Chemokine transport across human vascular endothelial cells

Leukocyte migration across vascular endothelium is mediated by chemokines that are either synthesized by the endothelium or transferred across the endothelium from the tissue. The mechanism of transfer of two chemokines, CXCL10 (interferon gamma inducible protein [IP]-10) and CCL2 (macrophage chemotactic protein [MCP]-1), was compared across dermal and lung microvessel endothelium and saphenous vein endothelium. The rate of transfer depended on both the type of endothelium and the chemokine. The permeability coefficient (Pe) for CCL2 movement across saphenous vein was twice the value for dermal endothelium and four times that for lung endothelium. In contrast, the Pe value for CXCL10 was lower for saphenous vein endothelium than the other endothelia. The differences in transfer rate between endothelia was not related to variation in paracellular permeability using a paracellular tracer, inulin, and immunoelectron microscopy showed that CXCL10 was transferred from the basal membrane in a vesicular compartment, before distribution to the apical membrane. Although all three endothelia expressed high levels of the receptor for CXCL10 (CXCR3), the transfer was not readily saturable and did not appear to be receptor dependent. After 30 min, the chemokine started to be reinternalized from the apical membrane in clathrin-coated vesicles. The data suggest a model for chemokine transcytosis, with a separate pathway for clearance of the apical surface

Expression of chemokines and their receptors by human brain endothelium: Implications for multiple sclerosis

Leukocyte migration into the CNS is mediated by chemokines, expressed on the surface of brain endothelium. This study investigated the production of chemokines and expression of chemokine receptors by human brain endothelial cells (HBEC), in vitro and in situ in multiple sclerosis tissue. Four chemokines (CCL2, CCL5, CXCL8 and CXCL10), were demonstrated in endothelial cells in situ, which was reflected in the chemokine production by primary HBEC and a brain endothelial cell line, hCEMC/D3. CXCL8 and CCL2 were constitutively released and increased in response to TNF and/or IFN . CXCL10 and CCL5 were undetectable in resting cells but were secreted in response to these cytokines. TNF strongly increased the production of CCL2, CCL5 and CXCL8, while IFN up-regulated CXCL10 exclusively. CCL3 was not secreted by HBECs and appeared to be confined to astrocytes in situ. The chemokine receptors CXCR1 and CXCR3 were expressed by HBEC both in vitro and in situ, and CXCR3 was up-regulated in response to cytokine stimulation in vitro. By contrast, CXCR3 expression was reduced in silent MS lesions. Brain endothelium expresses particularly high levels of CXCL10 and CXCL8, which may account for the predominant TH1-type inflammatory reaction seen in chronic conditions such as multiple sclerosis

CCL2 disrupts the adherens junction: implications for neuroinflammation

Alterations to blood-brain barrier (BBB) adhesion molecules and junctional integrity during neuroinflammation can promote central nervous system (CNS) pathology. The chemokine CCL2 is elevated during CNS inflammation and is associated with endothelial dysfunction. The effects of CCL2 on endothelial adherens junctions (AJs) have not been defined. We demonstrate that CCL2 transiently induces Src-dependent disruption of human brain microvascular endothelial AJ. β-Catenin is phosphorylated and traffics from the AJ to PECAM-1 (platelet endothelial cell adhesion molecule-1), where it is sequestered at the membrane. PECAM-1 is also tyrosine-phosphorylated, an event associated with recruitment of the phosphatase SHP-2 (Src homology 2 domain-containing protein phosphatase) to PECAM-1, β-catenin release from PECAM-1, and reassociation of β-catenin with the AJ. Surface localization of PECAM-1 is increased in response to CCL2. This may enable the endothelium to sustain CCL2-induced alterations in AJ and facilitate recruitment of leukocytes into the CNS. Our novel findings provide a mechanism for CCL2-mediated disruption of endothelial junctions that may contribute to BBB dysfunction and increased leukocyte recruitment in neuroinflammatory diseases