Eicosapentaenoic acid and gamma-linolenic acid induce apoptosis in HL-60 cells.

Enteral nutrition with eicosapentaenoic acid (EPA; 20:5 n-3) and gamma-linolenic acid (GLA; 18:3 n-6) decreased pulmonary inflammation by reducing neutrophil counts and chemotactic factors in bronchoalveolar lavage fluid during acute respiratory distress syndrome (ARDS). We hypothesize that the anti-inflammatory effects of EPA and GLA may be due, in part, to induction of neutrophil apoptosis. The purpose of this study was to determine whether EPA and GLA, alone or in combination, trigger apoptotic cell death in the human promyelocytic leukemia HL-60 cell line. HL-60 cells were incubated with 10, 20, 50, and 100 micromol/L EPA, GLA, or various combinations of EPA and GLA for 2, 4, 8, 12, and 24 hs. Oleic acid (18:1 n-9) was used as a fatty acid control. Flow cytometry using dual staining with propidium iodide and annexin V-FITC assessed apoptosis, necrosis, and viability. Apoptosis was verified by DNA fragmentation as assessed by agarose gel electrophoresis. EPA, GLA, and various combinations of EPA and GLA significantly induced apoptosis and reduced cell viability in HL-60 cells. Viability was significantly reduced to the same extent with the combination of 50 micromol/L EPA\20 micromol/L GLA compared with 100 micromol/L EPA. These data indicate that EPA and GLA, alone or in combination, reduce cell survival by induction of apoptosis. Thus, induction of apoptosis by select dietary n-3 (EPA) and n-6 (GLA) polyunsaturated fatty acids may be the mechanism of the resolution of pulmonary inflammation in ARDS.

Platelet-activating factor (PAF)-induced decreases in whole-body and skeletal muscle protein synthesis.

Platelet-activating factor (PAF) has been shown to reduce rat skeletal muscle amino acid uptake, which may restrict intracellular amino acid availability for protein synthesis and amino acid oxidation during endotoxemia. We investigated in rats the effect of PAF infusion on amino acid and protein metabolism by measuring (a) whole-body and tissue leucine kinetics; (b) plasma amino acid profile; and (c) muscle RNA activity (protein synthesis efficiency) and relative abundance of myofibrillar proteins. Fasted male Sprague-Dawley rats (250+/-20 g) were given a 4-h i.v. continuous infusion of L-(1-14C)-leucine to determine leucine kinetics during the infusion of PAF (2 microg/kg PAF as a priming i.v. bolus 1 h before a 4-h i.v. infusion of 2 microg/kg/h PAF) or vehicle. PAF infusion caused sustained hypotension, hyperglycemia, hematological alterations, and hyperlacticacidemia. Whole-body protein synthesis was decreased by 24% (P < 0.05) and leucine flux oxidized was increased by 23% (P < 0.05). Leucine flux was reduced, although not significantly (P = 0.07), in PAF-treated rats (n = 8) compared with controls (n = 8). PAF significantly decreased fractional protein synthesis in the rectus abdominus (33%), soleus (30%), and extensor digitorum longus (26%) muscles, but not in the liver. Plasma branched-chain amino acid levels decreased (approximately 30%, P < 0.05) in PAF-treated rats. Muscle RNA activity was 32% lower and myosin relative abundance declined whereas actin was unchanged in PAF-treated rats. PAF induced net protein catabolism as a result of elevated leucine oxidation at the expense of protein synthesis. PAF had the cumulative effects in the skeletal muscle of (a) attenuating amino acid uptake, (b) reducing protein synthesis efficiency, (c) decreasing fractional protein synthesis rate, and (d) decreasing myosin relative abundance. Thus, PAF may be an important mediator of decreased protein synthesis in skeletal muscle during endotoxic and septic shock.

Effect of enteral feeding with eicosapentaenoic acid, gamma-linolenic acid, and antioxidants in patients with acute respiratory distress syndrome. Enteral Nutrition in ARDS Study Group.

OBJECTIVES: Recent studies in animal models of sepsis-induced acute respiratory distress syndrome (ARDS) have shown that a low-carbohydrate, high-fat diet combining the anti-inflammatory and vasodilatory properties of eicosapentaenoic acid (EPA; fish oil), gamma-linolenic acid (GLA; borage oil) (EPA+GLA), and antioxidants improves lung microvascular permeability, oxygenation, and cardiopulmonary function and reduces proinflammatory eicosanoid synthesis and lung inflammation. These findings suggest that enteral nutrition with EPA+GLA and antioxidants may reduce pulmonary inflammation and may improve oxygenation and clinical outcomes in patients with ARDS. DESIGN: Prospective, multicentered, double-blind, randomized controlled trial. SETTING: Intensive care units of five academic and teaching hospitals in the United States. PATIENTS: We enrolled 146 patients with ARDS (as defined by the American-European Consensus Conference) caused by sepsis/pneumonia, trauma, or aspiration injury in the study. INTERVENTIONS: Patients meeting entry criteria were randomized and continuously tube-fed either EPA+GLA or an isonitrogenous, isocaloric standard diet at a minimum caloric delivery of 75% of basal energy expenditure x 1.3 for at least 4-7 days. MEASUREMENTS AND MAIN RESULTS: Arterial blood gases were measured, and ventilator settings were recorded at baseline and study days 4 and 7 to enable calculation of PaO2/FIO2, a measure of gas exchange. Pulmonary neutrophil recruitment was assessed by measuring the number of neutrophils and the total cell count in bronchoalveolar lavage fluid at the same time points. Clinical outcomes were recorded. Baseline characteristics of 98 evaluable patients revealed that key demographic, physiologic, and ventilatory variables were similar at entry between both groups. Multiple bronchoalveolar lavages revealed significant decreases (approximately 2.5-fold) in the number of total cells and neutrophils per mL of recovered lavage fluid during the study with EPA+GLA compared with patients fed the control diet. Significant improvements in oxygenation (PaO2/FIO2) from baseline to study days 4 and 7 with lower ventilation variables (FIO2, positive end-expiratory pressure, and minute ventilation) occurred in patients fed EPA+GLA compared with controls. Patients fed EPA+GLA required significantly fewer days of ventilatory support (11 vs. 16.3 days; p = .011), and had a decreased length of stay in the intensive care unit (12.8 vs. 17.5 days; p = .016) compared with controls. Only four of 51 (8%) patients fed EPA+GLA vs. 13 of 47 (28%) control patients developed a new organ failure during the study (p = .015). CONCLUSIONS: The beneficial effects of the EPA+GLA diet on pulmonary neutrophil recruitment, gas exchange, requirement for mechanical ventilation, length of intensive care unit stay, and the reduction of new organ failures suggest that this enteral nutrition formula would be a useful adjuvant therapy in the clinical management of patients with or at risk of developing ARDS.

Dietary fish oil and fish and borage oil suppress intrapulmonary proinflammatory eicosanoid biosynthesis and attenuate pulmonary neutrophil accumulation in endotoxic rats.

OBJECTIVE: Proinflammatory eicosanoids and cytokines are important mediators of local inflammation in acute lung injury. We determined if enteral nutrition with anti-inflammatory fatty acids, eicosapentaenoic acid, and gamma-linolenic acid would reduce the intrapulmonary synthesis of proinflammatory eicosanoids and cytokines and pulmonary neutrophil accumulation in a rat model of acute lung injury. DESIGN: Prospective, randomized, controlled, double-blind study. SETTING: Research laboratory at a university medical center. SUBJECTS: Male Long-Evans rats (250 g). INTERVENTIONS: Rats were randomly assigned to three dietary treatment groups and fed nutritionally complete diets (300 kcal/kg/day) containing 55.2% of the total calories from fat with either 97% corn oil, 20% fish oil, or 20% fish and 20% borage oil for 21 days. On day 22, bronchoalveolar lavage was performed 2 hrs after an intravenous injection of Salmonella enteritidis endotoxin (10 mg/kg) or saline. Bronchoalveolar lavage fluid was analyzed for leukotriene B4, leukotriene C4/D4, thromboxane B2, prostaglandin E2, 6 keto-prostaglandin F1alpha, tumor necrosis factor (TNF)-alpha, and macrophage inflammatory protein-2 (MIP-2). Lung myeloperoxidase activity (a marker for neutrophil accumulation) and phospholipid fatty acid composition were also determined. MEASUREMENTS AND MAIN RESULTS: Lung phospholipid concentrations of arachidonic acid were lower and the concentrations of eicosapentaenoic acid and docosahexaenoic acid were higher with fish oil and fish and borage oil as compared with corn oil. Dihomo-gamma-linolenic acid, the desaturated and elongated intermediate of gamma-linolenic acid, increased with fish and borage oil as compared with fish oil and corn oil. The levels of leukotriene B4, leukotriene C4/D4, 6-keto-prostaglandin F1alpha, and thromboxane B2 with corn oil were significantly increased with endotoxin as compared with saline. In contrast to the corn oil group, endotoxin did not significantly increase bronchoalveolar lavage levels of leukotriene B4, leukotriene C4/D4, and thromboxane B2 above those of saline-treated rats with fish oil and fish and borage oil. Lung myeloperoxidase activity was significantly increased in endotoxin-treated rats compared with those rats given saline in all dietary treatment groups. However, lung myeloperoxidase activity was significantly lower with either fish oil or fish and borage oil as compared with corn oil after endotoxin. Although endotoxin increased the levels of TNF-alpha and MIP-2 with all dietary treatment groups as compared with saline-treated rats, there were no significant differences in the levels of either cytokine between the dietary treatment groups. CONCLUSIONS: These results indicate that dietary fish oil and fish and borage oil as compared with corn oil may ameliorate endotoxin-induced acute lung injury by suppressing the levels of proinflammatory eicosanoids (but not TNF-alpha or MIP-2) in bronchoalveolar lavage fluid and reducing pulmonary neutrophil accumulation.

Effects of eicosapentaenoic and gamma-linolenic acid on lung permeability and alveolar macrophage eicosanoid synthesis in endotoxic rats.

OBJECTIVES: Proinflammatory eicosanoids (cyclooxgenase and lipoxygenase metabolites of arachidonic acid) released by alveolar macrophages play an important role in endotoxin-induced acute lung injury. We investigated the effect of prefeeding rats for 21 days with enteral diets that provided the anti-inflammatory fatty acids, eicosapentaenoic acid and gamma-linolenic acid (derived from fish oil and borage oil, respectively), as compared with an n-6 fatty acid-enriched diet (corn oil) on the following: a) lung microvascular protein permeability, arterial blood pressure, and platelet and white blood cells in a model of endotoxin-induced acute lung injury; b) alveolar macrophage prostaglandin and leukotriene synthesis; and c) liver and alveolar macrophage phospholipid fatty acid composition. DESIGN: Prospective, randomized, controlled, double-blind study. SETTING: Research laboratory at a university medical center. SUBJECTS: Male Long-Evans rats, weighing 250 g. INTERVENTIONS: Rats were randomized into four dietary treatment groups and fed nutritionally complete diets (300 kcal/kg/day), containing 55.2% of the total calories from fat with either 97% corn oil, 20% fish oil, 20% fish and 5% borage oil, or 20% fish and 20% borage oil for 21 days. On day 22, lung microvascular protein permeability, mean arterial pressure, and platelet and white blood cell counts were determined for 2 hrs after an intravenous injection of Salmonella enteritidis endotoxin (10 mg/kg). In a second group of prefed rats, the phospholipid fatty acid composition was determined in liver and alveolar macrophages. Alveolar macrophages were harvested by bronchoalveolar lavage and stimulated in vitro with a calcium ionophore (A23187), and the concentrations of leukotrienes B4 and B5, thromboxane A2, prostaglandin E2, and 6-keto-prostaglandin F1 alpha were measured in a third group of prefed rats. MEASUREMENT AND MAIN RESULTS: Lung permeability was greatest with corn oil and was significantly attenuated with 20% fish oil and 20% fish and 5% borage oil, and this effect approached significance with 20% fish and 20% borage oil (p = .06). The early and late hypotensive effects of endotoxin were attenuated with 20% fish oil, 20% fish and 5% borage oil, and 20% fish and 20% borage oil, as compared with corn oil. Concentrations of leukotriene B4, prostaglandin E2, and thromboxane B2 released from A23187-stimulated alveolar macrophages were significantly lower with 20% fish oil and 20% fish and 20% borage oil, as compared with corn oil. The increase in lung microvascular protein permeability with 20% fish and 20% borage oil was not significantly different than the lung microvascular protein permeability that was found in animals receiving 20% fish oil (p = .20) and 20% fish and 5% borage oil (p = .31). Alveolar macrophage and liver phospholipid concentrations of arachidonic acid were lower, and the concentrations of eicosapentaenoic acid and docosahexaenic acid were higher, with 20% fish oil, and 5% borage oil, and 20% fish and 20% borage oil, as compared with corn oil. Dihomo-gamma-linolenic acid, the desaturated and elongated intermediate of gamma-linolenic acid, was increased with 20% fish and 20% borage oil, as compared with 20% fish oil and 20% fish and 5% borage oil. CONCLUSIONS: The severity of pulmonary microvascular protein permeability and the degree of hypotension were reduced with fish or fish and borage oil diets, as compared with corn oil, in endotoxic rats. The reduced synthesis of the proinflammatory arachidonic acid-derived mediators, leukotriene B4, thromboxane B2, and prostaglandin E2 from stimulated alveolar macrophages was indicative of a decrease in arachidonic acid and an increase in eicosapentaenoic acid and docosahexaenoic acid in cell membrane phospholipids.

The effect of LPS on cytokine synthesis and lung neutrophil influx after hepatic ischemia/reperfusion injury in the rat.

OBJECTIVE: To determine if cytokine responses and lung injury induced by intravenous (i.v.) lipopolysaccharide (LPS) at 4 hr were enhanced in rats that had been previously subjected to 30 min of total liver ischemia (Pringle's maneuver) followed by 24 hr or 3 days of reperfusion. BACKGROUND: Many patients with liver trauma require occlusion of hepatic blood flow to control hemorrhage and facilitate repair. A significant number of these patients subsequently develop the systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction (MOD) characterized by the release of cytokines and tissue neutrophil influx. Macrophages, including Kupffer cells, may be activated by ischemic injury and dysregulation of their response to LPS may contribute to the development of SIRS and acute respiratory distress syndrome. METHODS: Adult male Sprague-Dawley rats were randomly divided into six groups: three groups received total hepatic ischemia for 30 min and three groups had a sham procedure. Twenty-four hours or 3 days after hepatic ischemia/reperfusion injury, rats were treated with LPS (5 mg/kg) or saline and monitored for 4 hr. We collected serum, bronchoalveolar lavage (BAL) fluid, and lung tissue. RESULTS: Serum and BAL cytokine concentrations were significantly increased by i.v. LPS; however, hepatic ischemia/reperfusion injury 24 hr or 3 days before iv LPS ameliorated this cytokine response. The LPS-induced pulmonary neutrophil influx and histopathological changes were similar in sham and hepatic ischemia/reperfusion-injured groups. CONCLUSIONS: Hepatic ischemia/reperfusion injury significantly attenuated the serum and BAL cytokine concentrations, but did not change pulmonary neutrophil influx or histopathological alterations in response to i.v. LPS.

Intestinal digestion, absorption, and transport of structured triglycerides and cholesterol in rats.

We compared the intestinal absorption of trilinolein (1,2,3-tri-[1-14C]linoleyl-sn-glycerol) with two different structured triglycerides containing one linoleic acid (C18:2) and two octanoic acids (C8:0), 1,3-dioctanoyl-2-[1-14C]linoleyl-sn-glycerol (2-linoleate) and 1,2-di[1-14C]octanoyl-3-linoleyl-sn-glycerol (1,2-octanoate), respectively. Lymphatic radioactive lipid output of 2-linoleate resembled that of trilinolein rats but remained significantly lower during the lipid infusion. Radioactive lipid was recovered along the entire small intestinal lumen, with a significantly higher amount of [14C]lipid recovered in the lower small intestine and cecum in the 2-linoleate group. Delayed uptake of radioactive 2-linoleate was not due to poor digestion. In contrast, 1,2-octanoate was efficiently digested, and both the free fatty acid (FFA) and the monoacylglycerol (MG) containing octanoate were rapidly absorbed. Irrespective of its position on the triglyceride molecule, 14C-labeled octanoate was poorly transported into lymph. In addition, intestinal luminal and mucosal recovery of [14C]octanoate was significantly lower in the 1,2-octanoate group compared with [14C]linoleate recovery in the 2-linoleate or trilinolein groups. Total recovery of infused radioactive lipid was significantly less in the 1,2-octanoate group than in the 2-linoleate or trilinolein groups. Thus radioactive octanoate in the form of FFA or 2-MG was rapidly absorbed and transported via the portal vein. The infusion of either 2-linoleate or 1,2-octanoate did not affect the absorption and lymphatic transport of cholesterol compared with trilinolein. In summary, the type of the fatty acid on the structured triglyceride molecule affects its digestion, absorption, and lymphatic transport. Structured triglycerides containing octanoic acid in the 1- and 3-positions and linoleic acid in the 2-position may not be advantageous to use as a sole source of dietary lipid, but should be supplemented with long-chain triglycerides.

The role of platelet-activating factor in alterations of system A amino acid transport in rat soleus and extensor digitorum longus muscles during endotoxic shock.

We investigated the role of platelet-activating factor (PAF) as a mediator of system A amino acid transport alterations in skeletal muscle during endotoxic shock. Male Sprague-Dawley rats (80-100 g) were injected with Salmonella enteritidis endotoxin (10 mg/kg intravenously (i.v.)) or PAF (4 micrograms/kg i.v.) and killed 5 or 1 h later, respectively. Control rats were injected with a vehicle. System A amino acid transport was assessed by measuring the cellular uptake of 1-14C-alpha-aminoisobutyric acid (AIB, amino acid analog) in isolated soleus and extensor digitorum longus (EDL) muscles, in vitro. AIB uptake in soleus and EDL from endotoxic rats was approximately 33% lower than control muscles. The i.v. injection of PAF reduced AIB uptake 9% in soleus and 15% in EDL as compared with muscles from control rats. The prophylactic administration of WEB 2086 (30 mg/kg i.v.), a PAF receptor antagonist, attenuated the endotoxin-induced inhibition of amino acid transport by 26% in EDL and 17% in soleus. PAF (1 microgram/mL) added to incubation media had no effect on AIB uptake in soleus and EDL of control rats. However, there was a reduction in AIB uptake in soleus and EDL obtained from rats 1 h after an i.v. injection of PAF (4 micrograms/kg) and after incubation in media containing PAF (1 microgram/mL). Addition of plasma to incubation media obtained from rats 1 h after the i.v. injection of endotoxin or PAF attenuated AIB uptake in soleus and EDL of control rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Sepsis-induced myofibrillar protein catabolism in rat skeletal muscle.

This study evaluated sepsis-induced changes in myosin heavy chain (Mhc) protein breakdown and synthesis in rat soleus muscles. Rats were anesthetized and their external jugular veins were cannulated. After 12-16 h, rats were implanted intraabdominally with a sterile fecal pellet, or a pellet containing bacteria (Escherichia coli, 150 CFU and Bacteroides fragilis 10(4) CFU). Thirty hours after implantations, rats were infused with 14C-Leu (60 x 10(3) Bq/h) through the jugular cannula for 4 h. Protein fractional synthetic rate coefficient (FSRC) was determined in muscles of different rat groups. In separate experiments, intact soleus muscles were removed from the three rat groups on days 1 and 2 after implantations, and processed for their wet weight, total protein and Mhc contents. No mortality occurred in sterile-implanted rats. Approximately 40-45% of all septic-implanted rats died on days 1-3, post-implantation. Whereas an approximately 15% (P < 0.01, days 1 or 2) decrease occurred in Mhc content in sterile-implanted rats compared to unoperated controls, septic insult resulted in a greater Mhc loss (a 27% decrease, P < 0.001). Rats' body weight, soleus wet weight and tolat muscle protein changes with sepsis relative to controls were also greater than in the sterile groups. The FSRC value in the septic-implanted rats was significantly lower (P < 0.05) than in non-septic rat muscle. TNF-alpha administration to the septic animals or their treatment with diltiazem did not have a significant effect on FSRC. Overall, these results indicate myosin as a major muscle protein subjected to net catabolism during sepsis, and that the net catabolic response was related to a more pronounced increased in Mhc degradation than the decrease in Mhc synthesis.

Effect of intravenous lipid emulsions enriched with gamma-linolenic acid on plasma n-6 fatty acids and prostaglandin biosynthesis after burn and endotoxin injury in rats.

OBJECTIVE: To study the effect of intravenous lipid emulsions enriched with gamma-linolenic acid on plasma fatty acids and series-2 prostaglandins to determine if the slow conversion of linoleic acid by delta-6-desaturase to gamma-linolenic acid could be bypassed to provide substrate for the formation of dihomo-gamma-linolenic acid, the immediate precursor for series-1 prostaglandins, in control and injured rats. Dihomo-gamma-linolenic acid can also compete with arachidonic acid for oxidative metabolism by cyclooxygenase to modulate series-2 prostaglandin biosynthesis. DESIGN: Prospective, randomized, controlled, double-blind study. SETTING: Research laboratory at a university medical center. SUBJECTS: Thirty-three control and thirty-one injured male Sprague-Dawley rats were randomized into one of four parenteral dietary treatment groups. INTERVENTIONS: Rats were injured by the combined actions of a 30% body surface area full-thickness skin burn and a nonlethal injection of endotoxin (1 mg/kg ip). The rats were parenterally fed 200 kcal/kg/day, 1.5 g nitrogen/kg/day, and 30% of nonprotein calories as lipid (20% soybean lipid emulsion enriched with 2.7%, 4.4%, or 6.1% gamma-linolenic acid derived from borage oil) for 3 days. Control rats were treated similarly but were not injured. A 20% soybean/safflower oil lipid emulsion was used as the control diet (0% gamma-linolenic acid). Plasma was analyzed on day 3 to determine the concentrations of total fatty acids, thromboxane B2, 6-keto-prostaglandin F1 alpha, and bicyclo-prostaglandin E. MEASUREMENTS AND MAIN RESULTS: Parenteral nutrition with 2.7%, 4.4%, and 6.1% gamma-linolenic acid increased the plasma percentages (mol%) of gamma-linolenic acid and dihomo-gamma-linolenic acid in a dose-dependent fashion in control and injured rats. Supplementation with gamma-linolenic acid did not increase the plasma percentage of arachidonic acid as compared with the 0% gamma-linolenic acid lipid emulsion in control and injured rats. The ratio of dihomo-gamma-linolenic acid to arachidonic acid was significantly increased in response to 4.4% and 6.1% gamma-linolenic acid in both the control and injured groups. The plasma ratio of thromboxane B2 to 6-keto-prostaglandin F1 alpha was substantially reduced with gamma-linolenic acid compared with 0% gamma-linolenic acid in injured rats. Bicyclo-prostaglandin E concentration was significantly higher with 2.7% gamma-linolenic acid in injured rats. Injured rats were protein catabolic, as evidenced by a net negative nitrogen balance and loss of body mass compared with controls, but neither group showed overt signs of intolerance to the diets. CONCLUSIONS: Supplementation of parenteral nutrition with gamma-linolenic acid had the following effects: a) increased plasma gamma-linolenic acid, dihomo-gamma-linolenic acid, and bicyclo-prostaglandin E; b) increased the plasma ratio of dihomo-gamma-linolenic acid to arachidonic acid; and c) favorably reduced the ratio of thromboxane B2 to 6-keto-prostaglandin F1 alpha in injured rats. These results reflect the potential capacity of gamma-linolenic acid-enriched lipid emulsions to have the following actions: a) to increase dihomo-gamma-linolenic acid, which is the fatty acid precursor of the antiaggregatory, anti-inflammatory eicosanoid, prostaglandin E1; and b) to modulate arachidonic acid-derived series-2 prostaglandins after injury.

Effect of platelet-activating factor on basal and insulin-mediated system A amino acid transport in rat soleus muscle.

We investigated the effect of platelet-activating factor (PAF) on basal and insulin-stimulated sodium-dependent neutral amino acid transport by system A in rat soleus muscle. Fasted male Sprague-Dawley rats (75-100 g) were given an intravenous injection of saline or PAF (4 micrograms/kg) and killed 1 hr later. Isolated soleus muscles were incubated in media containing 1-14C-alpha-aminoisobutyric acid (AIB; system A amino acid analog), 3H-inulin, and 0 (basal) or 100 mU/ml insulin. Na(+)-free media were assessed by Na(+)-independent AIB uptake. The rate of AIB uptake by muscle was corrected for uptake into extracellular (inulin) space. Basal cellular AIB uptake by soleus muscles from PAF-treated rats was 30.2% lower than in controls. Insulin increased the absolute rate of cellular AIB uptake above basal levels to the same extent in muscles of control and PAF-treated rats, although the rate of maximal insulin-stimulated AIB uptake was 20% lower in muscles of PAF-treated rats. Na(+)-independent AIB uptake (measured in Na(+)-free media) was the same in muscles of control and PAF-treated rats. The fact that AIB uptake in Na(+)-free media was the same in muscles of control and PAF-treated rats indicates that the reduction in basal and insulin-stimulated AIB uptake in muscles of PAF-treated rats was due to an alteration of system A Na(+)-dependent amino acid transport.

Methylprednisolone does not influence endotoxin translocation during cardiopulmonary bypass.

This study investigated whether the prophylactic administration of methylprednisolone sodium succinate (MPSS) could prevent an increase in plasma endotoxin levels during cardiac surgery with cardiopulmonary bypass. MPSS (1 g/patient) or saline was given intravenously with induction in the steroid (n = 6) and control (n = 7) groups, respectively. Blood samples were collected preinduction and postinduction, during and after cardiopulmonary bypass, and 1 and 24 hours postoperatively. Plasma endotoxin was determined by a chromogenic Limulus amebocyte lysate assay. There was an intraoperative increase in the level of plasma endotoxin that occurred primarily after initiation of cardiopulmonary bypass and removal of the aortic cross-clamp. Endotoxin at 1 and 24 hours postoperatively was lower than the peak intraoperative levels and approached the preinduction level in both groups. The pump prime and other administered fluids contained low levels of endotoxin that were at or below the preinduction or postinduction level of the patients. MPSS did not prevent or attenuate the degree of endotoxemia during cardiopulmonary bypass. The loss of normal gut mucosal barrier function during cardiopulmonary bypass may result in endotoxemia and/or bacterial translocation, either of which could initiate or contribute to postoperative complications.

Effect of endotoxic shock on basal and insulin-mediated Na+/K(+)-pump activity in rat soleus muscle.

This study evaluated basal and insulin-mediated Na+/K+ transport in skeletal muscle during endotoxic shock. Fasted male Holtzman rats (80-100 g) were killed 5 hr after the i.v. injection of saline (control) or 20 mg/kg Salmonella enteritidis endotoxin. 86Rb+ uptake into the cellular compartment of soleus muscle was determined in the presence and absence of 100 mU/ml insulin. The rate coefficient and rate of 22Na+ efflux and muscle intracellular 22Na+ content (in percentage of total muscle 22Na+) were determined in the presence and absence of 1 mM ouabain. The rate of basal cellular 86Rb+ uptake (cpm x 10(3).g wet weight-1.min-1) was not significantly different between control (77.7 +/- 2.9) and endotoxic soleus muscles (76.1 +/- 3.9). Insulin increased the rate of cellular 86Rb+ uptake by 19% in control (92.4 +/- 3.7) and 23% in endotoxic soleus muscles (93.8 +/- 4.1). Cellular 22Na+ content was 40% greater in endotoxic muscle (16.8 +/- 1.4) as compared to control muscles (12.0 +/- 0.9). The rate coefficient for ouabain-sensitive 22Na+ efflux in endotoxic muscles (0.028 min-1) was 1.7-fold greater than that in control muscles (0.017 min-1). The rate of ouabain-sensitive 22Na+ efflux was more than doubled in endotoxic muscles (0.470%/min) compared with control muscles (0.204%/min). Endotoxic shock did not alter insulin's ability to stimulate Na+/K+ transport in muscle. An increase in the electrogenicity of the Na+/K+ pump is consistent with an increased rate coefficient for 22Na+ efflux, with no change in the rate of 86Rb+ uptake in endotoxic soleus muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

Total parenteral nutrition with short- and long-chain triglycerides: triacetin improves nitrogen balance in rats.

Little is known about the long-term metabolic effects of parenteral administration of short-chain triglycerides. These studies were undertaken to investigate triacetin, the water-soluble triglyceride of acetate when it is incorporated into nutritionally balanced total parenteral nutrition formulas. Male Sprague-Dawley rats (n = 22) were fed an isovolemic, isocaloric and isonitrogenous diet for 7 d. The lipid energy represented 30% of the nonprotein energy with short-chain triglycerides representing 0, 50 or 90% of the lipid energy. Plasma acetate concentration was determined as well as indicators of protein metabolism: daily and cumulative nitrogen balance, whole body leucine kinetics and rectus muscle and liver fractional protein synthetic rates. No overt toxic effects were observed at any point during the study. As the proportion of short-chain triglycerides in the diet increased from 0 to 50 or 90% of the lipid energy, cumulative nitrogen balance increased 50 or 120%, respectively (P less than 0.05). Whole-body and tissue leucine kinetics (determined during the last 2.5 h of the 7-d study) were unaffected by the lipid composition of the diet. Plasma acetate concentration was not significantly different among groups. These results indicate that incorporation of the short-chain triglyceride, triacetin, in nutritionally balanced total parenteral nutrition formulas improves nitrogen balance with no overt toxic effects. These data indicate that triacetin may have a future role as a parenteral nutrient, and that further studies of its use are warranted.

Parenteral nutrition with short- and long-chain triglycerides: triacetin reduces atrophy of small and large bowel mucosa and improves protein metabolism in burned rats.

The effect of total parenteral nutrition with combinations of long-chain triglycerides (LCTs) and triacetin, the water-soluble triglyceride of acetate, on structural components of the gastrointestinal tract and protein metabolism was assessed in burned (30% body surface area) rats. Rats received isovolemic, isocaloric, and isonitrogenous diets that delivered 672 kJ.kg-1.d-1 (160 kcal.kg-1.d-1), 9.6 g amino acids.kg-1.d-1, and 30% nonprotein calories as 90% triacetin/10% LCTs, 50% triacetin/50% LCTs, or 100% LCTs for 7 d. Daily and cumulative nitrogen balances and whole-body leucine kinetics and fractional protein synthetic rates in rectus muscle and liver were determined on the last day of nutrition. DNA, protein, and total weight were determined in mucosal scrapings from segments of jejunum and colon. Plasma acetate concentrations were substantially higher in both triacetin groups. Parenteral nutrition with 50% triacetin and 50% LCTs promoted a positive nitrogen balance similar to that of 100% LCTs, increased protein in rectus muscle and liver, smaller and more numerous mucosal cells in jejunum and colon, and increased colonic mucosal weight compared with the other groups. Triacetin did not appreciably affect whole-body and tissue leucine kinetics. The equicaloric provision of triacetin and LCTs improved protein utilization and structural components of the small and large bowel and reduced the development of intestinal mucosal atrophy associated with conventional parenteral nutrition in burn injury.

Effect of total parenteral nutrition with xylitol on protein and energy metabolism in thermally injured rats.

The use of xylitol as an alternative carbohydrate calorie source in total parenteral nutrition may offer unique pharmacologic and nutritional properties in the therapy of the thermally injured. Male Sprague-Dawley rats (250 g) received a 15-second dorsal scald injury (25-30% BSA) and were parenterally fed isovolemic diets (60 ml/day) that provided 200 kcal/kg/d, 9.68 g of amino acids/kg/d, and 23.5% nonprotein calories (NPC) as fat for 3 days. The balance of NPC were provided as dextrose (Dex) or 50% xylitol:50% dextrose (Xyl/Dex). Rectus muscle and liver fractional protein synthetic rates (FSR, %/day), whole body leucine appearance (Flux), oxidation (OX), protein breakdown (PB), and synthesis (PS) were estimated using a 4-hour iv infusion of [1-14C]leucine on day 3. Mean values (+/- SE) for leucine kinetics (mumol leucine/hr/100 g), cumulative nitrogen balance (mg N) and plasma insulin concentration (Table I). (microU/mL). The partial replacement of dextrose calories with xylitol did not significantly alter whole body and tissue leucine kinetics, daily and cumulative nitrogen balance, insulin concentration, and energy expenditure (indirect calorimetry). These data indicate that xylitol may be useful as an alternative carbohydrate calorie source in parenteral nutrition to avoid possible deleterious side effects of glucose overfeeding in the critically ill but did not improve protein metabolism under the conditions of this study.

Enteral nutrition with structured lipid: effect on protein metabolism in thermal injury.

The effect of total enteral nutrition with structured and conventional lipids on protein and energy metabolism was assessed in gastrostomy-fed burned rats (30% body surface area) by measuring nitrogen balance, serum albumin, energy expenditure, and rectus muscle and liver fractional synthetic rates of protein (FSRs). Male Sprague-Dawley rats weighing 200 +/- 10 g received isovolemic diets that provided 50 kcal/d, 2 g/d amino acids, and 40% nonprotein calories as lipid for 3 d. The lipid source was either long-chain triglycerides (LCTs), medium-chain triglycerides (MCTs), structured lipid (SL), or a physical mix (PM) of the oils used in SL. Burned rats enterally fed either SL (p less than 0.01) or PM (p less than 0.05) yielded significantly higher daily and cumulative nitrogen balances and rectus muscle and liver FSRs than those fed either LCTs or MCTs. Rats fed SL or MCTs maintained higher serum albumin concentrations than rats fed either PM or LCTs. This study shows that the enteral administration of a mixed fuel system containing SL or its PM improves protein anabolism and attenuates net protein catabolism after thermal injury.

Enhanced skeletal muscle and liver protein synthesis with structured lipid in enterally fed burned rats.

We assessed the effects of total enteral nutrition with long-chain triacylglycerides (LCT), medium-chain triacylglycerides (MCT), or two structured lipids, modified dairy fat (MDF) and modified MCT (Captex 810B, Capital City Products, Columbus, OH), on protein and energy metabolism in hypermetabolic burned rats (25% to 30% body surface area). Male Sprague-Dawley rats (200 +/- 10 g) were continuously gastrostomy-fed isovolemic diets that provided 50 kcal/d, 2 g amino acids/d and 40% nonprotein calories as lipid for three days. Changes in body weight, nitrogen balance, serum albumin, indirect calorimetry, whole body leucine kinetics, and rectus muscle and liver protein kinetics were determined. Whole body leucine kinetics and tissue fractional protein synthetic rates (FSR, percent per day) were estimated using a four-hour constant intravenous infusion of L-[1-14C]leucine on day 3. The group of rats enterally fed MDF lost less body weight than the other groups (P less than or equal to .05). MDF and Captex 810B produced a positive and significantly greater (P less than or equal to .05) daily and cumulative nitrogen balance than either LCT or MCT. Oxygen consumption (P less than or equal to .05) and total energy expenditure (P less than or equal to .05) were elevated approximately 22% with MDF as compared with LCT or MCT. Rectus muscle FSR and absolute rate of protein synthesis were increased 19% with MDF (P less than or equal to .05) as compared with LCT or MCT.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of burn and first-pass splanchnic leucine extraction on protein kinetics in rats.

The effects of burn and first-pass splanchnic leucine extraction (FPE) on protein kinetics and energy expenditure were assessed by measuring O2 consumption, CO2 production, nitrogen balance, leucine kinetics, and tissue fractional protein synthetic rates (FSR-%/day) in enterally fed rats. Anesthetized male rats (200 g) were scalded on their dorsum with boiling water (25-30% body surface area) and enterally fed isovolemic diets that provided 60 kcal/day and 2.4 g of amino acids/day for 3 days. Controls were not burned. An intravenous or intragastric infusion of L-[1-14C]leucine was used to assess protein kinetics on day 3. FPE was taken as the ratio of intragastric to intravenous plasma leucine specific activity. There was a 69% reduction in cumulative nitrogen balance (P less than 0.001) and a 17-19% increase in leucine oxidation (P less than 0.05) and total energy expenditure (P less than 0.01) in burned rats. A 15% decrease in plasma leucine clearance (P less than 0.05) was accompanied by a 20% increase in plasma [leucine] (P less than 0.01) in burned rats. Burn decreased rectus muscle FSR from 5.0 +/- 0.4 to 3.5 +/- 0.5 (P less than 0.05) and increased liver FSR from 19.0 +/- 0.5 to 39.2 +/- 3.4 (P less than 0.01). First pass extraction of dietary leucine by the splanchnic bed was 8% in controls and 26% in burned rats. Leucine kinetics corrected for FPE showed increased protein degradation with burn that was not evident without FPE correction. This hypermetabolic burn model can be useful in the design of enteral diets that optimize rates of protein synthesis and degradation.

Studies in small bowel transplantation. Prevention of graft-versus-host disease with preservation of allograft function by donor pretreatment with antilymphocyte serum.

Donor pretreatment with antilymphocyte serum (ALS) effectively prevents graft-versus-host disease (GVHD) in a unidirectional (parent-to-F1 hybrid) rat small bowel transplantation model. ALS must be administered prior to or at the time of transplantation, and the intraperitoneal route is more effective than subcutaneous administration. Donor pretreatment with ALS uniformly prevents GVHD without impairing subsequent allograft function as measured by absorption of dietary energy and nitrogen, weight gain, and bowel morphology. These rodent studies suggest that ALS treatment of donors as well as recipients in small bowel transplantation may be a highly effective, simple, and easily applicable method to prevent or ameliorate GVHD in human small bowel transplantation.