ENDOTOXIN BINDING ON CAPILLARY ENDOTHELIUM AND MYOCYTE PLASMA-MEMBRANES IN PERFUSED RAT-HEART

This work uses a novel heart-perfusion technique to measure [H-3]-lipo polysaccharide ([H-3]-LPS) binding on capillary endothelium and myocyt e cell membranes in Sprague-Dawley rats. Free or serum-containing Ring er-Lock buffer was infused at a rate of 1 ml/min and in the presence o f 20 mM K+ and [H-3]-LPS through an aortic cannula, and the effluent w as collected through a catheter introduced into the right atrium cavit y. The capillary endothelial lining was removed by CHAPS treatment to expose the cardiac myocyte surface. A physical model describing 1:1 bi nding stoichiometry of [H-3]-LPS with its receptors is proposed and th e mathematical equations derived allow for the calculation of binding constants (k(n)), reversal constants (k(-n)), dissociation constants ( k(d)), and residency time constants (tau). The results showed that the presence of serum in the perfusate, slowed the binding of [H-3]-LPS w ith the endothelial lining and myocytes, but increased the residency t ime by 3- and 50-fold, respectively. Hence, the endothelium and myofib er may contain LPS receptors that can bind more strongly with the liga nd in association with sCD14-like and LBP-like molecules in rat serum. Thus it is postulated that the affinity of LPS to its receptor subtyp es is not strictly and specifically dependent on the CD14 binding prof ile.