The production of macrophage inflammatory protein‐2 induced by soluble intercellular adhesion molecule‐1 in mouse astrocytes is mediated by src tyrosine kinases and p42/44 mitogen‐activated protein kinase

Abstract

Severe traumatic brain injury stimulates the release of soluble intercellular adhesion molecule-1 (sICAM-1) into CSF. Studies in cultured mouse astrocytes suggest that sICAM-1 induces the production of macrophage inflammatory protein-2 (MIP-2). In the present study, we investigated the underlying mechanisms for MIP-2 induction. sICAM-1 induced MIP-2 in astrocytes lacking membrane-bound ICAM-1, indicating that its action is due to heterophilic binding to an undescribed receptor rather than homophilic binding to surface ICAM-1. Signal transduction may be mediated by src tyrosine kinases, as the src tyrosine kinase inhibitors herbimycin A and PP2 abolished MIP-2 induction by sICAM-1. Phosphorylation of p42/44 mitogen-activated protein kinase (MAPK), but not of p38 MAPK, occurred further downstream, as evidenced by western blot analysis combined with the use of herbimycin A and specific MAPK inhibitors. By contrast, induction of MIP-2 by tumour necrosis factor-a (TNF-a) involved both p42/44 MAPK and p38 MAPK. Following stimulation with either sICAM-1 or TNF-a, astrocyte supernatants promoted chemotaxis of human neutrophils and incubation of these supernatants with anti-MIP-2 antibodies more efficiently suppressed the migration induced by sICAM-1 than by TNF-a. These results show that sICAM-1 induces the production of biologically active MIP-2 in astrocytes by heterophilic binding to an undefined receptor and activation of src tyrosine kinases and p42/44 MAPK. Keywords: astrocytes, macrophage inflammatory protein-2, p42/44 mitogen-activated protein kinase, soluble intercellular adhesion molecule-1, src tyrosine kinases. Cell surface adhesion molecules play an important role in inflammation and immunosurveillance, mediating extravasation of leucocytes and modulating their effector functions. The migration of leucocytes from the bloodstream into inflamed tissue requires a cascade of events, starting with rolling of leucocytes along the endothelium (Kuby 1997). Upon activation by inflammatory signals such as chemokines, rolling progresses to firmer adhesion mediated by binding of leucocyte integrins to endothelial adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1). The firm adhesion of leucocytes to the endothelium is a prerequisite for their subsequent transendothelial migration. Address correspondence and reprint requests to Vivianne I. Otto, PhD, Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, BRC 417, 975 NE. 10th Street, Oklahoma City, OK 73104, USA. E-mail: vivianne-otto@ouhsc.eduAbbreviations used: fMLP, N-formyl-methionyl-leucyl-phenylalanine; ICAM-1, intercellular adhesion molecule-1; ICAM-1-/-, ICAM-1 knockout; IL, interleukin; LFA-1, leucocyte function associated antigen-1; Mac-1, macrophage antigen-1; MAPK, mitogen-activated protein kinase; MIP, macrophage inflammatory protein; PMN, polymorphonuclear granulocytes; RANTES, regulated upon activation, normal T-cell expressed and secreted; sICAM-1, soluble intercellular adhesion molecule...

ICAM-1: Contribution to Vascular Inflammation and Early Atherosclerosis

“…In static cell culture, sICAM-1 at physiologically relevant concentrations has been reported to induce expression of pro-inflammatory mediators (Otto, Heinzel-Pleines et al 2000;Otto, Gloor et al 2002). mRNA levels of IL-8 and MCP-1 were significantly increased 1hr after exposure to sICAM-1 in HUVECs ( figure 6A and B).…”

“…Two potential candidates for receptors have been suggested so far, namely the homophilic binding to membrane bound ICAM-1, or an unknown receptor of 49kD (Gho, Kleinman et al 1999;Otto, Gloor et al 2002). Otto et al showed that sICAM-1 did not bind to cell surface ICAM-1 using ICAM-1 deficient murine astrocytes but identified the involvement of an unidentified receptor (Otto, Gloor et al 2002).…”

“…Two potential candidates for receptors have been suggested so far, namely the homophilic binding to membrane bound ICAM-1, or an unknown receptor of 49kD (Gho, Kleinman et al 1999;Otto, Gloor et al 2002). Otto et al showed that sICAM-1 did not bind to cell surface ICAM-1 using ICAM-1 deficient murine astrocytes but identified the involvement of an unidentified receptor (Otto, Gloor et al 2002). It is possible that the level of expression of this unknown receptor on the cell surface could vary under different flow conditions, for example the receptor could be regulated through activation of a shear stress response element (SSRE) and therefore result in different expression levels on EC depending on the kinetics and/or type of shear stress to which the cells have been exposed.…”

“…However, a number of studies have also shown that addition of sICAM-1 at a physiologically relevant concentration activates proinflammatory cascades and causes angiogenesis in different in vitro and in vivo models. Incubation of murine brain microvascular EC or astrocytes with sICAM-1 lead to the increased secretion of the proinflammatory chemokine MIP-2 via activation of src tyrosine kinase and Erk phosphorylation (Otto, Heinzel-Pleines et al 2000;Otto, Gloor et al 2002). Gho and colleagues investigated the effects of sICAM-1 on EC angiogenesis.…”

“…Levels of phospho-Erk in HUVEC subjected to flow remained below phospho-Erk in equivalent static cultures for at least a further 24h (data not shown). sICAM-1 has been shown to induce phospho-Erk in mouse astrocytes in static culture (Otto, Gloor et al 2002), we therefore examined the effect of sICAM-1 in HUVEC pre-exposed to 10dyn/cm 2 flow. As seen in Figure 5B in HUVEC pre-exposed to flow for 8hr followed by sICAM-1 incubation caused a transient elevation of phospho-Erk after 15min ( Figure 5B 292±28% of untreated flowed HUVEC p<0.01).…”