26 DEFECTIVE FUNCTIONING OF RAS IN LUPUS T CELLS

A host of immunologic abnormalities have been thoroughly described in systemic lupus erythematosus. There is an increasing body of evidence that the production of autoantibodies is the result of abnormal regulation of T cells of the early monitored response to autoantigens. Available data in the literature suggest there is a possible defect in the down-regulation of the Ras pathway in lupus T cells leading to a failure to develop anergy and abnormally prolonged expression of CD40 ligand. Treatment of MRL/lpr mice with the Ras inhibitor S-farnesylthiosalicylic acid (FTS) reduced disease symptoms in the treated mice, indicating that modulation of Ras may represent a novel therapeutic approach in SLE. In this study we are investigating the expression of CD40 ligand in CD40+ T cells from lupus and healthy subjects before and after inactivation of Ras by FTS, a synthetic substance that detaches Ras from the inner cell membrane and induces its rapid degradation. The results of our study will help us understand if the defective functioning of Ras in lupus T cell is an intrinsic defect (by comparing the results with the ones from healthy subjects) versus an acquired one, possibly through autoantibodies. This is a continuation of a study that was previously undertaken to look at CD40 ligand expression.MethodsFreshly isolated lymphocytes from SLE and healthy controls were established in culture and a portion treated with FTS and treated and untreated samples were harvested by fixation at 3 and 6 hours post initiation of treatment (FTS). The fixed lymphocytes were immunolabeled for CD40L, and dual labeled with CD3 and CD20. The secondary fluorescence was quantified by laser scanning cytometry.ResultsThe initial study revealed that the expression of CD40L in lupus patients was modulated when Ras was inhibited by FTS treatment. The down-regulation of CD40L in SLE by FTS treatment occurred at the 4 hour harvest but was refractory at the 48 hour time point. The healthy controls showed no modulation of CD40L expression with or without FTS treatment. The current study used 3 and 6 hours ± FTS treatment and found that at 3 hours post treatment the SLE lymphocytes had down-regulated CD40L but the controls did not. At the 6 hour time point the SLE patient CD40L levels were increasing, whereas the control levels did not change.ConclusionsRas inactivation by FTS was able to markedly decrease the level of CD40L in lupus T cells but did not appear to modulate the level of CD40L i