The axonal repellent, Slit2, inhibits directional migration of circulating neutrophils

Abstract

In inflammatory diseases, circulating neutrophils are recruited to sites of injury. Attractant signals are provided by many different chemotactic molecules, such that blockade of one may not prevent neutrophil recruitment effectively. The Slit family of secreted proteins and their transmembrane receptor, Robo, repel axonal migration during CNS development. Emerging evidence shows that by inhibiting the activation of Rho-family GTPases, Slit2/Robo also inhibit migration of other cell types toward a variety of chemotactic factors in vitro and in vivo. The role of Slit2 in inflammation, however, has been largely unexplored. We isolated primary neutrophils from human peripheral blood and mouse bone marrow and detected Robo-1 expression. Using videomicroscopic live cell tracking, we found that Slit2 selectively impaired directional migration but not random movement of neutrophils toward fMLP. Slit2 also inhibited neutrophil migration toward other chemoattractants, namely C5a and IL-8. Slit2 inhibited neutrophil chemotaxis by preventing chemoattractant-induced actin barbed end formation and cell polarization. Slit2 mediated these effects by suppressing inducible activation of Cdc42 and Rac2 but did not impair activation of other major kinase pathways involved in neutrophil migration. We further tested the effects of Slit2 in vivo using mouse models of peritoneal inflammation induced by sodium periodate, C5a, and MIP-2. In all instances, Slit2 reduced neutrophil recruitment effectively (PϽ0.01). Collectively, these data demonstrate that Slit2 potently inhibits chemotaxis but not random motion of circulating neutrophils and point to Slit2 as a potential new therapeutic for preventing localized inflammation.

Fibroblasts secrete Slit2 to inhibit fibrocyte differentiation and fibrosis

“…Slit2 binds to the Robo1 receptor on monocytes, macrophages, neutrophils, and dendritic cells (35)(36)(37). We observed that Robo1 is expressed on fibrocytes and macrophages and that incubating PBMCs with Slit2 led to reduced fibrocytes, but no obvious changes in Robo1 levels on macrophages (Fig.…”

Regulation of immune cell responses by semaphorins and their receptors

“…1 For example, some axon guidance molecules, such as slits [2][3][4] and ephrins, [5][6][7][8] have been shown to regulate immune cell migration. In addition, T-cell-antigenpresenting cell contact sites, the so-called 'immunological synapse', is structurally similar to the 'neurological synapse' that connects pairs of neurons.…”

The Neurorepellent Slit2 Inhibits Postadhesion Stabilization of Monocytes Tethered to Vascular Endothelial Cells

“…In neuronal cells, Slit2 promotes recruitment of soluble Slit Robo GTPase-activating protein to the cytoplasmic tail of Robo-1, in turn preventing activation of the Rho family GTPases Rac and Cdc42 (20). We (17,21) and other investigators (14-16, 18, 19) showed that secreted Slit2 interacts with Robo-1 on the surface of human PBMC, neutrophils, VSMC, and monocytes to inhibit migration of these cells toward diverse inflammatory chemoattractant cues both in vitro and in vivo. We observed that binding of Slit2 to Robo-1 prevented activation of Cdc42 and Rac, thereby preventing the actin polymerization and cell polarization necessary for directional migration of leukocytes (17).…”

“…Slit and Robo are also expressed in mature organisms, and an isoform of Robo, Robo-1, was detected on the surface of cells involved in vascular injury, including neutrophils, vascular smooth muscle cells (VSMC), lymphocytes, and monocytes (14)(15)(16)(17)(18)(19). In neuronal cells, Slit2 promotes recruitment of soluble Slit Robo GTPase-activating protein to the cytoplasmic tail of Robo-1, in turn preventing activation of the Rho family GTPases Rac and Cdc42 (20).…”