title: FITC Conjugation Markedly Enhances Hepatic Clearance of N-Formyl Peptides.
abstract: In both septic and aseptic inflammation, N-formyl peptides may enter the circulation and induce a systemic inflammatory response syndrome similar to that observed during septic shock. The inflammatory response is brought about by the binding of N-formyl peptide to formyl peptide receptors (FPRs), specific signaling receptors expressed on myeloid as well as non-myeloid cells involved in the inflammatory process. N-formyl peptides conjugated with fluorochromes, such as fluorescein isothiocyanate (FITC) are increasingly experimentally used to identify tissues involved in inflammation. Hypothesizing that the process of FITC-conjugation may transfer formyl peptide to a ligand that is efficiently cleared from the circulation by the natural powerful hepatic scavenging regime we studied the biodistribution of intravenously administered FITC-fNLPNTL (Fluorescein-isothiocyanate- N-Formyl-Nle-Leu-Phe-Nle-Tyr-Lys) in mice. Our findings can be summarized as follows: i) In contrast to unconjugated fNLPNTL, FITC-fNLPNTL was rapidly taken up in the liver; ii) Mouse and human liver sinusoidal endothelial cells (LSECs) and hepatocytes express formyl peptide receptor 1 (FRP1) on both mRNA (PCR) and protein (Western blot) levels; iii) Immunohistochemistry showed that mouse and human liver sections expressed FRP1 in LSECs and hepatocytes; and iv) Uptake of FITC-fNLPNTL could be largely blocked in mouse and human hepatocytes by surplus-unconjugated fNLPNTL, thereby suggesting that the hepatocytes in both species recognized FITC-fNLPNTL and fNLPNTL as indistinguishable ligands. This was in contrast to the mouse and human LSECs, in which the uptake of FITC-fNLPNTL was mediated by both FRP1 and a scavenger receptor, specifically expressed on LSECs. Based on these results we conclude that a significant proportion of FITC-fNLPNTL is taken up in LSECs via a scavenger receptor naturally expressed in these cells. This calls for great caution when using FITC-fNLPNTL and other chromogen-conjugated formyl peptides as a probe to identify cells in a liver engaged in inflammation. Moreover, our finding emphasizes the role of the liver as an important neutralizer of otherwise strong inflammatory signals such as formyl peptides.
authors: Vascular Biology Research Group, Department of Medical Biology, Faculty of Health Sciences, University of Tromsø, Tromsø, Norway.
authors: Cristina Ionica
authors: CI
authors: Øie
authors: Molecular Inflammation Research Group, Department of Medical Biology, Faculty of Health Sciences, University of Tromsø, Tromsø, Norway.
authors: Igor
authors: I
authors: Snapkov
authors: D'Liver AS, Forskningsparken, Tromsø, Norway.
authors: Kjetil
authors: K
authors: Elvevold
authors: Molecular Inflammation Research Group, Department of Medical Biology, Faculty of Health Sciences, University of Tromsø, Tromsø, Norway.|Childhood Cancer Research Unit, Department of Women's and Children's Health, Karolinska Institutet, Astrid Lindgren Children's Hospital, Stockholm, Sweden.
authors: Baldur
authors: B
authors: Sveinbjørnsson
authors: Vascular Biology Research Group, Department of Medical Biology, Faculty of Health Sciences, University of Tromsø, Tromsø, Norway.
authors: Bård
authors: B
authors: Smedsrød
mesh_terms: D000818:Animals; D002196:Capillaries / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism*; D005452:Fluoresceins / Q000378:metabolism*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism*; D044042:Receptors, Formyl Peptide / Q000378:metabolism*
chemical_list: D005452:Fluoresceins; D009842:Oligopeptides; D044042:Receptors, Formyl Peptide; C061989:F-chemotactic peptide-fluorescein; C021642:F-chemotactic peptide; D016650:Fluorescein-5-isothiocyanate