We purified CD4 and CD8 T cells of SLE patients

and then determined the effect of oestrogen on these cell subsets separately. The result showed that 10−6 M of 17β-oestradiol repressed the PMA plus ionomycin-induced increase in DNA fragmentation in both cell subsets near to basal level (Fig. 1b), indicating that the protective effect of oestrogen on AICD is not different between CD4+ and CD8+ T cells. To address how oestrogen blocked the ACID of T cells, we next investigated whether oestrogen regulated FasL expression in T cells. Flow cytometry analysis revealed that treatment of 17β-oestradiol (10−8 M–10−6 M) decreased FasL selleck chemicals llc protein expression dose-dependently in SLE T cells stimulated with PMA plus ionomycin (Fig. 2a). In contrast, testosterone (10−8 M–10−6 M), a male sex hormone, increased FasL expression additively in these same types of cells (Fig. 2b). Additionally, 17β-oestradiol (10−8 M–10−6 M) abrogated the PMA plus ionomycin-induced up-regulation of FasL mRNA expression in SLE T cells in a dose-dependent manner (Fig. 3a). The Fas mRNA expression in T cells stimulated ACP-196 in vivo with PMA plus ionomycin was decreased similarly by 17β-oestradiol (Fig. 3a). Moreover, 17β-oestradiol also repressed FasL mRNA expression dose-dependently in an hFasL/L5178Y cell line

(kindly provided by Dr J.K. Min, Catholic University of Korea), in which human FasL mRNA was expressed stably (Fig. 3b). To test the specificity of the oestrogen effect, SLE T cells were pretreated with various concentrations (0·5 µM–5 µM) of tamoxifen, an oestrogen receptor antagonist, 1 h before the addition of 17β-oestradiol (10−6 M). As revealed in Fig. 4, tamoxifen cancelled the oestradiol-induced decrease dose-dependently in FasL mRNA expression in T cells stimulated with PMA plus ionomycin, indicating that oestrogen regulates FasL expression through a receptor-coupling event. Based on these data,

we speculated that oestrogen may inhibit the apoptosis of SLE T cells by suppressing FasL up-regulation in the course C1GALT1 of AICD. To address this issue, human FasL-expressing cells (hFasL/L5178Y) were co-cultured with a Fas-expressing cell line (D98AH2 cells, kindly provided by Dr J.H. Lee, Catholic University of Korea) in the presence of 17β-oestradiol. As shown in Fig. 5, 10−6 M of oestradiol inhibited apoptosis of Fas-expressing cells to a similar extent to 10 µg/ml of anti-FasL monoclonal antibody (mAb) treatment. Considering that 17β-oestradiol inhibited FasL expression in the hFasL/L5178Y cell line (Fig. 3b), these data suggest that oestradiol attenuates apoptotic death of Fas-expressing cells by suppressing FasL expression in effector cells.