Lyme disease is the most common vector-borne disease in the United States. It is caused by the spirochete Borrelia burgdorferi and transmitted by lxodes ticks. This renewal application examines the contribution of synovial gamma delta T lymphocytes in Lyme arthritis. Preliminary studies show that gamma delta T cells of the V delta 1 subset accumulate in Lyme arthritis synovial fluid (1 0-15 percent) and proliferate vigorously in response to lipidated but not delipidated Borrelia proteins in the presence of dendritic cells (DC) and IL-2. This may occur through Toll-like receptor 2 (TLR2), which is known to bind Borrelia lipoproteins. The synovial V delta 1 clones recognize the MHC class I-like molecules MICA and CD1b and express high and prolonged levels of surface Fas-ligand (FasL). Finally, we have observed that DC contain very high levels of the Fas death receptor inhibitor, FLIP, and are very resistant to Fas-induced death. In fact, Fas ligation on DC actually promotes upregulation of B7.1, B7.2, and CD4O, similar to TNF alpha. We have identified a signal pathway for this in which high levels of FLIP can divert signals to the MAP kinase, ERK, and NF-KB by binding to adaptor proteins that like to these pathways. In this manner death signals can be switched to signals for cell growth or proliferation. The model emerging from the preliminary studies is that lipoproteins from B. burgdoderi bind to TLR2 on DC to upregulate molecules such as MICA and CD1b that are stimulatory for synovial V delta 1 cells. The V delta 1 cells express high and prolonged surface FasL which is lytic to some synovial components, but may be stimulatory toward DC due to their high expression of FLIP. Each of the three aims studies one aspect of this model.
Aim 1 examines whether B. burgdorferi activates V delta 1 cells directly or indirectly through TLR2. We will examine cells expressing or not expressing TLR2 for their ability to activate V delta 1 clones in the presence of Borrelia proteins, either lipidated or delipidated.
Aim 2 studies two important aspects of synovial V delta 1 clones: how their high expression of surface FasL is regulated, and whether the actual gamma delta TCR is responsible for the response to MICA and CD1b.
Aim 3 will study development of DC from CD34+ precursors using a newly developed in vitro culture technique using Flt3 ligand. This will be paralleled by studies of FLIP expression, the ability of soluble FasL to promote cell death or growth/differentiation, depending on the levels of FLIP. The FasL stimulated DC will be analyzed for surface induction of CD1b, CD4O, B7.l/2, MICA, and their ability to stimulate the V delta 1 clones.
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