Effect of pentoxifylline on nitrogen balance and 3-methylhistidine excretion in parenterally fed endotoxemic rats.

Pentoxifylline interrupts early gene activation for tumor necrosis factor, interleukin-1, and interleukin-6 production and improves survival from experimental sepsis. These effects can alter nitrogen loss during critical illness. To determine the dose-dependent influence of pentoxifylline on nitrogen loss, 44 male Sprague-Dawley rats (220 to 265 g) were randomized to receive parenteral nutrition only (PN), PN plus continuous infusion of Escherichia coli 026:B6 lipopolysaccharide (LPS) at 9 mg x kg(-1) x d(-1), or PN plus LPS plus a continuous infusion of pentoxifylline at either 25 (PEN25) or 100 mg x kg(-1) x d(-1) (PEN100) for 48 h. Before randomization, all animals underwent intravenous cannulation and 40 h of PN adaptation. All animals received isocaloric, isonitrogenous PN (160 kcal x kg(-1) x d(-1) and 1.0 gN x kg(-1) x d(-1)) and were kept nil per os except for water ad libitum. Administration of LPS significantly worsened nitrogen balance for all three groups compared with PN control; however, pentoxifylline only modestly improved nitrogen balance compared with LPS (206 +/- 255, -497 +/- 331, -332 +/- 329, and -310 +/- 383 mg/48hr for the PN, LPS, PEN25, and PEN100 groups, respectively; P < 0.001). Pentoxifylline did not significantly change 3-methylhistidine urinary excretion compared with LPS (573 +/- 180, 705 +/- 156, 780 +/- 326, and 683 +/- 266 microg/48 h for the PN, LPS, PEN25, and PEN100 groups, respectively, P not significant). Pentoxifylline, given in therapeutic doses after an endotoxin challenge, modestly, but not significantly, improved nitrogen balance. Urinary 3-methylhistidine excretion was not influenced by pentoxifylline. A dose-dependent effect by pentoxifylline on these markers was not evident.

Alterations in N-acetylation of 3-methylhistidine in endotoxemic parenterally fed rats.

N-methylhistidine (3-meH) is endogenously released during muscle catabolism and serves as a marker of protein turnover. In rats > 85% of 3-meH is excreted in the urine as the N-acetyl derivative. It has been reported that the percent of non-acetylated 3-meH (NA-3-meH) varies minimally with stress. To further evaluate these reports we randomized 39 male Sprague-Dawley rats (157-213 g) to receive parenteral nutrition only (PN) or PN plus continuous infusion of Escherichia coli 026:B6 lipopolysaccharide at 6 (LPS-6) or 12 (LPS-12) mg.kg-1.d-1 for 48 h. All animals received isocaloric and isonitrogenous PN 24 h before and throughout the study with water ad libitum. Total 3-meH excretion was significantly increased (P < 0.05) in the LPS-6 (470 +/- 136 micrograms/48 h) and LPS-12 (557 +/- 171 micrograms/48 h) groups versus the PN (331 +/- 126 micrograms/48 h) group. NA-3-meH differed significantly between the LPS-12 (218 /+- 89 micrograms/48 h, LPS-6 (94 +/- 48 micrograms/48 h), and PN (39 +/- 12 micrograms/48 h) groups (P < 0.05). Percent NA-3-meH increased significantly from 12.7 +/- 3.9% in the PN group to 19.8 +/- 8.0 and 39.9 +/- 12.8% in the LPS-6 and LPS-12 groups, respectively (P < 0.05). No significant changes in acetyl 3-meH were found between groups. These data suggest that either saturation or inhibition of acetylation pathways occurs with increasing levels of stress. Due to the disproportionate increases in NA-3-meH and percent NA-3-meH during endotoxemia, only total 3-meH should be used as an indicator of protein turnover in rats.

The effect of alpha-adrenergic antagonism upon nitrogen loss during endotoxemia.

Sixty male Sprague-Dawley rats were randomized to receive parenteral nutrition (PN) only; PN plus continuous infusion of Escherichia coli 026:B6 lipopolysaccharide (PN + LPS) at 6 mg.kg-1.d-1; or PN plus LPS plus a continuous infusion of the alpha-adrenergic antagonist phentolamine (PN + LPS + PHEN) at 5 mg.kg-1.d-1 or 20 mg.kg-1.d-1 for 48 h. All animals received isocaloric, isonitrogenous PN. LPS significantly lowered nitrogen balance (mmol/48 h) from PN control; however, addition of PHEN substantially worsened nitrogen balance compared with LPS (14.2 +/- 3, 2.4 +/- 5.2, -1.6 +/- 4.5, -0.8 +/- 5.4, for the PN, PN + LPS, PN + LPS + PHEN5 and PN + LPS + PHEN20 groups, respectively; P < 0.0001). Urinary 3-methylhistidine/creatinine ratio (3-meH/creat) paralleled the nitrogen balance data (0.30 +/- 0.09, 0.45 +/- 0.12, 0.51 +/- 0.14, 0.60 +/- 0.12, respectively; P < 0.0001). The high-dose PHEN resulted in 82 +/- 9% blockade. To ascertain if any beneficial effect upon body protein loss is achieved during severe stress, 30 rats were given PN + LPS at 12 mg.kg-1.d-1 or PN + LPS12 + PHEN20. These data showed similar changes in nitrogen balance and 3-methylhistidine/creatinine with the use of PHEN during severe endotoxemia. alpha-adrenergic antagonism with PHEN worsens body protein loss as measured by nitrogen balance and 3-methylhistidine/creatinine in PN-fed endotoxemic rats.

Octreotide and potassium homeostasis.

Somatostatin infusion causes hyperkalemia in healthy subjects and in some animal models. The purpose of this investigation was to determine what effect octreotide has on potassium homeostasis during serious illness and if there is a dose-response relationship. Sixty-six male Sprague-Dawley rats (185-225 g) were randomized to receive parenteral nutrition (PN) only, PN plus continuous infusion of Escherichia coli lipopolysaccharide (LPS), or PN plus LPS plus octreotide 10, 100, or 1000 micrograms/kg/day for 48 hours. Before randomization all animals received isocaloric, isonitrogenous, isokalemic PN. A 24-hour urine was collected and a blood sample was taken at the end of the study immediately before euthanization. Data were analyzed by ANOVA and Duncan's multiple range test. Nonhemolyzed serum samples from 50 rats were available for study. Serum potassium concentrations were in the normal range for rats and did not differ significantly among the groups: 5.97 +/- 0.86, 5.96 +/- 1.58, 5.78 +/- 1.48, 5.79 +/- 1.67, 5.35 +/- 0.78 mEq/L, respectively. No differences among groups were found for fractional excretion of potassium or serum creatinine concentration. Octreotide administration in escalating dosages does not cause hyperkalemia in endotoxemic rats given intravenous potassium at a constant rate by PN.

Dose-dependent effect of octreotide on nitrogen retention and glucose homeostasis in response to endotoxemia in parenterally fed rats.

OBJECTIVE: This study compared the effect of different doses of octreotide on glucose and protein homeostasis in rats receiving concomitant lipopolysaccharide and parenteral nutrition infusions. METHODS: Sixty-six male Sprague Dawley rats (185 to 220 g) were randomized to receive parenteral nutrition only (PN), PN plus continuous infusion of Escherichia coli 026:B6 lipopolysaccharide at 6 mg/kg/day (LPS), PN plus LPS plus octreotide at 10 micrograms/kg/day (LPS + Oct 10), 100 micrograms/kg/day (LPS + Oct 100), or 1000 micrograms/kg/day (LPS + Oct 1000) for 48 hours. Prior to randomization all animals received isocaloric and isonitrogenous PN (170 kcal/kg/day as glucose and 1.1 g N/kg/day) and were kept nil per os except for water ad libitum. Nitrogen balance, urinary 3-methylhistidine/creatinine ratio, serum glucose concentration, and incidence of glycosuria were compared between groups. Serum urea nitrogen (SUN) changes were incorporated into the cumulative 48 hour nitrogen balance. ANOVA, Duncan's multiple range test, and Fisher's Exact Test were used for statistical analysis. RESULTS: Nitrogen balance (mg/48 hours) was significantly lower in all four groups receiving LPS +/- Oct when compared to the control group receiving PN alone. SUN (mg/dL) was significantly higher in all four groups receiving LPS +/- Oct when compared to control. There were no statistically significant differences in nitrogen balance or SUN among the four groups receiving LPS +/- Oct. The ratio of urinary 3-methylhistidine/ creatinine was significantly higher in the LPS + Oct 1000 group compared to the PN group (0.77 +/- 0.37 vs. 0.42 +/- 0.24, p < 0.05). Serum glucose concentrations and incidence of glycosuria among the five groups were not significantly different. CONCLUSIONS: Endotoxin significantly reduces nitrogen balance compared to controls fed PN. Octreotide does not significantly improve nitrogen retention or glucose homeostasis in endotoxemic parenterally fed rats.

Standardization of acid hydrolysis procedure for urinary 3-methylhistidine determination by high-performance liquid chromatography.

The N-acetylated form of N-methylhistidine (3-methylhistidine, 3-meH), a non-invasive marker of proteolysis, accounts for 80-90% of total 3-meH excretion (acetylated+non-acetylated 3-meH) in the rat. To determine total 3-meH excretion, samples require acid hydrolysis prior to determination by high-performance liquid chromatography. This study evaluated the stability of 3-meH at various times and temperatures of hydrolysis and determined the optimal conditions for hydrolysis of samples. Increasing temperature (120 degrees C) results in significant degradation of 3-meH with no appreciable change in concentration being noted at 80 degrees C. Hydrolysis at 100 degrees C for 1.5 to 4 h or 80 degrees C for 8 to 12 h is recommended for determining total 3-meH concentrations in rat urine.