Transplantation and immunologyVitamin E inhibits cyclosporin A and H2O2 promoted epstein–barr virus (EBV) transformation of human B cells as assayed by EBV oncogene LMP1 expression
Introduction
Since its introduction about two decades ago, cyclosporin A (CsA) has became the mainstay of transplant immunosuppression drug because of its potency and selectivity in targeting T cell activation and maturation. One of its side effects is the increasing incidence of post-transplant lymphoproliferative disorder (PTLD),and shortening of the latency period of development of PTLD 1, 2. It is generally believed that the increasing incidence of PTLD is largely the result of suppression of T cell activity by CsA, resulting in uninhibited proliferation of Epstein-Barr virus (EBV)-immortalized B cells 3, 4. Significant emphasis, therefore, has been placed on the impaired immunity for prevention and therapy of PTLD 4, 5, resulting in an increased risk of transplanted organ rejection. However, CsA also exerts a direct oxidative effect on cells by suppression of mitochondria permeability transition pore [6], and this process has been reported in different cell types 7, 8. We have reported that CsA and H2O2 directly promoted EBV immortalization of human peripheral blood B cells in vitro in a concentration-dependent manner and that this promotion of EBV-B cells transformation was inhibited by antioxidants [11]. Subsequently we reported that CsA or H2O2 caused lipid peroxidation in human B cells as assayed by lipid hydroperoxide level in cell extract and that this oxidative process was also blocked by antioxidants and we correlated this effect with an increased colony number and 3H-thymidine incorporation in EBV-infected human B cells, which was again inhibited by antioxidant Vitamin E [12]. LMP-1 is a true oncogene encoded by EBV and has been reported to be essential for transformation of B cells. Therefore, we asked whether direct oxidative stress induced by either CsA or H2O2 will promote LMP-1 expression in EBV-infected B cell population and whether this process will be blocked by dietary antioxidant vitamin E as well. Results of this investigation are presented in this article, confirming our previous reports that oxidative stress plays an important role in EBV-induced B cell transformation and that this process may be mediated via EBV oncogene LMP-1.
Section snippets
Cells and EBV
Human peripheral blood B cells and splenocytes were isolated as described previously 11, 12. The lymphoblastoid cell line LCLD#1 is established in our laboratory by immortalized human peripheral blood B cells with EBV strain B95-8 as described previously [13]. The B95-8 strain of EBV was purchased from American Type Culture Collection (ATCC, Rockville, MD). EBV generation procedures were according to the provider's protocol and their published methods [14].
Culture medium
Standard culture medium of RPMI-1640
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