Improving the Results of Heart Transplantation: The Role of NRF-2

Improving the Results of Heart Transplantation: The Role of NRF-2

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By: Hirouki Kawajiri, MD, Arash Ghashghai, H.BSc, Laura Tumiati, H.BSc, Vivek Rao, MD, PhD, FRCS(C), F.A.H.A

Division of Cardiovascular Surgery, Toronto General Hospital, University Health Network, University of Toronto

The prevalence of heart failure has been increasing worldwide.1 Although many treatment approaches have been developed, heart transplantation remains the best option for end-stage heart failure patients. However, donor scarcity is still an unsolved issue, and physicians are sometimes forced to enlarge the donor pool by accepting marginal organs, which include organs from elderly or ill patients. Damage caused to organ tissue when the blood supply is returned after being deprived of oxygen, known as ischemia reperfusion injury, is the major factor complicating heart transplantation.It causes various disease processes including inflammation, apoptosis and acceleration of allograft (organ or tissue transplant from the same species) rejection, or chronic allograft dysfunction. Since marginal organs are maximally prone to ischemia reperfusion injury-mediated damage,2 strategies that reduce ischemia reperfusion injury will significantly improve short- and long-term graft function and survival in these extended criteria cases and may be a potential solution for the donor scarcity issue.

Cardiac allograft vasculopathy (CAV), a form of chronic rejection that narrows the blood vessels of the transplanted heart is the most important cause of long-term morbidity in heart transplantation, in addition to malignancy. CAV is detectable by angiography in 8% of survivors within the first year, in 32% within the first five years, and in 43% within the first eight years after heart transplantation.3 The pathogenesis for the development of CAV involves immune and non-immune mediated mechanisms that lead to endothelial cell injury, followed by intimal hyperplasia, vascular remodeling, and subsequent coronary stenosis.4,5 Although many non-immune pathogenic factors contribute to the development of CAV, ischemia reperfusion injury has been shown to have an important impact.6,7 The use of immunosuppressive drugs, namely cyclosporine, has been also incriminated in the development of CAV.8

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that binds to the antioxidant response elements of genes encoding certain antioxidant enzymes, such as superoxide dismutase, and plays a key role in cellular defense against oxidative stress by enhancing the removal of cytotoxic electrophiles or reactive oxygen species.9 In addition to protection against oxidative stress, recent studies have suggested that Nrf2 responds to pro-inflammatory stimuli and rescues cells andtissues from inflammatory injuries. Several pro-inflammatory cytokines such as tumor necrosis factor-alpha, interleukin (IL)-1, IL-2, IL-6, and IL-12 are overproduced when redox-sensitive nuclear factor-kB (NF-kB) is activated by oxidative stress. Reactive oxygen species generated by these cytokines can further induce NF-kB activation and overproduce these cytokines. This vicious cycle of oxidative stress and overproduction of pro-inflammatory cytokines can be disintegrated by activation of Nrf2.

Since oxidative stress and following inflammation plays a critical role in mediating cellular apoptosis and tissue injury during ischemia reperfusion injury, we hypothesized that Nrf2 acts to protect the donor heart from ischemia reperfusion injury and following CAV in heart transplantation. In an in vitro study, we showed that cyclosporine decreased the Nrf2 nuclear/cytosol ratio, Nrf2 phosphorylation, and superoxide dismutase activity, which were rescued by the Nrf2 inducer, tert-Butylhydroquinone (tBHQ). Using rats, we then showed that cyclosporine exposure impaired endothelial function , which was also rescued by tBHQ. These results suggested that cyclosporine-mediated endothelial dysfunction, as it pertains to CAV, may be related to Nrf2.

We then extended our experiments to Nrf2 knockout mice and our unique murine neck heart transplantation model (Figure 1). We stored the donor heart in cold solution for two hours to induce ischemia reperfusion injury before transplant. The knock out donor heart function assessed by echocardiography after 24 hours was significantly impaired compared with wild-type donor hearts. We have also shown that the Nrf2 inducer sulforaphane prevented donor hearts from ischemia reperfusion injury in wild-type. However, this protective effect was absent in mice.

These results suggested that Nrf2 protects against ischemia reperfusion injury in heart transplantation, and the Nrf2 inducer sulforaphane may represent novel therapy to prevent ischemia reperfusion injury in heart transplantation. Now we are planning a chronic rejection murine heart transplantation study to explore the role of Nrf2 in the development of CAV.

We believe that our results are novel and highlight the protective role of the transcription factor Nrf2 against ischemia reperfusion injury and cardiac allograft vasculopathy in heart transplantation. We continue to investigate methods to help solve clinical issues surrounding heart transplantation and improve both short and long term outcomes.

Figure 1: Murine neck heart transplantation model.

References

1. Barker WH, Mullooly JP, Getchell W. Changing incidence and survival for heart failure in a well-defined older population, 1970-1974 and 1990-1994. Circulation. 2006;113: 799-805.

2. Subramaniam K. Early graft failure after heart transplantation: prevention and treatment. Int Anesthesiol Clin. 2012;50: 202-27.

3. Taylor DO, Edwards LB, Boucek MM, et al. Registry of the International Society for Heart and Lung Transplantation: twenty-third official adult heart transplantation report: 2006. J Heart Lung Transplant. 2006;25:869-79.

4. Ramzy D, Rao V, Brahm J, et al. Cardiac allograft vasculopathy: a review. Can J Surg. 2005;48: 319.

5. Libby P, Tanaka H. The pathogenesis of coronary arteriosclerosis (‘‘chronic rejection’’) in transplanted hearts. Clin Transplant. 1994;8: 313.

6. Murata S, Miniati DN, Kown MH, et al. Superoxide dismutase mimetic M40401 reduces ischemia-reperfusion injury and graft coronary artery disease in rodent cardiac allografts. Transplantation. 2004;78:1166-71.

7. Tanaka M, Mokhtari G, Terry R, et al. Prolonged cold ischemia in rat cardiac allografts promotes ischemia-reperfusion injury and the development of graft coronary artery disease in a linear fashion. J Heart Lung Transplant. 2005;24:1906-14.

8. Jeanmart H, Malo O, Nickner C et al. Comparative study of CSA and FK506 versus newer immunosuppressive drugs MMF and rapamycin on coronary endothelial function in vitro. J Heart Lung Transplant. 2001;20:235.

9. Lee JM, Johnson JA. An important role of Nrf2-ARE pathway in the cellular defense mechanism. J Biochem Mol Biol. 2004;37:139-43.