Purpose The expression of CD40 ligand (CD40L) in individuals with systemic lupus erythematosus (SLE) is prolonged when compared to expression on normal T cells. A defect in the down-regulation of the Ras, an intracellular G-protein involved in cell proliferation and differentiation, has been implicated in abnormally prolonged expression of CD 40L on lupus T cells. Inactivation of Ras with farnesylthiosalicylic acid (FTS) in lupus T cells leads to a temporary down-regulation of CD40L expression. SLE is characterized by antinuclear antibodies directed against DNA as well as to individual nucleosides. Guanosine has been demonstrated to be the most immunogenic nucleoside. Serum antiguanosine antibody levels correspond well with disease activity in patients with SLE. Antiguanosine antibodies in murine and human lupus have been shown to have the internal image of the G-binding protein Ras. In this study we investigated whether the murine antiguanosine antibody (4H2) was internalized in living cells and whether it modulated Ras mediated T lymphocytes signal transduction and CD40L expression in normal individuals. This is a continuation of a study that was previously undertaken to look at CD40L expression and the involvement of Ras.
Methods Antiguanosine antibody was isolated from murine hybridoma supernatant and quantified by Bradford Assay. Freshly isolated lymphocytes from healthy controls and SLE patients were obtained with histopaque. Cells were then divided into plus/minus treatment with 4H2 and FTS. The lymphocytes were incubated, fixed, and labeled with anti-CD40L.
Results Our previous studies showed that the expression of CD40L in lupus patients was modulated when Ras was inhibited by FTS treatment but this did not occur in the healthy group.
Conclusion In previous studies, Ras was implicated in the sustained elevation of CD40L on lymphocytes from patients with SLE. In this continuation study 4H2 antiguanosine antibody was used to determine if the Ras-mimicking potential of this internalizing antibody was capable of modulating CD40L levels on healthy lymphocytes. The results thus far show that the addition of FTS did not alter the levels of CD40L on healthy lymphocytes. Whether the 4H2 antiguanosine antibody elevated the levels of CD40L was inconclusive. Currently studies are under way to expand the concentrations of 4H2 tested and to vary the times of incubation.
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