjuvant peritonitis had 66% survival as opposed to 15% survival of protein depleted rats. Protein-depleted (DEP-oral) rats refed with regular diets had a 60% survival which was comparable to normal controls. Rats repleted with intravenous hyperalimentation had a mortality comparable to protein-depleted controls. The data confirm that protein depletion is associated with loss of host defense mechanisms. Although refeeding by regular diet resulted in restoration of host defense, repletion by parenteral nutrition, in this model, did not improve survival.

Adjuvants, Immunologic↗
PubMed1980

Epidermal growth factor is a major growth-promoting agent in human milk.

Human milk stimulates DNA synthesis in cell cultures in which growth has been arrested. The mitogenic activity of milk is neutralized by the addition of antibody to human epidermal growth factor. The results identify epidermal growth factor as a major growth-promoting agent in breast milk.

Animals↗
PubMed1980

Epidermal growth factor-receptor-protein kinase interactions. Co-purification of receptor and epidermal growth factor-enhanced phosphorylation activity.

Membranes may be prepared from A-431 human epidermoid carcinoma cells which have the ability to bind 125I-labeled epidermal growth factor (EGF) in a specific manner and which, in the presence of EGF, catalyze the phosphorylation of a number of endogenous membrane proteins. The activation of the membrane associated protein kinase by EGF appears to be a reversible phenomenon. The membrane preparation may be solubilized by a number of nonionic detergents with the retention of both 125I-labeled EGF-binding activity and EGF-enhanced phosphorylation of specific membrane proteins. The solubilized membrane preparation may be purified by affinity chromatography using EGF covalently linked to Affi-Gel. The purified preparation retains both EGF-binding activity and EGF-enhanced phosphorylation activity. Analysis of the affinity-purified preparation by sodium dodecyl sulfate-gel electrophoresis indicates the presence of one major protein band of molecular weight 150,000 and several trace bands. The evidence suggests that the major 150,000 protein band is the receptor for EGF and is a substrate of the phosphorylation reaction. The co-purification of EGF-binding activity and EGF-stimulated phosphorylation activity suggests an inherent close relationship.

Animals↗
PubMed1980

Production and characterization of antibody blocking epidermal growth factor:receptor interactions.

Membranes were prepared from the human epithelioid carcinoma cell line A-431 which has approx. 2 . 10(6) epidermal growth factor receptors per cell. This membrane preparation which retained a high epidermal growth factor binding specific activity was used as an antigen to produce antisera in rabbits. Double-immunodiffusion experiments demonstrated that the immune serum contained precipitating antibodies to several components of detergent solubilized A-431 membranes. The immunoglobulin G fraction of this immune sera inhibited 125I-labeled epidermal growth factor binding to receptors in: (1) intact human and mouse cells; (2) membrane preparations from A-431 cells and human placenta, and (3) solubilized A-431 membranes. Inhibition of 125I-labeled epidermal growth factor binding was observed with divalent and monovalent fragments of immunoglobulin G prepared from the immunoglobulin G fraction. Also, the immunoglobulin G fraction blocked growth factor binding to membranes at low temperature (5 degrees C). Anti-A-431 antibody blocked the induction of DNA synthesis in quiescent fibroblasts by epidermal growth factor in a manner similar to that of anti-epidermal growth factor antibody. Addition of either anti-A431 or anti-epidermal growth factor antibodies to fibroblasts at times up to 5 h after the addition of epidermal growth factor completely reversed the hormone's mitogenic potential. At later times (after 12 h) addition of either antibody was without effect on the stimulation of DNA synthesis by epidermal growth factor. Anti-A-431 antibody did not block the induction of DNA synthesis in fibroblasts by fibroblast growth factor or serum.

Animals↗
PubMed1980

Protein phosphorylation in human placenta. Stimulation by epidermal growth factor.

Membranes prepared from normal human term placenta possess a protein kinase activity which phosphorylates endogenous substrates in the presence of [gamma-32P]ATP. This kinase activity requires either Mg2+ or Mn2+, is enhanced by glycerol and appears to be cyclic-nucleotide-independent. Addition of epidermal growth factor to the placental membrane preparation increases the level of total phosphorylation by approx. 35%. When analyzed by sodium dodecyl sulfate gel electrophoresis and autoradiography, the placental proteins whose degree of phosphorylation was enhanced by epidermal growth factor had apparent molecular weights of 170000, 150000, and 25000. This report documents the presence of protein kinase activity in human placenta and demonstrates that epidermal growth factor can enhance protein phosphorylation in normal human tissue.

Adenosine Triphosphate↗
PubMed1980

Autoregulation of erythropoiesis by 2,3 diphosphoglycerate during hyperalimentation.

Protein depletion in rats is associated with hemoconcentration, diminution of red cell mass, and suppression of erythropoiesis. In a rat model, repletion with standard diets or total parenteral nutrition was associated with gain in body weight and regeneration of red cell mass. Hematocrit values did not reflect the degree of red cell mass diminution associated with protein depletion. Intravenous repletion of normal and protein-depleted rats, unlike oral refed cohorts, was associated with elevated 2,3 diphosphoglycerate (DPG) values and a 'relative anemia.' It is postulated that elevated red cell 2,3 DPG lowers the hypoxic stimulus for erythropoietin production by enhancing oxygen unloading to the tissues, thus partially compensating for the anemia associated with total parenteral nutrition.

Anemia↗