Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) of unknown etiology. It is the most common non-traumatic cause of neurologic disability in young adults. Although three drugs have been approved to treat the symptoms of MS, they are not a cure. Development of more effective treatment strategies will be improved by a greater understanding of the pathogenic events in this disease. The pathology of MS is characterized by perivascular infiltration of mononuclear cells. Several lines of evidence suggest that costimulation independent (memory) myelin basic protein (MBP) reactive T cells exist preferentially in MS patients' blood and may be involved in initiating the inflammatory response. T cell activation requires at least two signals: antigen presented in the context of major histocompatibility complex (MHC) class II and a second, costimulatory, signal of binding the CD28 receptor on the T cell. In MS, it is thought that the activated T cell then preferentially produces specific Th1 cytokines that upregulate adhesion molecules on endothelial cells and promote migration into the CNS, where demyelination occurs. Chemokines (CK) are chemoattractant cytokines that mediate tissue specific migration of T cells that express certain chemokine receptors (CKR). In MS, CK and CKR expression are thought to be an important mechanism by which antigen driven T cells migrate to sites of inflammation, but there is little data to support this concept. The investigators will examine CKR expression, function, and correlation with cytokine profiles through the following specific aims:
Specific Aim 1. To determine whether MBP reactive memory T cells express more CCR5 and CXCR3 than MBP reactive naive T cells or TT reactive memory cells: a) using primary proliferation assays to isolate the MBP and TT reactive memory T cells that are costimulation independent (do not require CD28 signal) from MS patients, as compared to ONDs and controls; and b) quantifying CKR expression on these cells (by FACS ) and function (by migration studies).
Specific Aim 2. To evaluate whether memory MBP reactive T cells that express CCR5 and CXCR3 produce more of the Th1 cytokines: a) isolating T cell subsets with specific CKR profiles by FACS and migration analysis; and b) quantifying cytokine production by intracytoplasmic staining (using flow cytometry).
Specific Aim 3. To investigate whether VCAM binding to VLA-4 on T cells alters the expression of CKR on MBP reactive T cells by: co-culturing MBP reactive T cells with VCAM expressing endothelial cells or VCAM-Ig, and analyzing CKR expression by FACS or ELISA. The results from these aims will enhance our understanding of MBP reactive T cells as a model for the pathogenic events in MS.
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