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* Snyder Institute of Infection Immunity and Inflammation, Department of Physiology and Biophysics, University of Calgary, Calgary, Alberta, Canada;
Department of Medical Cell Biology, Uppsala University, Uppsala, Sweden;
Department of Biomedical Engineering, Genome and Biomedical Sciences Facility, University of California, Davis, CA 95616; and
Institute of Molecular Biotechnology of the Austrian Academy of Science, Vienna, Austria
Mac-1-dependent crawling is a new step in the leukocyte recruitment cascade that follows LFA-1-dependent adhesion and precedes emigration. Neutrophil adhesion via LFA-1 has been shown to induce cytoskeletal reorganization through Vav1-dependent signaling, and the current study investigates the role of Vav1 in the leukocyte recruitment process in vivo with particular attention to the events immediately downstream of LFA-1-dependent adhesion. Intravital and spinning-disk-confocal microscopy was used to investigate intravascular crawling in relation to endothelial junctions in vivo in wild-type and Vav1–/– mice. Adherent wild-type neutrophils almost immediately began crawling perpendicular to blood flow via Mac-1 until they reached an endothelial junction where they often changed direction. This pattern of perpendicular, mechanotactic crawling was recapitulated in vitro when shear was applied. In sharp contrast, the movement of Vav1–/– neutrophils was always in the direction of flow and appeared more passive as if the cells were dragged in the direction of flow in vivo and in vitro. More than 80% of Vav1–/– neutrophils moved independent of Mac-1 and could be detached with LFA-1 Abs. An inability to release the uropod was frequently noted for Vav1–/– neutrophils, leading to greatly elongated tails. The Vav1–/– neutrophils failed to stop or follow junctions and ultimately detached, leading to fewer emigrated neutrophils. The Vav1–/– phenotype resulted in fewer neutrophils recruited in a relevant model of infectious peritonitis. Clearly, Vav1 is critical for the complex interplay between LFA-1 and Mac-1 that underlies the programmed intravascular crawling of neutrophils.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 This work was supported by the Canadian Institutes for Health Research, a CIHR group grant, and the Swedish Research Council (57P-20680, 57x-20675). P.K. is an Alberta Heritage Foundation for Medical Research Scientist and the Snyder Chair in Critical Care Medicine. M.P. is a Canadian Association of Gastroenterology Postdoctoral Fellow.
2 M.P. and B.H. contributed equally.
3 Address correspondence and reprint requests to Dr. Paul Kubes, Immunology Research Group, University of Calgary, 3330 Hospital Drive Northwest, Calgary, Alberta T2N 4N1, Canada. E-mail address: pkubes{at}ucalgary.ca
4 Abbreviations used in this paper: WT, wild type; PGN, peptidoglycan.
5 The online version of this article contains supplemental material.
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