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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.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.

Original publication

DOI

10.1007/s00109-014-1194-6

Type

Journal article

Journal

Journal of molecular medicine (Berlin, Germany)

Publication Date

11/2014

Volume

92

Pages

1169 - 1178

Addresses

Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.

Keywords

Leukocytes, Cell Membrane, Macrophages, Animals, Mice, Inbred C57BL, Mice, Knockout, Mice, Inflammation, Leukotriene B4, Intercellular Signaling Peptides and Proteins, Receptors, Chemokine, Chemokines, Microscopy, Fluorescence, Flow Cytometry, Chemotaxis, Female, Male, Complement C5a, Chemokine CCL2, Chemokine CCL5, Chemokine CCL3