Regulation of CCL2 and CCL3 expression in human brain endothelial cells by cytokines and lipopolysaccharide.
BACKGROUND
Chemokines are emerging as important mediators of CNS inflammation capable of activating leukocyte integrins and directing the migration of leukocyte subsets to sites of antigenic challenge. In this study we investigated the expression, release and binding of CCL2 (MCP-1) and CCL3 (MIP-1alpha) in an in vitro model of the human blood-brain barrier.


METHODS
The kinetics of expression and cytokine upregulation and release of the beta-chemokines CCL2 and CCL3 were studied by immunocytochemistry and enzyme-linked immunosorbent assay in primary cultures of human brain microvessel endothelial cells (HBMEC). In addition, the differential binding of these chemokines to the basal and apical endothelial cell surfaces was assessed by immunoelectron microscopy.


RESULTS
Untreated HBMEC synthesize and release low levels of CCL2. CCL3 is minimally expressed, but not released by resting HBMEC. Treatment with TNF-alpha, IL-1beta, LPS and a combination of TNF-alpha and IFN-gamma, but not IFN-gamma alone, significantly upregulated the expression and release of both chemokines in a time-dependent manner. The released CCL2 and CCL3 bound to the apical and basal endothelial surfaces, respectively. This distribution was reversed in cytokine-activated HBMEC resulting in a predominantly basal localization of CCL2 and apical distribution of CCL3.


CONCLUSIONS
Since cerebral endothelial cells are the first resident CNS cells to contact circulating leukocytes, expression, release and presentation of CCL2 and CCL3 on cerebral endothelium suggests an important role for these chemokines in regulating the trafficking of inflammatory cells across the BBB in CNS inflammation.
Chui
Ray
R
Division of Neuropathology, Department of Pathology and Laboratory Medicine, Vancouver General Hospital and the University of British Columbia, Vancouver, BC, Canada.
Dorovini-Zis
Katerina
K
D000328:Adult; D000704:Analysis of Variance; D001921:Brain / Q000166:cytology*; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism*; D054405:Chemokine CCL3 / Q000235:genetics / Q000378:metabolism*; D016207:Cytokines / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008875:Middle Aged; D021381:Protein Transport / Q000187:drug effects; D018709:Statistics, Nonparametric; D013997:Time Factors
D018932:Chemokine CCL2; D054405:Chemokine CCL3; D016207:Cytokines; D008070:Lipopolysaccharides