Journal of Autoimmunity (1999) 13, 383–392 Article No. jaut.1999.0331, available online at http://www.idealibrary.com on Acceleration of Autoimmune Diabetes by Cyclophosphamide is Associated with an Enhanced IFN- Secretion Pathway Vitaly Ablamunits, Francisco Quintana, Tamara Reshef, Dana Elias and Irun R. Cohen Department of Immunology, The Weizmann Institute of Science, 76100 Rehovot, Israel Received 25 May 1999 Accepted 3 September 1999 Key words: Autoimmune diabetes, cyclophosphamide, interferon-gamma Cyclophosphamide (CY), an alkylating cytostatic drug, is known for its ability to accelerate a number of experimental autoimmune diseases including spontaneous diabetes in NOD mice. The mechanism(s) by which CY renders autoreactive lymphocytes more pathogenic is largely unknown, but it has been postulated that the drug preferentially depletes regulatory (suppressor) T cells. It has been suggested that in cell-mediated autoimmune diseases, Th2-like lymphocytes secreting IL-4 and/or IL-10 provide protection, while Th1-like cells secreting IFN- are pathogenic. In this study, we analysed the effects of CY on autoimmune diabetes and cytokines in two mouse models: the spontaneous diabetes of NOD mice and the diabetes induced in C57BL/KsJ mice by multiple injections of low dose streptozotocin (LD-STZ). In both models, CY induced severe lymphopenia and accelerated the progression to hyperglycemia. This was associated with changes in splenic cytokine patterns indicating a shift towards the IFN--secreting phenotype. We provide here evidence that IFN- producers are relatively resistant to depletion by CY and that Th0 clones can be shifted towards Th1. However, direct exposure of T lymphocytes to CY may not be a necessary condition for exacerbation of diabetes; NOD.scid mice treated with CY before adoptive transfer of NOD splenocytes developed diabetes at a higher rate than did controls. Thus, the acceleration of diabetes by CY seems to be a complex event, which includes the relatively high resistance of IFN- producers to the drug, their rapid reconstitution, and a Th1 shift of surviving T cell clones. © 1999 Academic Press Introduction Cyclophosphamide (CY) is a cytostatic drug widely used for the treatment of a number of neoplastic and inflammatory diseases [1]. In various experimental systems, the drug has been demonstrated to affect dramatically immune responses: administration of CY has been reported to potentiate tumour rejection, but to inhibit allograft rejection, oral tolerance and some autoimmune diseases [2–5]. In other autoimmune disease models, such as experimental autoimmune encephalomyelitis (EAE) and insulin-dependent diabetes mellitus (IDDM), CY has been shown to promote susceptibility to the disease [6, 7]. In the non-obese diabetic (NOD) mouse, a spontaneous model of IDDM, the disease may be accelerated by one or two injections of CY at a dose of 200–300 mg/ kg [7, 8]. Since the disease induced by CY can be transferred only into irradiated recipients and hyper- glycemia can be prevented by the administration of spleen cells of young non-diabetic NOD mice, it has been suggested that CY preferentially depletes regulatory (suppressor) cells [8, 9]. The nature of these suppressors is largely unknown. There is growing evidence that Th2-like cells secreting IL-4 and IL-10 provide protection, while pathogenic cells are Th1-like and secrete IFN- [10, 11]. Indeed, it has been demonstrated that administration of CY to NOD increases the numbers of IFN- producers in the islet infiltrate by an order of magnitude and renders these lymphocytes more pathogenic in adoptive transfer into NOD.scid mice [12–14]. In this study, we tested the effects of CY on the development of diabetes in the spontaneous disease in NOD mice and in the disease induced by LD-STZ in KsJ mice. The latter is considered autoimmune since it is associated with insulitis [15], in contrast to diabetes induced by high dose STZ that causes a direct toxic death of islet -cells. Therefore, we were interested to learn whether CY affects LD-STZ-induced diabetes in a way similar to that observed in the spontaneous Correspondence to: Prof. Irun R. Cohen Department of Immunology, The Weizmann Institute of Science, 76100, Rehovot, Israel. Fax: (972)–8–934–4103. E-mail: irun.cohen@weizmann.ac.il 383 0896–8411/99/080383+10 $30.00/0 © 1999 Academic Press