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J J Spitzer

Publications and source records attributed to J J Spitzer.

At least 37 records · Page 2Linked to original sources
PubMed1995

Nitro containing L-arginine analogs interfere with assays for nitrate and nitrite.

We evaluated the effect of in vivo and in vitro administration of nitro-containing and nitro-deficient L-arginine-derived nitric oxide (NO) synthase inhibitors on the measurement of NO in plasma, urine and HEPES buffered physiologic salt solution (PSS) by ozone chemiluminescence and by the modified Griess reaction. In vivo administration of 1, 5, 25, 40 or 50 mg/kg of NG-nitro-L-or D-arginine methyl ester (LNAME, DNAME), NG-nitro-L-arginine (LNA) or aminoguanidine (AG) to rats and mice increased NO in urine and plasma as determined by chemiluminescence using 2.3% vanadium chloride in 2N HCI at 100 degrees C as the redox reagent. In vivo administration of 1 and 10 mg/kg/day of NG-imino-ethyl-L-ornithine (LNIO) or 3 amino-1,2,4 triazine (AT) reduced plasma and urine NO. Addition of LNAME, DNAME and LNA (100 nM to 1 mM) to the redox solution produced a concentration response curve for NO in the chemiluminescence assay similar to that produced by standard solutions of sodium nitrite and nitrate. LNMMA produced a small NO signal but only at concentrations equal to or exceeding 0.1 mM. LNIO, AT and AG did not give any NO signal even at concentrations exceeding 1 mM. Conversion of plasma or urine nitrate to nitrite with cadmium gave elevated values of plasma nitrite by the Greiss assay when LNAME or LNA was the NO synthase inhibitor. We conclude that in vivo and in vitro use of LNAME and LNA and in vivo use of high doses of aminoguanidine interfere with the assay of NO2- and NO3- with the modified Griess reaction and with chemiluminescence. We suggest that LNAME and LNA not be used in vivo or in vitro when total RNI is measured with these assays.

Animals↗
PubMed1995

Liver sinusoid during chronic alcohol consumption in the rat: an electron microscopic study.

Transmission and scanning electron microscopic studies were performed on the liver sinusoid, with emphasis on sinusoidal endothelial cells, in rats fed a liquid diet containing either alcohol or dextrin (control) for 14 weeks. Animals were also treated with either Gram-negative bacterial lipopolysaccharide (LPS; 100 micrograms/100 g body weight, intravenously) or sterile saline (control). All specimens were prepared after perfusion fixation of the liver. Livers of rats fed dextrin-containing liquid diet displayed the ultrastructural features typical of the sinusoid and its endothelial cells. Livers from alcohol-fed animals, however, were characterized by massive loss of sieve-plate architecture of the sinusoidal endothelium, which was virtually replaced with a meshwork of enlarged openings with diameters frequently exceeding 1 micron. Morphological evidence of Kupffer cell activation could also be seen along with significant fatty infiltration of the hepatocyte. Conversely, LPS administration to dextrin-fed animals induced an apparent decrease in fenestration of the sinusoidal endothelial cell, accompanied by morphological evidence of enhanced endocytotic activity and cytoplasmic swelling. The changes seen 3 hr after LPS administration were markedly advanced at 24 hr. LPS administration to alcohol-fed rats accentuated the alterations observed after alcohol treatment alone. Additionally, the presence of platelets in the sinusoid as well as adhering to the hepatocyte microvilli in the space of Disse, along with the presence of Ito and Kupffer cell activation, greater than that observed in the alcohol-treated rats, is morphological evidence consistent with the disruption of vascular integrity in the liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Tumor necrosis factor-alpha cell-surface receptors of liver parenchymal and nonparenchymal cells during acute and chronic alcohol administration to rats.

Tumor necrosis factor-alpha (TNF-alpha) has been shown to contribute to the alcohol [ethanol (ETOH)]-induced alteration of hepatic function. Therefore we tested the hypothesis that the hepatic action of TNF-alpha could be due, at least in part, to alterations in TNF-alpha cell-surface receptors of hepatic parenchymal (hepatocytes) and nonparenchymal (Kupffer and sinusoidal endothelial) cells. Rats were either acutely treated with ETOH by a primed, continuous 7-hr intravenous infusion of 20% (w/v) ETOH (30 mg/100 g body weight/h) or chronically fed an ETOH-containing liquid diet (5.2% ETOH, w/v, with ETOH as 36% of total calories) for 14 weeks. Control rats in the acute group were infused with sterile saline, whereas control rats in the chronic group were fed liquid diet containing dextrin to replace ETOH in isocaloric amounts. Three hr before killing, the rats were injected intravenously with Gram-negative bacterial lipopolysaccharide [(LPS) 100 micrograms/100 g body weight] or saline. Hepatocytes, Kupffer cells, and sinusoidal endothelial cells were isolated after liver perfusion with collagenase (without pronase), separated by centrifugal elutriation, and used to determine the affinity (Kd) and capacity (Bmax) of binding sites, using recombinant human-[125I]TNF-alpha as the ligand. Two binding sites were detected on Kupffer cells and sinusoidal endothelial cells isolated from control animals: a high-affinity (Kd1, in the range of 150-200 pM), low-capacity (Bmax, in the range of 2-3 fmol/10(6) cells) binding site and a low-affinity (Kd2, in the range of 2-9 nM), high-capacity (Bmax2, in the range of 3-15 fmol/10(6) cells) binding site.(ABSTRACT TRUNCATED AT 250 WORDS)

Alcoholic Intoxication↗
PubMed1995

Chronic alcohol administration stimulates nitric oxide formation in the rat liver with or without pretreatment by lipopolysaccharide.

This study examines the effect of chronic alcohol consumption on nitric oxide release from the liver of rats with or without lipopolysaccharide (LPS) (Escherichia coli) treatment. Reactive nitrogen intermediates (RNIs) in plasma were monitored with an NOx Analyzer, and nitric oxide (NO) production was measured as nitrite or nitrite + nitrate accumulation in perfusates of the perfused liver, and in supernatants of the freshly isolated hepatic cells after incubation for 3 hr in Hank's balanced salt solution buffer containing 1 mM L-arginine. RNI concentration in plasma of control rats was 32.0 +/- 3.4 microM (mean +/- SE). Livers from diet-fed control rats produced RNIs at the barely detectable rate of 7.8 +/- 1.5 nmol/hr x g wet liver. Six hr after administration of LPS (1 mg/kg, i.v.), plasma RNI levels in diet-fed control rats increased to 426.9 +/- 29.4 microM, and RNI release from the perfused liver was also markedly elevated to 97.7 +/- 7.7 nmol/hr x wet g liver, indicating hepatic NO release as a potentially important source for the increased RNI in plasma. The presence of NG-monomethyl-L-arginine (0.5-1 mM) or the absence of L-arginine in the perfusate inhibited LPS-induced stimulation of RNI release. EGTA (1 mM) had little effect, indicating that the increased RNI release was likely to be due to inducible NO synthase activity. The release of RNIs by freshly isolated Kupffer cells increased 13-fold, and this small cell mass contributed almost half of the hepatic RNI production under these conditions. Plasma ALT concentration was elevated after LPS administration, indicating incipient liver damage.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Oxidoreductases↗
PubMed1995

Acute endotoxin tolerance downregulates superoxide anion release by the perfused liver and isolated hepatic nonparenchymal cells.

This work is based on the hypothesis that low-dose lipopolysaccharide (LPS) suppresses the stimulatory and priming effects of a subsequent high-dose endotoxin on the formation of toxic oxygen-derived radicals by the perfused liver and isolated hepatic nonparenchymal cells. Such effects may in turn contribute to hyposensitivity to the lethal effect of large doses of endotoxin. Male Sprague-Dawley rats received a nonlethal ("low-dose") intravenous injection of Escherichia coli LPS (0.5 mg/kg body weight) 12 to 120 hours before they were challenged by a "large dose" of endotoxin (10 mg/kg). Three hours after LPS challenge, the livers were perfused, and superoxide release was determined. Nonparenchymal cells were also isolated for the determination of superoxide anion formation in vitro. There was a low rate (0.14 +/- 0.1 nmol/min/g liver weight) of superoxide generated by the perfused livers from rats that received the low-dose LPS 1 to 5 days previously. Control livers generated less than 0.08 nmol superoxide. A high rate (1.3 +/- 0.1 nmol/min/g) of superoxide release was measured in the perfused liver 4 hours after treatment of previously untreated control rats with large-dose LPS. This was attenuated to 0.7 +/- 0.04 nmol/min/g by an injection of low-dose LPS before challenge. This attenuation was time dependent; it failed to manifest at 12, 24, or 120 hours after low-dose LPS. Isolated endothelial cells, Kupffer cells, and sequestered hepatic neutrophils from rats given a high-dose LPS also generated significant amounts of superoxide both in the presence or absence of agonists, i.e., phorbol myristate acetate or opsonized zymosan.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Antineutrophil monoclonal antibody (1F12) alters superoxide anion release by neutrophils and Kupffer cells.

The formation of oxygen-derived radicals by phagocytes is regulated by chemotactic agents, cytokines, and adhesion molecules, such as CD11b/CD18 (Mac-1). In the rat system, we investigated the effect of monoclonal antibody 1F12 against rat neutrophils on hepatic sequestration of neutrophils and superoxide release by hepatic phagocytes. Within 15 min after 1F12 injection, there was profound neutropenia, which persisted for 24 h. The majority of the "lost" neutrophils were sequestered in the liver 4 h after treatment. Zymosan-induced superoxide release in vitro by isolated hepatic neutrophils from 1F12-treated rats was significantly attenuated at 4 and 24 h. The phorbol myristate acetate mediated superoxide release was inhibited 24 h after treatment. Superoxide anion release by normal adherent neutrophils in the presence of agonists was also inhibited by 1F12 in vitro. The in vivo administration of 1F12 primed the Kupffer cells to release superoxide. In vitro treatment of Kupffer cells with 1F12 also stimulated superoxide release. Monoclonal antibody WT.3 (also directed against rat neutrophils), which does not cause neutropenia, did not alter superoxide generation by neutrophils and Kupffer cells. These results indicate that 1F12 may be useful in attenuating inflammation and tissue injury associated with neutrophil activation. However, the activation of Kupffer cells to release toxic oxygen-derived metabolites may predispose the liver to injury in certain pathological conditions.

Animals↗
PubMed1994

The roles of nitric oxide and hydrogen peroxide production in lipopolysaccharide-induced intestinal damage.

The importance of gastrointestinal injury in endotoxin-induced shock and multiple organ failure is of great interest. In this paper we describe a method to assess the degree of intravascular congestion and bleeding into the wall of the intestine by determining the hemoglobin content of the tissue. After validating this method, we used it to study the mechanism of jejunal injury induced by intravenous injection of Escherichia coli lipopolysaccharide (LPS, 50 mg/kg bw), the role of nitric oxide release in maintaining the integrity of endothelial cells, and the participation of H2O2 production in the LPS-induced intestinal damage in rats. Our results show that after the administration of LPS at the dose of 50 mg/kg intravenously, the hemoglobin content of the jejunum (17.8 mg/100 mg tissue) increased 7.7-fold over that of control animals (2.3 mg/100 mg), reflecting a serious degree of congestion, bleeding, and damage in the gastrointestinal tract. Administration of nitro-L-arginine methyl ester (L-NAME) not only enhanced this injury, but also markedly decreased the dose of LPS necessary to induce intestinal damage. Infusion of L-arginine (300 mg/kg bolus plus infusion 600 mg/kg.h intravenously) protected the intestine against LPS or LPS plus L-NAME. Inhibition of basal nitric oxide release by L-NAME produced significant changes in cardiovascular variables, but failed to induce a significant bleeding damage. However, when inhibition of NO release was combined with enhanced H2O2 production by a small dose of LPS, a serious bleeding damage was observed. This was accompanied by a marked decrease in mesenteric blood flow and cardiac output. High dose of LPS induced the above effects, and thus could be responsible for the bleeding damage, while low dose of LPS that fails to inhibit nitric oxide, did not induce any intestinal bleeding. It seems that inhibition of NO release and stimulation of H2O2 production are both involved in the LPS-induced bleeding damage.

Animals↗
PubMed1994

Epinephrine-induced changes in hepatic glucose production after ethanol.

Previous studies indicate that catecholamines play an important role in mediating the glucose metabolic response to endotoxin. Because acute ethanol (EtOH) intoxication impairs this response, the present study was initiated to ascertain whether EtOH attenuates the lipopolysaccharide response by decreasing the increment in plasma catecholamines after endotoxin or by decreasing the responsiveness of rats to epinephrine. All studies were performed on chronically catheterized fasted rats infused intravenously with either EtOH or an equal volume of saline. In the first series of experiments, intravenous administration of Escherichia coli endotoxin increased, to the same extent, the plasma concentrations of epinephrine and norepinephrine in both saline- and EtOH-infused rats. In the second study, rats were infused with [3-3H]glucose to assess whole body glucose metabolism and the ability of EtOH to alter the glucose metabolic response to epinephrine. The exogenous infusion of a maximally stimulating dose of epinephrine (1 microgram.min-1.kg-1) into saline-infused control animals for 3 h produced a marked hyperglycemia that resulted from a sustained increase in the rate of hepatic glucose production and a reduction in the metabolic clearance rate for glucose. EtOH infusion did not prevent the epinephrine-induced hyperglycemia but blunted the stimulatory effect of epinephrine on glucose production. The differences in glucose metabolism between saline- and EtOH-treated rats could not be explained by changes in plasma insulin or glucagon concentrations. Furthermore, the ability of EtOH to impair the epinephrine-induced increase in glucose production was still evident in rats treated with 4-methylpyrazole, an inhibitor of alcohol dehydrogenase.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Acute alcohol administration attenuates insulin-mediated glucose use by skeletal muscle.

The aim of the present work was to test the effect of acute in vivo alcohol administration (180-190 mg/dl plasma for 3 h) on glucose utilization by tissues under basal conditions or after a hyperinsulinemic (100-130 microU/ml) euglycemic clamp in fasted rats. In vivo glucose use by individual tissues was assessed by the tracer 2-deoxy-D-glucose technique. Alcohol administration to saline-infused rats markedly inhibited glucose use by skeletal muscles, including the soleus, white and red quadriceps, and gastrocnemius, as well as by the heart. Ethanol infusion, however, had no effect on glucose use by the diaphragm, lung, liver, skin, ileum, brain, and adipose tissue. The insulin-stimulated glucose use was also inhibited by alcohol selectively in the muscles, with no effect on other tissues tested, except a moderate inhibition in the brain. Ethanol inhibited muscle glucose use by an average of approximately 50% under both basal and insulin-stimulated conditions. However, because insulin treatment more than doubled basal glucose use by these muscles, the 50% inhibition by ethanol treatment represents a greater inhibition of absolute glucose use under insulin-stimulated rather than under basal conditions. Our data demonstrate that acute alcohol intake attenuates basal and hormone-induced glucose utilization in a tissue-specific fashion. The inhibitory effect of alcohol on skeletal muscle glucose use could contribute to the previously observed decreased glucose recycling in humans after acute alcohol intake.

Animals↗
PubMed1994

Inhibition of nitric oxide formation in vivo enhances superoxide release by the perfused liver.

Nitric oxide, a known scavenger of toxic oxygen-derived radicals, has been shown to have a protective effect against tissue injury in endotoxemia. Based on the hypothesis that under normal physiological conditions, a balance between superoxide and nitric oxide exists in vivo, this work examines hepatic superoxide release after nitric oxide formation is inhibited in vivo. Male Sprague-Dawley rats were treated intravenously with N omega-nitro-L-arginine methyl ester (L-NAME; 50 mg/kg body wt), an inhibitor of nitric oxide synthase. One hour later, superoxide anion release by the perfused liver was determined. Results show that a significant amount of superoxide was released after L-NAME treatment. Likely sources of this radical are the Kupffer cells. Inhibition of nitric oxide formation in vivo did not enhance superoxide release by hepatocytes or sinusoidal endothelial cells. The effect of L-NAME treatment on superoxide release in endotoxemic rats was also examined 12 h after lipopolysaccharide treatment, when toxic oxygen-derived radical formation could not be detected. Inhibition of nitric oxide release in vivo in these rats enhanced the formation of superoxide anion. The interaction between nitric oxide and superoxide anion under normal conditions may represent an important protective mechanism of the host against free radical damage.

Animals↗
PubMed1994

Modulation of hepatic sinusoidal endothelial cell function by Kupffer cells: an example of intercellular communication in the liver.

We tested the hypothesis that Kupffer cells modulate sinusoidal endothelial cell function in the liver. Rats were treated with Kupffer cell-depleting agents (gadolinium chloride and liposome-encapsulated dichloromethylene diphosphonate) or with inhibitors of phospholipase A2 or leukotriene A4 synthase (dexamethasone and diethylcarbamazine, respectively). Hyaluronan uptake by the isolated, perfused liver was measured as an index of the functional state of the sinusoidal endothelial cell. Plasma hyaluronan concentration was also determined. Three hours after Escherichia coli lipopolysaccharide administration (100 micrograms/100 gm body wt, intravenously) plasma hyaluronan levels were significantly increased (280% to 320%), whereas hepatic hyaluronan uptake was markedly decreased (approximately 76%). Pretreatment with gadolinium chloride (0.5 mg/100 gm body wt, intravenously, 21 hr before saline solution or lipopolysaccharide administration), liposome-encapsulated dichloromethylene diphosphonate (40 mumol/100 gm body wt, intravenously, 44 hr before saline solution or lipopolysaccharide injection), dexamethasone (40 micrograms/100 gm body wt, intravenously, 1 hr before saline solution or lipopolysaccharide administration) or diethylcarbamazine (repeated doses, 10 mg/100 gm body wt, intravenously, 1 hr before saline solution or lipopolysaccharide injection) counteracted the lipopolysaccharide inhibitory effect on hepatic hyaluronan uptake. With the exception of gadolinium chloride, all other agents also prevented the lipopolysaccharide-induced increase in plasma hyaluronan concentration. Gadolinium chloride only attenuated the lipopolysaccharide effect on plasma hyaluronan level. Taken together with earlier results from our laboratory, these data indicate that: (a) Kupffer cell activation by lipopolysaccharide results in suppression of hyaluronan uptake by sinusoidal endothelial cells and (b) such modulation of endothelial cell function is likely mediated by products of the lipoxygenase pathway of arachidonate metabolism.

Animals↗
PubMed1993

Alcohol administration attenuates LPS-induced expression of inducible nitric oxide synthase in Kupffer and hepatic endothelial cells.

This study investigates the effects of in vivo ethanol (primed infusion, causing 170-190 mg% plasma alcohol for 12 hours) and/or LPS (12 hours after injection of E. coli LPS 1 mg/kg bw.) on the mRNA expression of inducible nitric oxide synthase (NOS II) in hepatic cells measured by competitive PCR technique, and on hepatic release of reactive nitrogen intermediates (RNI, NO2- + NO3-). Perfused livers from alcohol- or saline-infused animals did not release measurable amounts of RNI. Under these conditions small amounts of NOS II mRNA were expressed in Kupffer and endothelial cells, while it was not detectable in parenchymal cells. LPS treatment along with markedly elevating hepatic RNI release increased NOS II mRNA levels by 35- and 200-fold, in endothelial and Kupffer cells, respectively. LPS injection and alcohol infusion to the same animal decreased hepatic RNI release by about 70% and almost completely inhibited the LPS-induced, elevated NOS II mRNA in Kupffer or endothelial cells. No similar changes were observed in the parenchymal cells. These data suggest that the primary target of in vivo LPS in upregulating hepatic NO release are the nonparenchymal cells. Furthermore, alcohol inhibits the LPS-induced response which may influence immune-related hepatic function.

Amino Acid Oxidoreductases↗
PubMed1993

Primed pentose cycle activity supports production and elimination of superoxide anion in Kupffer cells from rats treated with endotoxin in vivo.

Glucose use and pentose cycle activity were determined in freshly isolated rat Kupffer cells 3 h after an i.v. injection of Escherichia coli endotoxin (0.1 mg/kg body weight), by using [1-14C], [6-14C] and [2-3H]glucose. Endotoxin treatment in vivo caused a 5-fold increase in the basal glucose uptake in Kupffer cells. Pentose cycle activity was elevated from 8.7 to 13.6 nmol/h per 10(7) cells after endotoxin. In vitro treatment of the cells from saline- and endotoxin-treated animals with phorbol ester (10(-6) M) increased pentose cycle activity 2-fold and 8-fold, respectively. Phorbol ester caused a 50% increase in glucose uptake in both groups. t-Butyl hydroperoxide (0.5 mM) caused a similar increase in pentose cycle activity as phorbol ester. Glucose oxidation in the Krebs cycle was also doubled after endotoxin. KC from endotoxin-treated animals produced O2- spontaneously, and were primed to produce additional large amounts of O2- upon phorbol ester treatment. Addition of t-butyl hydroperoxide inhibited O2- production by Kupffer cells. Depletion of glutathione by N-ethylmaleimide (0.1 mM), or inhibition of NADPH oxidase by diphenyliodonium (0.1 mM) inhibited both the pentose cycle activity and the O2- production. Increasing the concentration of exogenous glucose in the cell medium elevated the glycolytic rate, while pentose cycle flux was not affected either under basal conditions or following subsequent challenges by phorbol ester or t-butyl hydroperoxide. Our data suggest that the endotoxin-induced elevated glucose use in Kupffer cells is accompanied by a primed state of the pentose cycle. This condition supports superoxide and macromolecule synthesis and could also represent a potentiated protective mechanism against oxidative cellular injury during bacterial infections.

Animals↗
PubMed1993

Association of galactosamine-induced hepatitis in the rat with hyperhyaluronanaemia and decreased hyaluronan uptake by the isolated, perfused liver.

Plasma hyaluronan (HA) concentration and the rate of HA uptake by the isolated, perfused liver were measured in rats treated with saline, D-galactosamine (GaI-NH2, 50 mg/100 g body wt), gadolinium chloride (GdCl3) (0.5 mg/100 g body wt), and GdCl3 + GaI-NH2. GdCl3 was given 24 hr before GaI-NH2 or saline. Plasma L-alanine:2-oxoglutarate aminotransferase (EC 2.6.1.2), a marker for hepatocyte damage, was increased by 8 hr and remained elevated for 24 hr after GaI-NH2 injection. GdCl3 did not affect plasma enzyme levels when given alone or in association with, but prior to, GaI-NH2. Plasma HA levels were increased (200%) within 24 hr after GaI-NH2 administration. A plateau was reached at 8 hr, which was maintained for at least 24 hr. Although GdCl3 alone did not affect plasma HA levels, it slightly delayed the increase in HA concentration in GaI-NH2-treated rats. Livers, isolated 24 hr after GaI-NH2 treatment, exhibited a severe depression (approximately 67%) of HA uptake. GdCl3 did not prevent this suppression. The data presented indicate that: (1) one of the sinusoidal endothelial cell-dependent functions of the liver, i.e. removal of HA from the blood stream, is profoundly impaired during galactosamine-induced hepatitis, and (2) the adverse effect of GaI-NH2 on this sinusoidal endothelial cell function may not be dependent on CdCl3-suppressible Kupffer cell functions.

Alanine Transaminase↗
PubMed1993

Brief endotoxemia markedly increases expression of GLUT1 glucose transporter in Kupffer, hepatic endothelial and parenchymal cells.

Expression of various glucose transporter isoforms was studied in hepatic cells from fasted rats 3h after an injection of E. coli LPS (1mg/kg bw., i.v.). Glucose transporter isoform content of plasma membranes from hepatic cells was determined by western blot analysis using polyclonal antibodies. The predominant glucose transporter isoform expressed in parenchymal cells was GLUT2. GLUTS-1 and -4 were also observed to be present; however, GLUT4 protein was expressed to a relatively minor extent. GLUT3 was not detectable. LPS injection resulted in a 50% decrease in GLUT2 while GLUT1 protein doubled. GLUT4 was not altered after LPS. In the plasma membranes of Kupffer and endothelial cells only the GLUT1 isoform was detected. LPS treatment resulted in a 7- and 4-fold increase in the GLUT1 protein content in these cells. These data indicate that the predominant glucose transporter of hepatic nonparenchymal cells is the GLUT1 isoform and its synthesis and/or membrane translocation is augmented in response to LPS. The observed alterations in the contents of glucose transporters indicate adaptive changes to endotoxemia in the glucose consuming and glucose producing populations of hepatic cells.

Animals↗
PubMed1993

Augmented glucose use and pentose cycle activity in hepatic endothelial cells after in vivo endotoxemia.

Glucose use and pentose cycle activity were determined in freshly isolated rat hepatic endothelial cells 3 hr after an intravenous injection of Escherichia coli lipopolysaccharide (0.1 mg/kg body weight), by use of [1-14C]glucose, [6-14C]glucose and [2-3H]glucose. Lipopolysaccharide treatment in vivo increased glucose use fivefold, whereas glucose oxidation in the pentose cycle was elevated from 0.2 to 1.5 nmol/hr/10(7) cells. In vitro incubation of endothelial cells from saline- and lipopolysaccharide-treated animals in the presence of phorbol 12-myristate 13-acetate (10(-6) mol/L) increased pentose cycle activity twofold and eightfold, respectively. Phorbol 12-myristate 13-acetate caused only a 40% to 60% increase in glycolysis in both groups. Addition of t-butyl hydroperoxide (0.5 mmol/L), a substrate for glutathione peroxidase, caused a 24-fold and 16-fold increase in the glucose flux through the pentose cycle in cells from saline- and lipopolysaccharide-treated rats, respectively. Oxidation of glucose through the Krebs cycle was also increased several-fold after t-butyl hydroperoxide administration. Depletion of cellular glutathione by N-ethylmaleimide (0.1 mmol/L) inhibited the phorbol 12-myristate 13-acetate-induced or t-butyl hydroperoxide-induced increase in the pentose cycle activity with no marked effects on glycolysis. Diphenyleneiodonium (0.1 mmol/L), an inhibitor of superoxide and nitric oxide synthesis inhibited the phorbol 12-myristate 13-acetate-induced increased pentose cycle activity with no effects on the t-butyl hydroperoxide-induced response.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗