cigarettes induce even greater oxidative stress - as adjudi-cated by cellular HO-1 levels - than standard cigarettes.

doi:10.1016/j.clim.2008.03.123

Autoimmune Neurologic Diseases

F.12. Immune Regulatory Role for CNS-specific,Autoreactive CD8 T Cells in Multiple SclerosisEthan Baughman, Vinodh Pillai, Elliot Frohman, NitinKarandikar. UT Southwestern Med Ctr, Dallas, TX

Multiple sclerosis (MS) is an inflammatory, demyelinatingdisease of the central nervous system (CNS). MS is thought tobe a T-cell-mediated disease, although the major focus ofprior research has been on CD4 T cells. We have shownpreviously that MS patients and healthy subjects harbor CNS-specific CD8 Tcells with a mixed functional profile. The roleof these CD8 Tcells in the pathogenesis and/or regulation ofMS is still unclear. We have also shown that untreated MSpatients show an overall deficit in the immune suppressiveability of CD8 T-cells, which is corrected following glatir-amer acetate (Copaxone) therapy. In the current study, weaddressed the role of CNS-targeted CD8 Tcells in MS patientsby developing a novel, flow cytometry-based in vitroimmune suppression assay. First, we confirmed the validityof this approach by replicating the suppression by CD4+CD25+ regulatory T cells (“T-regs”) from healthy subjects.We observed a deficit of CD4+CD25+ T-reg-mediated sup-pression in untreated MS patients. Moreover, MS patients alsohad an overall deficit in CD8 T-cell-mediated suppression inCopaxone- or anti-CD3-stimulated cultures. Interestingly,CD8 T cells from MS patients and healthy controls subjectsthat were targeted against CNS autoantigens, but not thosespecific for control antigens, exhibited regulatory ability, inthat they suppressed the proliferation of CD4+CD25- T cells.In conclusion, our results not only confirm that MS is a diseaseof perturbed immune regulation but also provide the firstevidence that CNS-specific CD8 Tcells may have a regulatoryrole in this disease. These studies were supported, in part, bygrants from the NIH and the National MS Society.

doi:10.1016/j.clim.2008.03.124

F.13. Inhibition of Autoimmune Response toMultiple Antigens in EAE by a Single RecombinantTCR Ligand (RTL)Sushmita Sinha,1 Gregory Burrows,2 Arthur Vandenbark,1

Halina Offner.1 1Portland VA Medical Center, Portland, OR;2Oregon Health and Science University, Portland, OR

Recombinant T cell receptor ligands (RTLs) can preventand reverse clinical and histological signs of experimentalautoimmune encephalomyelitis (EAE) in an antigen specificmanner, and are currently in clinical trials for treatment ofsubjects with multiple sclerosis (MS). The efficacy of RTLtreatment in MS will depend on whether a single RTL is

sufficient in down modulating autoimmune response tomultiple antigens. To test this, RTLs containing PLP-139-151 (401) and PLP-84-104 (403) peptides were used to treatEAE induced by mixed peptides (PLP139-151, 178-191, 84-104) or syngenic whole spinal cord homogenate (WSCH).Single RTL was capable of reversing clinical and histologicalsigns of EAE given that one of the immunizing peptides waspresent in the RTL molecule used for the treatment. BothRTLs, 401 & 403, were very effective in treating WSCHinduced EAE as well. CNS cells from the WSCH experimentshowed proliferative responses to PLP-139-151 and PLP-43-61, which were significantly reduced with RTL treatment. Invitro, WSCH stimulated splenocytes from the RTL treatedgroup had significantly reduced levels of IL-17, TNF-α, IL-2and increased levels of IL-4, IL-10 and IL-13. We speculatethat increased anti-inflammatory cytokine production, par-ticularly IL-10, by RTL-treated splenocytes inactivatesmacrophages and dendritic cells, leading to reduction inpro-inflammatory cytokines and inhibition of MHC class IIupregulation by these cells and further dampening theongoing autoimmune response. Our data clearly demonstratethat antigen specific anti-inflammatory responses generatedby an RTL molecule are capable of suppressing autoimmuneresponses against multiple epitopes, further supporting theirpotential efficacy as therapy for MS.

doi:10.1016/j.clim.2008.03.125

F.14. Autoantibodies to Myelin OligodendrocyteGlycoprotein in Pediatric MSKatherine McLaughlin,1 Tanuja Chitnis,2 Brenda Banwell,3

Amit Bar-Or,4 Jens Kuhle,5 Ludwig Kappos,5 Kevin Rostasy,6

Daniela Pohl,6 Susan Wong,7 Norma Tavakoli,7 SilviaTenembaum,8 Bettina Franz,1 Kevin O'Connor,2 DavidHafler,2 Kai Wucherpfennig.1 1Dana-Farber Cancer Institute,Boston, MA; 2Brigham and Women's Hospital, Boston, MA;3The Hospital for Sick Children, Toronto, ON, Canada;4Montreal Neurological Institute, Montreal, QC, Canada;5Basel University Hospital, Basel, Switzerland; 6Georg-AugustUniversity, Goettingen, Germany; 7Wadsworth Center,New York State Department of Health, Albany, NY;8Dr. J.P. Garrahan National Pediatric Hospital,Buenos Aires, Argentina.

We previously reported a flow cytometric assay fordetection of antibodies to folded myelin oligodendrocyteglycoprotein (MOG) in patient samples. We have now appliedthis technique to determine the relative frequencies ofautoantibodies to MOG in pediatric and adult-onset multiplesclerosis. Sera were incubated with Jurkat cells expressingfull-length human MOG fused to GFP or GFP alone. Bound IgGwas detected with a biotinylated secondary antibody andstreptavidin-PE. The amount of specific antibody wasexpressed as the ratio of PE fluorescence on the MOG cellsand controls, and a ratio of 5 was considered positive. Todemonstrate specificity, antibodies were purified from seraon beads coated with recombinant MOG or a control protein.Antibodies to MOG were rarely found in adult or pediatriccontrol sera. While the frequency of anti-MOG was low inadult-onset MS (4.5%), 28/131 pediatric MS sera (21.4%)

S47Abstracts