Peroxynitrite (ONOO(-)), a toxic product of the free radicals nitric oxide and superoxide, has been implicated in the pathogenesis of CNS inflammatory diseases, including multiple sclerosis and its animal correlate experimental autoimmune encephalomyelitis (EAE). In this study we have assessed the mode of action of uric acid (UA), a purine metabolite and ONOO(-) scavenger, in the treatment of EAE. We show that if administered to mice before the onset of clinical EAE, UA interferes with the invasion of inflammatory cells into the CNS and prevents development of the disease. In mice with active EAE, exogenously administered UA penetrates the already compromised blood-CNS barrier, blocks ONOO(-)-mediated tyrosine nitration and apoptotic cell death in areas of inflammation in spinal cord tissues and promotes recovery of the animals. Moreover, UA treatment suppresses the enhanced blood-CNS barrier permeability characteristic of EAE. We postulate that UA acts at two levels in EAE: 1) by protecting the integrity of the blood-CNS barrier from ONOO(-)-induced permeability changes such that cell invasion and the resulting pathology is minimized; and 2) through a compromised blood-CNS barrier, by scavenging the ONOO(-) directly responsible for CNS tissue damage and death.

The loss of blood-brain barrier (BBB) integrity in CNS inflammatory responses triggered by infection and autoimmunity has generally been associated with the development of neurological signs. In the present study, we demonstrate that the clearance of the attenuated rabies virus CVS-F3 from the CNS is an exception; increased BBB permeability and CNS inflammation occurs in the absence of neurological sequelae. We speculate that regionalization of the CNS inflammatory response contributes to its lack of pathogenicity. Despite virus replication and the expression of several chemokines and IL-6 in both regions being similar, the up-regulation of MIP-1β, TNF-α, IFN-γ, and ICAM-1 and the loss of BBB integrity was more extensive in the cerebellum than in the cerebral cortex. The accumulation of CD4- and CD19-positive cells was higher in the cerebellum than the cerebral cortex. Elevated CD19 levels were paralleled by κ-L chain expression levels. The timing of BBB permeability changes, κ-L chain expression in CNS tissues, and Ab production in the periphery suggest that the in situ production of virus-neutralizing Ab may be more important in virus clearance than the infiltration of circulating Ab. The data indicate that, with the possible exception of CD8 T cells, the effectors of rabies virus clearance are more commonly targeted to the cerebellum. This is likely the result of differences in the capacity of the tissues of the cerebellum and cerebral cortex to mediate the events required for BBB permeability changes and cell invasion during virus infection.

The costimulatory requirements required for peripheral blood T regulatory cells (Tregs) are unclear. Using cell-based artificial APCs we found that CD28 but not ICOS, OX40, 4-1BB, CD27, or CD40 ligand costimulation maintained high levels of Foxp3 expression and in vitro suppressive function. Only CD28 costimulation in the presence of rapamycin consistently generated Tregs that consistently suppressed xenogeneic graft-vs-host disease in immunodeficient mice. Restimulation of Tregs after 8–12 days of culture with CD28 costimulation in the presence of rapamycin resulted in >1000-fold expansion of Tregs in <3 wk. Next, we determined whether other costimulatory pathways could augment the replicative potential of CD28-costimulated Tregs. We observed that while OX40 costimulation augmented the proliferative capacity of CD28-costimulated Tregs, Foxp3 expression and suppressive function were diminished. These studies indicate that the costimulatory requirements for expanding Tregs differ from those for T effector cells and, furthermore, they extend findings from mouse Tregs to demonstrate that human postthymic Tregs require CD28 costimulation to expand and maintain potent suppressive function in vivo.

No abstract