T-cell interaction with ICAM-1/ICAM-2 double-deficient brain endothelium in vitro: the cytoplasmic tail of endothelial ICAM-1 is necessary for transendothelial migration of T cells

Abstract

IntroductionInteraction of T cells with the vascular endothelium is a critical step during T-cell extravasation from blood into tissue in immunosurveillance and inflammation. Cell adhesion molecules (CAMs) expressed on the T-cell and endothelial cell surfaces play essential roles in this intercellular interaction. The intercellular adhesion molecule 1 (ICAM-1) and ICAM-2, members of the immunoglobulin superfamily, and their ␤2-integrin ligand lymphocyte function-associated antigen 1 (LFA-1), expressed on all leukocytes, 1,2 are important for firm attachment of leukocytes to, and their subsequent migration across, the endothelium. 3 A role of endothelial ICAM-1 and ICAM-2 in LFA-1-mediated T-cell adhesion to endothelial cells as well as their transendothelial migration (TEM) has been firmly established by a number of in vitro studies. [4][5][6][7][8] In vivo expression patterns of ICAM-1 and ICAM-2 are distinct but overlapping. Overall tissue distribution of ICAM-2 is more restricted than that of ICAM-1. Both ICAMs are expressed at low levels on most leukocytes. ICAM-2 is constitutively expressed on all vascular endothelium, including high endothelial venules at much higher levels than ICAM-1. 9 ICAM-1 expression is strongly inducible by inflammatory cytokines, whereas ICAM-2 was reported to be down-regulated by inflammatory cytokines. 10 On the basis of the endothelial expression patterns of ICAM-1 and ICAM-2, it has been hypothesized that ICAM-2 mediates leukocyte traffic into noninflamed tissue, that is, lymphocyte recirculation during immunosurveillance, whereas upregulated levels of ICAM-1 may increase leukocyte extravasation at sites of inflammation. 11 Strikingly, ICAM-1-or ICAM-2-deficient mice are both viable and show relatively mild defects in their immune responses. ICAM-1-deficient mice suffer from a moderate leukocytosis while activation and migration of leukocytes to places of inflammation are reduced, resulting in impaired immune and inflammatory responses. 12 On the other hand, they are resistant to septic shock 13 and show attenuated ischemia reperfusion injury 14 owing to reduced leukocyteendothelial interaction. Interestingly, ICAM-2-deficient mice do not show any impairment in lymphocyte homing or development of leukocytes, but rather demonstrated a critical role for ICAM-2 in eosinophil trafficking. 15 Recently, an additional function for human endothelial ICAM-2 in mediating the TEM of dendritic cells (DCs) via interaction with a novel DC-specific ligand DC-specific ICAMgrabbing nonintegrin (DC-SIGN) has been demonstrated. 16 Analysis of the involvement of endothelial ICAM-1 and ICAM-2 in T-cell extravasation in vivo has been complicated even in ICAM-1 Ϫ/Ϫ or ICAM-2 Ϫ/Ϫ mice by the fact that both molecules are also involved in immune cell activation. To delineate the functional involvement of endothelial ICAM-1 versus ICAM-2 in TEM of T lymphocytes in vitro, we have established endothelial cell lines deficient for both ICAM-1 and ICAM-2 from ICAM-1 Ϫ/Ϫ ICAM-2 Ϫ/Ϫ mice. Here we show that the ...

Migration of immature mouse DC across resting endothelium is mediated by ICAM‐2 but independent of β2‐integrins and murine DC‐SIGN homologues

“…as the principal mediator while endothelial ICAM-2 is of minor importance [28,33,34]. These results suggest that ICAM-2 may be an important molecular player in mediating homeostatic extravasation of immature DC under non-inflamed conditions in vivo.…”

“…We analyzed the in vitro migration of immature and mature DC across resting endothelium using murine endothelioma lines that had been established previously [27,28]. The endothelioma line bEnd.5 was derived from wild-type mice and expresses moderate levels of ICAM-1 and high levels of ICAM-2 ( We first analyzed transendothelial migration (TEM) across wild-type endothelium by adding 1Â10 6 immature or mature DC to monolayers of resting bEnd.5 cells seeded ontoTranswell filters.…”

“…To this end we used the anti-CD18 mAb GAME-46 [29], which had formerly been shown to be effective in blocking functional interactions between b 2 -integrins and both, ICAM-1 and ICAM-2 [30,31]. In order to demonstrate functionality of the GAME-46 mAb we analyzed the TEM of T cells, which is largely dependent on endothelial ICAM-1 and ICAM-2 and is known to be mediated by the lymphocytes' b 2 -integrins [28]. Fig.…”

“…Three endothelioma cell lines were used: wild-type bEnd.5, bEndI1/2.1 (ICAM-1 -/ICAM-2 -), bEndI1/ 2.1-ICAM1 (ICAM-1 + /ICAM-2 -) and bEndI1/2.1-ICAM2 (ICAM-1 -/ICAM-2 + ). These cells have been described before [27,28]. CHO and 293T cells were propagated according to American Type Culture Collection standard protocols.…”

Comparison of Immortalized bEnd5 and Primary Mouse Brain Microvascular Endothelial Cells as in vitro Blood–Brain Barrier Models for the Study of T Cell Extravasation

“…Specific knowledge about the molecular mechanisms of immune cell extravasation across the BBB has been provided by a number of in vitro studies that used immortalized brain endothelial cell lines from not only rats and mice-as reviewed by Turowski et al (2005)-but also from humans (Afonso et al, 2007;Bahbouhi et al, 2009). In particular, we have previously used polyoma middle T-oncogen-immortalized mouse brain endothelioma cell lines (bEnd) to show that vascular endothelial cell adhesion molecule (VCAM)-1 is involved in T cell adhesion to, but not in T cell diapedesis across the BBB (Laschinger and Engelhardt, 2000), whereas endothelial intercellular cell adhesion molecule (ICAM)-1 and ICAM-2 are essential for T cell diapedesis across the BBB under static conditions in vitro (Lyck et al, 2003;Reiss et al, 1998;Reiss and Engelhardt, 1999).…”