title: Contrasting in vitro vs. in vivo effects of a cell membrane-specific CC-chemokine binding protein on macrophage chemotaxis.
abstract: Chemokines (CK) provide directional cues that mediate the recruitment of leukocytes to sites of inflammation. Broad-spectrum blockade of the CC-CK family, using the vaccinia virus 35K protein, has been shown to cause a potent reduction of systemic inflammation in models of atherosclerosis, vein graft disease and arthritis. We have used a cell membrane-targeted form of 35K, Mem35K, to probe whether cell-associated blockade of chemokine response is sufficient to reduce cell recruitment in inflammation. In Tie2cre mice, activation of a flox-stop Mem35K transgene directed conditional expression of Mem35K in leukocytes and endothelial cells, confirmed by Western blotting, flow cytometry and immunofluorescence microscopy. This conditional Mem35K expression was sufficient to increase cell surface CCL5 binding and reduce chemotaxis in vitro to CCL5, CCL2 and CCL3 but not to non-CC-CK chemoattractants, LTB4, C5a or chemerin. However, in vivo monocyte recruitment into the peritoneum driven by zymosan or CC-chemokine injection, which was demonstrated to be CC-CK dependent using CCR2-/- mice, was not reduced by Mem35K expression, despite the expression of functional Mem35K protein. These findings highlight differing requirements for cell-associated anti-inflammatory activity in in vitro and in vivo models.
  
  
  CONCLUSIONS
  Mem35K is a cell-associated CC-chemokine binding protein. Conditional Mem35K transgenic mice show expression Mem35K in leukocytes. Mem35K blocks in vitro primary macrophage chemotaxis specifically towards CC-chemokines. Mem35K expression is not sufficient to reduce inflammation in vivo. The requirements for anti-inflammatory activity in vitro and in vivo are different.
authors: Division of Cardiovascular Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK.|Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
authors: Eileen
authors: E
authors: McNeill
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
authors: Asif J
authors: AJ
authors: Iqbal
authors: Division of Cardiovascular Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK.|Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
authors: Jyoti
authors: J
authors: Patel
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
authors: Gemma E
authors: GE
authors: White
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
authors: Daniel
authors: D
authors: Regan-Komito
authors: Sir William Dunn School of Pathology, University of Oxford, Oxford, UK.
authors: David R
authors: DR
authors: Greaves
authors: Division of Cardiovascular Medicine, John Radcliffe Hospital, University of Oxford, Oxford, OX3 9DU, UK. keith.channon@cardiov.ox.ac.uk.|Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK. keith.channon@cardiov.ox.ac.uk.
authors: Keith M
authors: KM
authors: Channon
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D018932:Chemokine CCL2 / Q000378:metabolism; D054405:Chemokine CCL3 / Q000378:metabolism; D018946:Chemokine CCL5 / Q000378:metabolism; D018925:Chemokines / Q000378:metabolism*; D002633:Chemotaxis; D015936:Complement C5a / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D007249:Inflammation; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D007962:Leukocytes / Q000166:cytology; D007975:Leukotriene B4 / Q000378:metabolism; D008264:Macrophages / Q000166:cytology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D019707:Receptors, Chemokine / Q000378:metabolism
chemical_list: C491768:Ccl2 protein, mouse; C496534:Ccl3 protein, mouse; C503460:Ccl5 protein, mouse; D018932:Chemokine CCL2; D054405:Chemokine CCL3; D018946:Chemokine CCL5; D018925:Chemokines; D036341:Intercellular Signaling Peptides and Proteins; D019707:Receptors, Chemokine; C523686:chemerin protein, mouse; D007975:Leukotriene B4; D015936:Complement C5a
keywords: Chemokine; Inflammation; Macrophage; Therapy; Transgenic