title: Induction of cytokine tolerance requires internalization of Chylomicron-Bound LPS into hepatocytes.
abstract: BACKGROUND
  Chylomicron-bound LPS (CM-LPS) renders hepatocytes unresponsive to stimulation by proinflammatory cytokines, a process termed cytokine tolerance. We have shown that cytokine tolerance is a time- and dose-dependent process requiring functional low-density lipoprotein receptors (LDLR). Thus, we hypothesized that cytokine tolerance directly correlates with the internalization of CM-LPS complexes, and inhibition of lipoprotein binding and/or internalization inhibits the induction of cytokine tolerance in hepatocytes.
  
  
  METHODS
  We correlated the rate of internalization of radioiodinated CM-LPS complexes with hepatocellular NO production as a measure of cytokine responsiveness. In additional studies, we used four different strategies to inhibit binding/internalization of CM-LPS via LDLR and then determined the effect of each strategy on the induction of cytokine tolerance.
  
  
  RESULTS
  There was a strong inverse correlation between the internalization of CM-LPS and the responsiveness of hepatocytes to proinflammatory cytokines (r(2) = -0.997). Furthermore, the greater the degree of LDLR inhibition, the less susceptible hepatocytes were to the induction of cytokine tolerance by CM-bound LPS. Accordingly, cytokine tolerance induction was inhibited in hepatocytes with decreased membrane expression of LDLR as compared to control cells (69 versus 12% control; P = 0.005). Competitive inhibition of CM-LPS binding prevented internalization of CM-LPS and resulted in loss of the cytokine-tolerant phenotype. Whereas CM-LPS successfully induced cytokine tolerance in ldlr(-/-) hepatocytes, it only occurred after a prolonged pretreatment period of 8 h. CM-LPS complexes containing apolipoprotein (apo) E(2) also required a prolonged pretreatment period to induce a level of cytokine tolerance comparable to that induced by CM-LPS complexes containing either apo E(3) or E(4).
  
  
  CONCLUSIONS
  Lipoprotein-bound LPS inhibits the responsiveness of hepatocytes to proinflammatory cytokines in a manner directly correlated with the internalization of LPS. Furthermore, inhibition of lipoprotein binding/internalization prevents this LPS-mediated induction of cytokine tolerance in rodent hepatocytes.
authors.affiliation: UCSF Surgical Research Laboratory, San Francisco General Hospital, San Francisco, CA 94110, USA.
authors.forename: F Behzad
authors.initials: FB
authors.lastname: Kasravi
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Brecht
authors.forename: Karl H
authors.initials: KH
authors.lastname: Weisgraber
authors.forename: Hobart W
authors.initials: HW
authors.lastname: Harris
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D002914:Chylomicrons / Q000378:metabolism; D016207:Cytokines / Q000276:immunology*; D015536:Down-Regulation / Q000276:immunology; D017353:Gene Deletion; D022781:Hepatocytes / Q000276:immunology / Q000378:metabolism*; D007108:Immune Tolerance; D007457:Iodine Radioisotopes; D008070:Lipopolysaccharides / Q000493:pharmacokinetics*; D008077:Lipoproteins, LDL / Q000378:metabolism; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism; D018805:Sepsis / Q000276:immunology* / Q000378:metabolism*
chemical_list: D002914:Chylomicrons; D016207:Cytokines; D007457:Iodine Radioisotopes; D008070:Lipopolysaccharides; D008077:Lipoproteins, LDL; D011973:Receptors, LDL