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]-lipopolysaccharide ([H-3]-LPS) binding on capillary endothelium and myocyte cell membranes in Sprague-Dawley rats. Free or serum-containing Ringer-Lock buffer was infused at a rate of 1 ml/min and in the presence of 20 mM K+ and [H-3]-LPS through an aortic cannula, and the effluent was collected through a catheter introduced into the right atrium cavity. The capillary endothelial lining was removed by CHAPS treatment to expose the cardiac myocyte surface. A physical model describing 1:1 binding stoichiometry of [H-3]-LPS with its receptors is proposed and the 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 with the endothelial lining and myocytes, but increased the residency time by 3- and 50-fold, respectively. Hence, the endothelium and myofiber may contain LPS receptors that can bind more strongly with the ligand in association with sCD14-like and LBP-like molecules in rat serum. Thus it is postulated that the affinity of LPS to its receptor subtypes is not strictly and specifically dependent on the CD14 binding profile.