James P. Filkins

Altered glucose transporter mRNA abundance in a rat model of endotoxic shock.

To better understand molecular mechanisms of glucose transport in shock, we studied glucose transporter isoform mRNA abundance after injection of S. enteritidis endotoxin (40 mg/kg) or saline. Six to 8 hours after injection, endotoxin-treated animals compared to controls became hypoglycemic (44 +/- 6 vs. 111 +/- 4 mg/dl) and lactacidemic (5.9 +/- 0.5 vs. 1.3 +/- 0.1). At such times, tissue RNA was isolated and hybridized to Riboprobes for GLUT1 (erythrocyte), GLUT2 (liver), and GLUT4 (muscle/fat) glucose transporter isoforms and expressed as percent of control. GLUT1 mRNA abundance was increased in fat (660%, p less than .05), soleus muscle (314%, p less than .05), and liver (871%, p less than .001) of endotoxin-treated rats. Soleus muscle GLUT4 mRNA levels were increased (+33%, p less than .02), while liver GLUT2 mRNA levels were markedly decreased (-58%, p less than .01). The overall increase in GLUT1 mRNA abundance accompanied by lowered liver GLUT2 mRNA levels may either cause or reflect profoundly altered glucose transport.

Effects of Lead Acetate on Sensitivity to Shock, Intravascular Carbon and Endotoxin Clearances, and Hepatic Endotoxin Detoxification

Summary Lead acetate iv at 5 mg/300 g to male rats of the Holtzman strain sensitized to the lethal shock induced by endotoxin, Noble-Collip tumbling trauma, hind limb ischemia, bowel ischemia, and horse serum anaphylaxis. Lead treatment depressed the intravascular phagocytic clearance of colloidal carbon as evaluated at 15, 30, 60, 120 and 240 min. By use of an endotoxin bioassay based on lethality in lead-treated assay rats, the intravascular removal of a test dose of 0.5 mg of endotoxin was not altered by lead treatment as compared to normal and sodium acetate treated controls. The ability of liver homogenates to detoxify endotoxin was not altered by lead administration at 0.5, 1, 2, 3, or 8 hr prior to sacrifice. The in vitro addition of lead to homogenates incubated with endotoxin did not alter detoxification. It is suggested that the mechanism of lead sensitization to endotoxin probably does not reflect alteration in its rate of clearance or detoxification but rather may relate to leads ability to alter metabolic responses during endotoxemia.

In Vivo vs In Vitro Effects of Endotoxin on Glycogenolysis, Gluconeogenesis, and Glucose Utilization

Summary Salmonella enteritidis lipopoly-saccharide administered iv to male rats of the Holtzman strain resulted in increased glycogenolysis and decreased gluconeogene-sis in isolated hepatocytes, increased glucose uptake in isolated hemidiaphragms, and enhanced glucose oxidation in epididy-mal fat pads. Endotoxin added in vitro to the identical test systems failed to affect the rates of glycogenolysis, gluconeogenesis, glucose uptake, and glucose oxidation. A mediated as opposed to a direct mechanism of endotoxin action is supported.

Factors Modifying Susceptibility to Bacterial Endotoxin: The Effect of Lead and Cadmium

AbstractThe current interest in the biologic activity of endotoxins and the factors modifying host response to bacterial endotoxins stem, in part, from the fact that endotoxemia is of increasing clinical concern.1 It has been estimated that as many as 300, 000 patients are hospitalized each year due to Gram-negative bacteremia, and that the fatality rate of these patients may run as high as 30 to 50%.1 Experimental models in which there is an induction of endotoxin hypersensitivity have been employed to elucidate the fundamental mechanisms essential for survival in endotoxemia. Factors modifying host response to endotoxin are also of interest from a toxicological point of view, since such studies may delineate potential, detrimental interaction of toxic environmental or clinically employed agents with endotoxins. Some of the diverse factors employed to induce sensitization of animals to endotoxins include carbon tetrachloride,2 an attenuated strain ofMycobacterium, Bade Calmette Guerin,3 actinomycin D4: g...

Hypoglycemia and Depressed Hepatic Gluconeogenesis During Endotoxicosis in Lead-sensitized Rats

Summary The administration of Salmonella enteritidis endotoxin to male rats rendered hypersensitive by lead acetate resulted in profound hypoglycemia, lactacidemia, and depletion of liver glycogen. Lead acetate depressed gluconeogenesis as studied in isolated hepatocytes. Possible mechanisms for leads action on gluconeogenesis and the significance of this metabolic lesion in lead hypersensitivity to shock are discussed.

Blood Endotoxin Inactivation after Trauma and Endotoxicosis

Summary In contrast to the inability to demonstrate endotoxin detoxifying activity in either control rat blood, blood cells, plasma, or serum, the induction of mild endotoxemia or trauma resulted in a prompt and marked detoxifying activity in blood plasma or serum, but not in the cellular elements. The functional significance of the blood anti-endotoxin system was demonstrated by a capability for passive transfer and by its loss during severe traumatic shock. A role for the RES in elaboration of the blood anti-endotoxin system is postulated.

Characterization of gluconeogenesis in enzymatically isolated parenchymal cells of rat liver.

Summary In vitro studies of the structural and metabolic integrity of dispersed parenchymal cells prepared by perfusion of isolated rat livers with collagenase and hyaluronidase were performed. A high percentage of trypan blue exclusion, a low degree of soluble enzyme release into the incubation media, and a constant rate of gluconeogenesis from lactate, as well as other precursors verified the viability and integrity of the isolated hepatocytes. Addition of 10 mM alanine, lactate, and pyruvate produced 5- to 10-fold increases in the rate of gluconeogenesis as compared to the endogenous level in cells from fasted rats. Prolongation of the fast from 24 hr up to 168 hr resulted in no significant changes in the gluconeogenic rates when expressed per gram of cell protein. This finding supports the concept that substrate presentation provides control of hepatic gluconeogenesis during prolonged fasts. The authors gratefully acknowledge the technical assistance of Mrs. Chio-hoon Tan.

Reticuloendothelial system function and glucose-insulin dyshomeostasis in sepsis

Circulating glucose levels, peripheral glucose utilization, and hepatic gluconeogenesis were compared in late endotoxicosis and severe septic shock in rats. Endotoxin was administered intravenously as 5 mg of Salmonella enteritidis lipopolysaccharide B. Sepsis was induced in the peritoneal cavity by use of the cecal ligation and puncture technique. Late endotoxicosis and severe sepsis were comparable in hypoglycemia, increased peripheral glucose use, and depression of gluconeogenesis. Immunoreactive insulin was lower in endotoxicosis than in sepsis; both models demonstrated elevations in serum nonsuppressible insulin-like activity. Endotoxic pancreata secreted excessive insulin, as did pancreata obtained after blockade of the reticuloendothelial system (RES). Macrophage-conditioned media induced a hypersecretory state of the beta cells in donor pancreata. The RES serves as a source of secretory products, i.e., gluco-regulatory monokines, which affects insulinization of tissues in sepsis and thus underwrites the hypoglycemia of late endotoxicosis and severe sepsis.

Effects of exogenous ATP on glucoregulation in vivo.

Summary The effect of exogenous ATP treatment (10 μmoles iv) on glucoregulation was evaluated in male Holtzman rats. ATP treatment decreased lethality of insulin in doses of 1 and 2 U sc, blunted the hypoglycemic response to 0.05, 0.10 and 0.20 U insulin sc, increased the iv glucose disappearance rate, depressed the in vivo oxidation of 400 mg glucose, and inhibited the insulin response of epididymal fat pads to insulin in vitro. The data are consistent with the Chang-Cuatrecasas hypothesis that exogenous ATP inhibits insulin-mediated effects on glucose metabolism via phosphorylation of a membrane component.

Hepatic Detoxification of Endotoxin after Adrenalectomy

Summary Bilateral adrenalectomy of 300±20 g male Holtzman rats resulted in extreme sensitization to endotoxin shock produced by iv Salmonella enteritidis Boivin lipopolysaccharide. Within 60 min after adrenalectomy, resistance to endotoxin as reflected in LD50 values decreased from 800 μg/100 g to 0.59 μg/100 g and by 168 hr after adrenalectomy the LD50 had diminished to 0.25 μg/100 g. Within 15 minutes after adrenalectomy endotoxin resistance had diminished to a LD50 of 50 μg/100 g. Liver homogenates from either acute (60 min) or chronic (168 hr) adrenalectomized and sham-operated control rats were assessed for endotoxin inactivating ability using a lead-sensitized rat bioassay. The ability of liver to detoxify endotoxin was not significantly altered by either acute or chronic adrenalectomy. Endotoxin inactivating ability also was measured in livers removed from either control or chronic adrenalectomized rats in the terminal phase of endotoxin shock; no appreciable loss of hepatic endotoxin inactivating ability was manifested in either group as compared to control or chronic adrenalectomized rats which were not exposed to endotoxin shock. The data suggest that the defect in host resistance to endotoxin imposed by adrenalectomy is probably not due to an impairment of the hepatic intracellular system for endotoxin inactivation.