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Biomedical subjects

J E Fischer

Publications and source records attributed to J E Fischer.

At least 145 records · Page 8Linked to original sources
PubMed1995

Effect of hepatocytes from normal or endotoxin treated animals on the production of tumour necrosis factor, interleukin-6, and prostaglandin-E2 by macrophages incubated in vitro with various fatty acids.

OBJECTIVE: To find out if the in vitro production of tumour necrosis factor (TNF), interleukin 6 (IL-6) and prostaglandin E2 (PGE2) by macrophages (P3888D1) were influenced by the addition to the culture media of hepatocytes from normal guinea pigs and from guinea pigs after infusion with endotoxin with and without fatty acids. DESIGN: Laboratory experiment. SETTING: University hospital, United States MATERIAL: 12 male Hartley guinea pigs. MAIN OUTCOME MEASURES: Concentrations of TNF-alpha, IL-6, and PGE2. RESULTS: Hepatocytes inhibited the production of TNF by macrophages. Those from animals pretreated with endotoxin caused a large increase in the production of PGE2 and affected the production of IL-6 only in the presence of fatty acids. Fatty acids differed from each other in their effects on production of TNF-alpha and PGE2, but had no effect on the production of IL-6. CONCLUSION: These results support the concept that hepatocytes and macrophages may interact in vivo, and this interaction may be influenced both by fatty acids that arise from lipid metabolism and from endotoxin present during sepsis and translocation.

Animals↗
PubMed1995

Burn injury stimulates multiple proteolytic pathways in skeletal muscle, including the ubiquitin-energy-dependent pathway.

BACKGROUND: Burn injury is associated with increased muscle protein breakdown. However, the role of different intracellular proteolytic pathways in burn-induced muscle proteolysis is not known. STUDY DESIGN: A 30 percent total body surface area burn injury was inflicted on rats. Total and myofibrillar proteolysis was determined in incubated extensor digitorum longus muscles as release of tyrosine and 3-methylhistidine, respectively. Lysosomal proteolysis was assessed by using the lysosomotropic agents leupeptin and methylamine. Calcium-dependent proteolysis was determined by incubating muscles in the absence or presence of calcium or by blocking the calcium-dependent proteases calpain I and II. Energy-dependent proteolysis was determined in muscles depleted of adenosine triphosphate (ATP) by 2-deoxyglucose and 2,4-dinitrophenol. Muscle ubiquitin messenger RNA (mRNA) was determined by Northern blot analysis to assess ATP-ubiquitin-dependent proteolysis. RESULTS: Calcium-dependent total protein breakdown was stimulated in muscles from burned rats. However, the sensitivity to calcium in vitro was not increased after burn. The lysosomal and energy-dependent components of total protein breakdown were doubled in muscles from burned rats and the energy-dependent myofibrillar protein breakdown was increased almost seven-fold. Ubiquitin mRNA was increased in muscles from burned rats. CONCLUSIONS: Burn injury stimulates multiple proteolytic pathways in skeletal muscle. The ubiquitin-energy-dependent pathway may be particularly important for the breakdown of myofibrillar proteins.

Animals↗
PubMed1995

Influence of burn injury on protein metabolism in different types of skeletal muscle and the role of glucocorticoids.

BACKGROUND: Negative nitrogen balance after burn injury mainly indicates muscle catabolism, but the exact influence of burn on protein synthesis and breakdown in different types of skeletal muscle and the role of glucocorticoids in this metabolic response are unknown. STUDY DESIGN: A 30 percent body surface area flame burn was inflicted on rats. Protein turnover rates were measured in vitro in the white fast-twitch extensor digitorum longus (EDL) muscle and the red slow-twitch soleus muscle. To test the role of glucocorticoids, groups of rats were treated with the glucocorticoid receptor antagonist RU 38486 or vehicle. RESULTS: Burns resulted in reduced protein synthesis and stimulated protein breakdown, in particular myofibrillar protein breakdown, and the changes were substantially more pronounced in the EDL than in the soleus muscle. A burn-induced increase in muscle proteolysis was abolished by treatment with RU 38486, whereas the reduced protein synthesis was not affected by the glucocorticoid receptor antagonist. CONCLUSIONS: The results suggest that the white fast-twitch muscle is more sensitive to the effects of burn injury than the red slow-twitch muscle. Burn-induced muscle proteolysis may be mediated, at least in part, by glucocorticoids, whereas protein synthesis is probably regulated by other mechanisms.

Animals↗
PubMed1995

Sepsis stimulates polyamine biosynthesis in the liver and increases tissue levels of ornithine decarboxylase mRNA.

The influence of sepsis on polyamine metabolism in the liver was studied in rats. Sepsis was induced by cecal ligation and puncture; control rats were sham-operated. Sepsis resulted in increased concentrations in liver tissue of putrescine and spermidine and stimulated activity of the enzymes ornithine decarboxylase (ODC) and s-adenosylmethionine decarboxylase. A similar metabolic response was seen following the subcutaneous injection of 1 mg/kg of endotoxin or following the e intraperitoneal injection of 100 micrograms/kg of human recombinant tumor necrosis factor (TNF)-alpha or interleukin-1 alpha (IL-1 alpha). ODC mRNA levels determined by Northern blots were increased in liver tissue of septic rats, suggesting that the increase in ODC activity may be regulated at the transcriptional level although increased stability of the messenger could give rise to similar results. Treatment of rats with either TNF antiserum, recombinant IL-1 receptor antagonist, or the glucocorticoid receptor antagonist RU 38486, did not prevent the sepsis-induced increase in hepatic ODC activity. The data suggest that sepsis stimulates the biosynthesis of polyamines in liver tissue and that this response to sepsis may not primarily be mediated by TNF, IL-1, or glucocorticoids. The biological role of increased liver polyamines during sepsis, in particular their relationship with the synthesis of acute phase proteins, remains to be determined.

Animals↗
PubMed1994

Characterization of the synthesis and expression of the GTA-kinase from transformed and normal rodent cells.

The murine transformed cell line YC-8 and beta-adrenergic receptor agonist (isoproternol) treated rat and mouse parotid gland acinar cells ectopically express cell surface beta 1-4 galactosyltransferase during active proliferation. This activity is dependent upon the expression of the GTA-kinase (p58) in these cells. Using total RNA, cDNA clones for the protein coding region of the kinase were isolated by reverse transcriptase-PCR cloning. DNA sequence analysis failed to show sequence differences with the normal homolog from mouse cells although Southern blot analysis of YC-8, and a second cell line KI81, indicated changes in the restriction enzyme digestion profile relative to murine cell lines which do not express cell surface galactosyltransferase. The rat cDNA clone from isoproterenol-treated salivary glands showed a high degree of protein and nucleic acid sequence homology to the GTA-kinase from both murine and human sources. Northern blot analysis of YC-8 and a control cell line LSTRA revealed the synthesis of a major 3.0 kb mRNA from both cell lines plus the unique expression of a 4.5 kb mRNA in the YC-8 cells. Reverse transcriptase-PCR of LSTRA and YC-8 confirmed the increased steady state levels of the GTA-kinase mRNA in YC-8. In the mouse, induction of cell proliferation by isoproterenol resulted in a 50-fold increase in steady state mRNA levels for the kinase over the low level of expression in quiescent cells. Expression of the rat 3' untranslated region in rat parotid cells in vitro led to an increased rate of DNA synthesis, cell number an ectopic expression of cell surface galactosyltransferase in the sense orientation. Antisense expression or vector alone did not alter growth characteristics of acinar cells. A polyclonal antibody monospecific to a murine amino terminal peptide sequence revealed a uniform distribution of GTA-kinase over the cytoplasm of acinar and duct cells of control mouse parotid glands. However, upon growth stimulation, kinase was detected primarily in a perinuclear and nuclear immunostaining pattern. Western blot analysis confirmed a translocation from a cytoplasmic localization in both LSTRA and quiescent salivary cells to a membrane-associated localization in YC-8 and proliferating salivary cells.

Amino Acid Sequence↗
PubMed1994

[D-TRP32]neuropeptide Y: a competitive antagonist of NPY in rat hypothalamus.

Neuropeptide Y (NPY) is a potent orexigenic peptide. Structure-activity studies have revealed that nearly the entire sequence of NPY is required to elicit feeding responses. Therefore, in order to develop antagonistic peptides for NPY-induced feeding, we synthesized full-length analogs of NPY, substituting D-Trp in the C-terminal receptor binding region, and screened their activity in rat hypothalamus. Although [D-Trp36]NPY and [D-Trp34]NPY inhibited isoproterenol-stimulated hypothalamic membrane adenylate cyclase activity, [D-Trp32]NPY exhibited no intrinsic activity. Furthermore, [D-Trp32]NPY inhibited [125I]NPY binding to rat hypothalamic membranes with a potency comparable to that of NPY. The presence of 30 and 300 nM concentrations of [D-Trp32]NPY shifted the inhibitory dose-response curve of NPY on isoproterenol-stimulated hypothalamic membrane adenylate cyclase activity parallel to the right with comparable KB values. Moreover, in vivo experiments in rats revealed that [D-Trp32]NPY (10 micrograms) significantly attenuated the 1-h feeding response induced by NPY (1 microgram). Several other substitutions at position 32 including 2-D-Nal resulted in agonist activity, suggesting that there are strict structural requirements to induce the antagonistic property in NPY. These findings confirm that [D-Trp32]NPY is a competitive antagonist of NPY in both in vitro and in vivo systems. Analogs based on [D-Trp32]NPY may have potential clinical application, since NPY has been implicated in the pathophysiology of a number of feeding disorders including obesity, anorexia, and bulimia.

Adenylyl Cyclase Inhibitors↗
PubMed1994

Endotoxin stimulates interleukin-6 production in intestinal epithelial cells. A synergistic effect with prostaglandin E2.

OBJECTIVE: To test the hypothesis that endotoxin stimulates the release of interleukin-6 (IL-6) from intestinal epithelial cells and that this effect of endotoxin is regulated by prostaglandin E2 (PGE2). DESIGN: A rat intestinal crypt cell line, IEC-6, was cultured in the presence of lipopolysaccharide (LPS), 0.1 to 1.0 microgram/mL, and/or PGE2, 1 mumol/L. In other experiments, indomethacin, 20 mumol/L, was added to LPS-treated cells to block the effects of prostaglandins. Control wells contained medium alone. Levels of IL-6 were determined by the B9 murine hybridoma bioassay. Polymerase chain reaction was performed on RNA from control and LPS-treated cells to examine IL-6 message. RESULTS: Lipopolysaccharide and PGE2 induced IL-6 release from IEC-6 cells in a dose- and time-dependent fashion, and the substances interacted synergistically. Addition of indomethacin blunted the effect of endotoxin on IL-6 production, consistent with a stimulatory role of PGE2. Polymerase chain reaction demonstrated increased IL-6 messenger RNA in endotoxin-treated cells. CONCLUSIONS: Endotoxin and PGE2 stimulate IL-6 production in IEC-6 cells and interact synergistically. The endotoxin-stimulated IL-6 release may be regulated at the transcriptional level.

Animals↗
PubMed1994

Possible role of neuropeptide Y in experimental cancer anorexia.

The efficacy of NPY to elicit feeding in TB rats was reduced prior to the onset of overt anorexia, with the feeding response decreasing further as anorexia developed. Hypothalamic concentration of NPY was reduced in TB rats, with the magnitude of the decrease paralleling the degree of anorexia. Binding affinity of NPY to hypothalamic membranes taken from TB rats suggested decreased binding affinity with no change in receptor number. Infusing ammonium salts at a concentration and rate necessary to increase blood ammonia levels to the degree observed in TB rats, produced anorexia and decreased NPY feeding. These results suggest that NPY feeding systems are abnormal in TB rats and that hyperammonemia may be of primary importance in this dysfunction.

Ammonia↗
PubMed1994

Sepsis in rats stimulates cellular proliferation in the mucosa of the small intestine.

BACKGROUND/AIMS: Increased protein synthesis in intestinal mucosa during sepsis may reflect increased cell turnover. The influence of sepsis and endotoxemia on cellular proliferation in the mucosa of the small intestine was studied. METHODS: Sepsis was induced in rats by cecal ligation and puncture. Control rats were sham-operated. Other rats were treated with endotoxin (total dose, 2 mg/kg), human recombinant tumor necrosis factor alpha, or human recombinant interleukin 1 alpha at a dose of 100 micrograms/kg each. Villus height and crypt depth in the jejunum were measured as the number of cells along the side of the villus and crypt, respectively. Cellular proliferation was assessed by measuring the rate of [3H]thymidine incorporation into DNA of the jejunal mucosa and performing autoradiographic studies after intravenous administration of [3H]thymidine. RESULTS: Sepsis resulted in reduced villus height, increased crypt depth, and increased incorporation of [3H]thymidine into DNA in the jejunal mucosa. Autoradiography after administration of [3H]thymidine showed labeled cells almost exclusively in the crypts; the number of labeled cells per crpt was higher in septic than in control rats. Administration of endotoxin or recombinant interleukin 1 alpha, but not recombinant tumor necrosis factor alpha, stimulated the incorporation of [3H]-thymidine into DNA in the jejunal mucosa. CONCLUSION: Sepsis and endotoxemia stimulate cellular proliferation in the mucosa of the small intestine. This response to sepsis and endotoxemia may be partly mediated by interleukin 1.

Animals↗
PubMed1994

Reduced muscle protein breakdown in septic rats following treatment with interleukin-1 receptor antagonist.

1. The role of interleukin-1 (IL-1) in sepsis-induced muscle proteolysis was assessed by treating septic rats with recombinant IL-1 receptor antagonist (rIL-1ra). 2. In initial experiments, we tested the effectiveness of IL-1ra in preventing muscle proteolysis induced by administration of IL-1. 3. When normal rats were treated with rIL-1 alpha (three intraperitoneal doses of 100 micrograms/kg body weight each over 16 hr), total and myofibrillar muscle protein breakdown rates, measured as release of tyrosine and 3-methylhistidine, respectively, by incubated extensor digitorum longus muscles, were significantly increased. 4. This metabolic response to IL-1 alpha was completely abolished by rIL-1ra, administered as three intraperitoneal doses of 3 mg/kg body weight each over 16 hr. 5. In subsequent experiments, sepsis was induced in rats by cecal ligation and puncture (CLP); non-septic rats were sham-operated. 6. Treatment of septic rats over 16 hr with a total dose of 25 mg/kg body weight of rIL-1ra reduced, but did not normalize, the increased muscle protein breakdown rates seen during sepsis. 7. When the dose of rIL-1ra was more than doubled and given as a constant infusion at a rate of 4.2 mg/kg body weight/hr for 16 hr, the increased rate of muscle proteolysis in septic rats was normalized. 8. The present study offers the first direct evidence that IL-1 is involved in the regulation of muscle proteolysis during sepsis.

Animals↗
PubMed1994

Hypothalamic concentration and release of neuropeptide Y into microdialysates is reduced in anorectic tumor-bearing rats.

Hypothalamic concentration of neuropeptide Y was decreased significantly in anorectic tumor-bearing rats, while NPY level was increased significantly in matched carcass weight control rats as compared with freely-feeding controls. In vivo microdialysis of the perifornical hypothalamic area of tumor-bearing rats prior to the development of anorexia revealed no alteration in NPY in dialysates. Following the development of anorexia, however, tumor-bearing rats exhibited significant reduction in NPY concentration in dialysates as compared with either matched carcass weight or freely-feeding control group. These results suggest that hypothalamic NPY concentration and release are decreased selectively in anorectic tumor-bearing rats. Since NPY also elicits less feeding in tumor-bearing rats, dysfunction of hypothalamic NPY feeding mechanisms may be of primary importance in cancer anorexia.

Animals↗
PubMed1994

Sepsis increases production of total secreted proteins, vasoactive intestinal peptide, and peptide YY in isolated rat enterocytes.

The effect of sepsis on the synthesis of endogenous and secretory proteins, including vasoactive intestinal peptide (VIP) and peptide YY (PYY), was determined in enterocytes from jejunum of rats. Sepsis was induced by cecal ligation and puncture (CLP). Control rats were sham-operated. Total endogenous and secreted protein synthesis was assessed in incubated jejunal enterocytes by measuring incorporation of 3H-phenylalanine into protein. Release of VIP and PYY into the medium of incubated enterocytes and cellular levels of the gut peptides were measured by radioimmunoassay. Sixteen hours after CLP, synthesis rates of both endogenous and secreted proteins were increased, and this effect of sepsis was most pronounced in cells from the lower parts of the villi and crypts. Enterocytes from septic rats released more VIP and PYY into the incubation medium, and approximately half of the peptides they released were newly synthesized VIP and PYY. Intracellular levels of VIP and PYY were increased as early as 4 hours after induction of sepsis. Our results suggest that sepsis stimulates the synthesis of endogenous and secretory proteins, including certain gut peptides, in small intestine mucosa. This is consistent with previous observations of increased circulating levels of VIP, PYY and other gastrointestinal hormones during sepsis. The biological significance of increased synthesis of gut peptides and other intestinal proteins during sepsis remains to be determined.

Analysis of Variance↗
PubMed1994

Effect of sepsis on mucosal protein synthesis in different parts of the gastrointestinal tract in rats.

1. In a previous study we found that the protein synthesis rate was increased by 50-60% in the mucosa of the jejunum and ileum during sepsis in rats. It is not known if sepsis affects protein turnover in other parts of the gastrointestinal tract as well. 2. In the present study, the influence of sepsis on mucosal protein synthesis in different parts of the gastrointestinal tract, from the stomach to the rectum, was determined in rats. 3. Sepsis was induced by caecal ligation and puncture; control rats underwent sham-operation. Protein synthesis rate was measured in vivo after administration of a flooding dose of [14C]leucine. 4. Basal mucosal protein synthesis rates were lower in the colon than in the rest of the gastrointestinal tract. Sixteen hours after caecal ligation and puncture, the protein synthesis rates were increased by 40-85% in the mucosa of the small and large intestine and the rectum, whereas in the gastric mucosa, the protein synthesis rate was reduced by approximately 40%. 5. The results suggest that mucosal protein synthesis rates differ in the various regions of the gastrointestinal tract, and that the metabolic response to sepsis is different in the stomach than in the rest of the gastrointestinal tract. The finding of a reduced protein synthesis rate in the gastric mucosa may partly explain the tendency to gastric stress ulcers and bleeding seen clinically in sepsis.

Animals↗
PubMed1994

Insulin reverses ammonia-induced anorexia and experimental cancer anorexia.

Previous experiments suggest that experimental cancer-induced anorexia is associated with hyperammonemia and that daily injections of insulin may attenuate the anorexia for several days. In the present study, we determined whether similar daily insulin treatments would correct anorexia induced by the infusion of ammonium salts and compared this feeding response with that of insulin-treated tumor-bearing (TB) rats. Daily treatment of control and anorectic TB rats with systemically administered insulin for six days increased feeding in all control rats and 40% of the TB rats. All insulin-treated groups exhibited equal degrees of hypoglycemia irrespective of anorexia. Basal concentrations of lactate and glucagon were elevated in saline-treated TB rats. Plasma lactate levels were normalized by insulin treatment, whereas glucagon was normalized only in the TB rats that fed to insulin and increased further in TB rats that did not feed to insulin. Elevated hypothalamic tyrosine was reduced in insulin-treated TB rats that ate, and 5-hydroxy-indoleacetic acid was increased further when the rats did not eat. Insulin also blocked anorexia resulting from the intravenous infusion of ammonium salts. Hypothalamic concentrations of tyrosine and tryptophan were increased by the ammonia infusion and reduced significantly in insulin-treated infused rats. These results indicate that insulin treatment can reverse experimental cancer-induced anorexia and hyperammonemia-induced anorexia. Neurochemical changes associated with these treatments are also similar, but not identical.

Ammonia↗
PubMed1994

Sepsis stimulates nonlysosomal, energy-dependent proteolysis and increases ubiquitin mRNA levels in rat skeletal muscle.

We tested the role of different intracellular proteolytic pathways in sepsis-induced muscle proteolysis. Sepsis was induced in rats by cecal ligation and puncture; controls were sham operated. Total and myofibrillar proteolysis was determined in incubated extensor digitorum longus muscles as release of tyrosine and 3-methylhistidine, respectively. Lysosomal proteolysis was assessed by using the lysosomotropic agents NH4Cl, chloroquine, leupeptin, and methylamine. Ca(2+)-dependent proteolysis was determined in the absence or presence of Ca2+ or by blocking the Ca(2+)-dependent proteases calpain I and II. Energy-dependent proteolysis was determined in muscles depleted of ATP by 2-deoxyglucose and 2.4-dinitrophenol. Muscle ubiquitin mRNA and the concentrations of free and conjugated ubiquitin were determined by Northern and Western blots, respectively, to assess the role of the ATP-ubiquitin-dependent proteolytic pathway. Total and myofibrillar protein breakdown was increased during sepsis by 50 and 440%, respectively. Lysosomal and Ca(2+)-dependent proteolysis was similar in control and septic rats. In contrast, energy-dependent total and myofibrillar protein breakdown was increased by 172% and more than fourfold, respectively, in septic muscle. Ubiquitin mRNA was increased severalfold in septic muscle. The results suggest that the increase in muscle proteolysis during sepsis is due to an increase in nonlysosomal energy-dependent protein breakdown, which may involve the ubiquitin system.

Animals↗