Regulation of immunoproteasome expression by human brain endothelium : functional significance in MHC class I expression and CD8+ T cell adhesion to the cerebral endothelium

Brain inflammatory diseases are characterized by the entry of blood-borne leukocytes into the brain across the blood-brain barrier (BBB) that normally restricts their entry into the central nervous system. Recent studies indicate that infiltrating brain antigen-specific CD8+ T cells clonally expand in multiple sclerosis (MS) lesions. Previous studies from this laboratory showed that cytokines induce human brain microvessel endothelial cells (HBMEC) to express or upregulate several endothelial cell adhesion molecules (ICAM -1, VCAM-1, PECAM-1 and E-selectin), which mediate leukocyte adhesion and transendothelial migration. In addition, cytokine stimulated HBMEC are induced to express class II MHC and co-stimulatory molecules and provide secondary signals for T cell proliferation. In the present study, we investigated the role of the proteasome in the regulation of MHC class I expression by cerebral microvessel endothelial cells and CD8+ T cell adhesion to the BBB endothelium. Primary cultures of HBMEC were treated with: 1) IFN-y to increase the cellular population of the 20S immunoproteasome, a protease that generates MHC class I specific peptides and 2) TNF-a to upregulate adhesion molecules. Interferon-Y induced upregulation of the immunoproteasome subunits (31 i and (35i at the mRNA and the protein levels. The constitutive expression of class IMHC by HBMEC was upregulated after treatment with IFN-Y for 48h. Treatment with the proteasome inhibitor lactacystin downregulated the IFN-y-induced class I upregulation. The minimal adhesion of CD8+ T cells to HBMEC was significantly upregulated by IFN-y treatment of HBMEC and downregulated by both lactacystin and MHC class I blocking antibodies. In addition, both the expression of ICAM-1, VCAM-1 and E-selectin and CD8+ T cell adhesion to HBMEC were upregulated by TNF-a and downregulated by co-incubation with lactacystin. These studies indicate that upregulation of the inducible proteasome subunits in HBMEC by proinflammatory cytokines leads to upregulation of MHC class I expression and antigen non-specific adhesion of CD8+ T cells to cerebral endothelium. Adhesion of CD8+ T cells is mediated by adhesion molecules expressed by HBMEC, which are also downregulated following proteasome inhibition. We therefore suggest that the proteasome plays an important role in the regulation of CD8+ T cell adhesion to the blood-brain barrier endothelium during brain inflammatory diseases.Medicine, Faculty ofPathology and Laboratory Medicine, Department ofGraduat