Rationale for the Use of Antioxidant Vitamins in Clinical Organ Transplantation

Transplantation of a solid organ from an organ donor has emerged as a treatment option for many diseases that otherwise mean certain death or long-term dependency on life support systems for the patient. The organs are usually obtained from brain-dead donors or from living related or living unrelated donors. Despite the shortage of organs for transplantation, careful attention is paid to the fact that only functionally intact, undamaged organs are used for transplantation. However, even with major improvements in the logistics of organ transplantation, surgical technique, preservation techniques, and preservation solutions, every transplantation starts with an inevitable insult on the graft: ischemia and reperfusion. While ischemia temporarily deprives the organ of oxygen and metabolic substrates and prevents the removal of harmful metabolic by-products, reperfusion of oxygenated blood initiates a complex series of events that have the potential to exacerbate the ischemic tissue damage. A pivotal feature of reperfusion injury is the generation of chemotactic mediators and the infiltration of inflammatory cells: inhibition of leukocyte accumulation and/or adhesion to endothelial cells affords protection from tissue injury, in transplanted and nontransplanted organs alike (1-4). In the first clinical trial ever to successfully use blocking monoclonal antibodies for therapeutic purposes in man, Haug and co-workers (5) demonstrated that administration of antibodies against the intracellular adhesion molecule 1 significantly improved the early function of renal allografts and reduced the incidence of allograft rejection episodes. Role of reactive oxygen species in postischemic transplant injury. Reactive oxygen species (ROS*), which are generated during ischemia and reperfusion, play a key role in the recruitment of inflammatory cells and the subsequent microvascular dysfunction. They exert direct tissue damage(peroxidative attack on fatty acids, denaturation of proteins, DNA damage), stimulate the formation of inflammatory mediators with powerful chemotactic potential, up-regulate adhesion molecules on leukocytes and endothelial cells, and stimulate leukocytes to release a host of proteolytic enzymes and ROS(3, 4, 6, 7). The generation and deleterious action of ROS in transplanted organs have been demonstrated experimentally (8-11) and in clinical organ transplantation(12-19). In most of the clinical studies, serum levels of malondialdehyde (an indirect marker of lipid peroxidation) were increased immediately after reperfusion of the transplanted organs. As a consequence of the ROS generation and action, several studies reported a pronounced consumption of the antioxidant serum capacity in experimental animals (9-11) and in human organ recipients (15, 17, 18). Conversely, boosting the antioxidant defense system by pretreatment with antioxidants and ROS scavengers significantly attenuated ischemia/reperfusion injury in transplanted organs, resulting in extended graft and host survival(8, 11, 20). In 1992, Rabl and co-workers(12) were the first to take these findings into the clinic. In a prospective, placebo-controlled, double-blind study, they pretreated 30 renal allograft recipients with a mixture of antioxidant vitamins. In vitamin-treated patients, lipid peroxidation was significantly reduced after transplantation, and early allograft function was significantly better than that in placebo-treated controls, as assessed by lower creatinine levels and improved creatinine clearance rates (12). Similar results have been reported by Land and co-workers after pretreatment of renal allograft recipients with superoxide dismutase or water-soluble, antioxidant vitamin C (14, 19). How does ischemia / reperfusion injury affect short-and long-term allograft function? It has been suggested that ischemia and reperfusion up-regulate the expression of MHC molecules, particularly class II, thus rendering postischemic allografts more immunogenic (21). Indeed, temporary ischemia was found to significantly up-regulate the expression of MHC class I and class II expression in mouse kidneys, when compared with the contralateral nonischemic kidney (22). It was also found to up-regulate MHC expression in canine lung allografts(23). Increased expression of MHC antigens leads to a more effective T-cell recognition and the appropriate effector responses(24). The cytotoxic T-cell response of the recipient against the transplanted organ could furthermore be enhanced by another reperfusion-induced event: the ROS-mediated inactivation of nitric oxide(25). Nitric oxide has several important roles in the regulation of cardiovascular parameters (26). Among other important properties, it counteracts platelet aggregation and leukocyte adhesion to endothelial cells (27, 28), and also significantly inhibits the development of allospecific cytotoxic lymphocyte response (28). Consequently, inhibition of nitric oxide significantly increases the mitogen-induced proliferation of lymphocytes(29). These mechanisms may explain the mode of action by which superoxide dismutase and/or vitamin C prevents acute rejection episodes in renal allograft recipients (14, 19). However, Land and co-workers observed a significant effect of superoxide dismutase not only on renal allograft function in the early posttransplant period, but also on long-term allograft survival. They ascribed this observation to the ability of superoxide dismutase to affect chronic allograft rejection(14, 30). Chronic rejection of transplants has been defined as progressive functional deterioration, occurring months to years after grafting and associated with characteristic morphologic changes in the vascular tree (31-33). The vascular histopathology results from infiltration of the intima by mononuclear cells, migration and proliferation of smooth muscle cells and fibroblasts from the media into the intima, and subsequent deposition of extracellular matrix(33). The mediators involved in leukocyte recruitment into the vessel wall are similar to the mediators involved in leukocyte adhesion during ischemia reperfusion injury. Likewise, the adhesion molecules expressed over the vascular lesions in chronic allograft rejection conform largely to the adhesion molecules expressed during ischemia/reperfusion injury of rat and human allografts (34). Several inflammatory mediators (and ROS) that are generated in postischemic tissue have been implicated in smooth muscle proliferation(35-38). Yilmaz and co-workers(39) demonstrated that prolonged warm ischemia of rat renal allografts predisposes to allograft loss due to chronic atherosclerosis and vascular obliteration, and Wanders et al. (40) observed in an aortic transplantation model in rats that the duration of transplant ischemia significantly correlated with the severity of transplant arteriosclerosis. Finally, compelling clinical evidence for an impact of ischemia/reperfusion injury on long-term allograft survival has been obtained in a recent study that retrospectively correlated the histomorphological severity of ischemia/reperfusion injury in early endomyocardial biopsies with the long-term graft survival. In this study, the histomorphological degree of peritransplant myocardial damage (attributed to ischemia/reperfusion injury) emerged as the strongest predictor of the later development of accelerated transplant atherosclerosis (41). These and other considerations have been proposed to explain the protective effect of ROS inhibition on long-term allograft function (30). Should antioxidant vitamins be included in the dietary / pharmacological regimen of transplant recipients? As mentioned above, antioxidant vitamins have been used both in experimental models and in clinical renal transplantation to effectively inhibit the severity of reperfusion injury and to improve short-term allograft function and survival(8-11, 19). Unfortunately, no study has focused on the long-term outcome of allograft transplantation and the potential impact of antioxidant vitamins on allograft atherosclerosis. It is known from experimental models, as well as from data obtained from human allograft recipients, that antioxidant vitamins are consumed during reperfusion injury of transplanted grafts(8, 9, 15, 17, 18), and that the lipid peroxidation observed after reperfusion can be effectively prevented with antioxidant vitamins, both in animal models (10, 11) and in patients (12, 19). It is furthermore known from animal studies that antioxidant vitamins significantly improve early allograft function and both graft and animal survival(10, 11, 20). Despite ample knowledge about these interrelations, little attention is paid to the antioxidant status in patients undergoing organ transplantation. This is dramatically emphasized by recent reports on antioxidant vitamin levels in liver allograft recipients. Even before transplantation, the circulating levels of glutathione, antioxidant vitamins A and E, carotene, and lycopene were significantly lower in these patients than in normal controls. In some patients, carotene and lycopene levels were so low that they could not be measured(15, 17). These data are particularly problematic in light of the availability and safety of antioxidant vitamin supplements. For example, vitamin C (ascorbate) and vitamin E (tocopherol) afford effective protection from ROS in vivo (7). Their efficacy in humans has been documented in a growing number of reports on clinical trials and epidemiological surveys demonstrating significant protection from coronary artery disease (42, 43). Both of these major antioxidant vitamins have an excellent safety record in doses that far exceed the doses required to reach adequate antioxidant protection in vivo(44). The water-soluble vitamin C inactivates aqueous peroxyl radicals and other ROS and thus protects from lipid peroxidation(45), and the lipid-soluble vitamin E reacts with lipid peroxyl radicals, interrupts the radical chain reaction, and thus prevents further lipid peroxidation. Although theoretically both vitamins C and E can function as pro-oxidants, the widespread use in diverse clinical conditions, including ischemia/reperfusion situations (46, 47), has demonstrated that these mechanisms are either irrelevant in vivo or are by far outweighed by the protective effect of the antioxidant action. For example, 500 mg of vitamin C has been safely injected intravenously into renal allograft recipients immediately before reperfusion of the renal graft, resulting in a significant protection from lipid peroxidation(19). The only limitation to the use in transplant recipients may be vitamin K deficiency coagulopathy, which may be exacerbated by high doses of vitamin E (44). Based on these considerations, we believe that clinical studies are warranted to thoroughly test the impact of antioxidant vitamins in transplant recipients on parameters of short-term and long-term function and survival after clinical organ transplantation.