Detection of endotoxin in triglyceride-rich lipoproteins in vitro

Abstract

Numerous investigations have been performed in which volunteers have received infusions of triglyceride-rich lipoproteins without apparent screening of the infusates for bacterial endotoxin. This study was designed to examine the capacity of triglyceride-rich lipoproteins to mask their endotoxin content in vitro as measured by a chromogenic modification of the standard Limulus assay. Lipoproteins and lipoprotein-deficient plasma were isolated from normal human plasma by sequential ultracentrifugation under apyrogenic conditions. Individual lipoproteins and a synthetic lipid emulsion were suspended in 10% lipoprotein-deficient plasma. Samples were then incubated at 37 degrees C for 4 hours with increasing concentrations of E. coli (055:B5) endotoxin and assayed for detectable endotoxin activity. The capacity to inhibit detection of endotoxin in 10% lipoprotein-deficient plasma was significantly increased (10 to 100 times) by the addition of VLDL (1.0 mg triglyceride/ml), chylomicrons (1.0 mg triglyceride/ml), or the synthetic lipid emulsion (2.5 mg triglycerides/ml). These data demonstrate that triglyceride-rich lipoproteins, and the synthetic lipid emulsion, can markedly inhibit the detection of endotoxin by the Limulus assay in vitro. In addition to the potential of harm to experimental subjects, infusion of endotoxin could vitiate kinetic studies by direct alteration of lipoprotein metabolism and by inducing changes in hepatic blood flow. Thus experimental protocols that involve the infusion of humans with triglyceride-rich lipoproteins should include detailed testing for the presence of endotoxin.