T. KAWAMURA1,2, T. MOMOZANE1,2, S. FUNAKI1, Y. SHINTANI1, M. INOUE1, M. MINAMI1, K. SUGIMURA2, O. IIDA2, H. FUCHINO2, N. KAWAHARA2, H. TAKEMORI2, M. OKUMURA1 1Osaka University Graduate School Of Medicine,General Thoracic Surgery,Suita, OSAKA, Japan 2National Institute Of Biomedical Innovation,Ibaraki, OSAKA, Japan
Introduction: Oxidative stress is thought to be involved in various disease states related to organ transplantation such as ischemia/reperfusion injury and its abrogation in affected patients is critical. Carnosol is a plant-derived phenol that functions as an antioxidant and its mechanism of action involves activation of nuclear factor E2 p45-related factor 2 (Nrf2), which plays a key role in regulation of genes driven by antioxidant response element (ARE). Although it is considered to have potential for treatment of oxidative stress, disadvantages include instability in water and high cost, making it difficult to employ in clinical practice. In the present study, we investigated the lung protective effects of carnosol, extracted from Callicarpa longissima, and its derivatives in vivo using a warm lung ischemia model and in vitro with a lung cell line.
Methods: C57BL/6J mice received 0.01% carnosol chow for 2 weeks, after which the left lungs were excised and cultured in DMEM at 37˚C. The concentration of lactate dehydrogenase (LDH) in medium was measured as a marker of lung damage. Following pre-treatment with carnosol, we assessed the expression of heme oxygenase (HO)-1 and H2O2 treatment tolerance in a lung cell line (NCI-H1975), and also evaluated activation of Nrf2 using a cell-based ARE-luciferase reporter assay. Furthermore, we investigated the stability of carnosol derivatives (methylated-, acetylated-, propionylated-carnosol) in a water solution.
Results: In the excised ischemic lungs, carnosol was shown to increase the amount of HO-1 protein and delayed the leakage of LDH (Fig. A). In NCI-H1975 cell cultures, pretreatment with carnosol induced HO-1, increased the amount of intracellular Nrf2, and protected against H2O2 treatment (Fig. B). Carnosol also increased ARE-luciferase activity in a dose-dependent manner. In addition, our results showed that carnosol induced HO-1 to a much greater degree than the same concentration of tBHQ, a representative Nrf2 inducer (Fig. C). Together, these findings suggest that carnosol induces HO-1 via multiple signaling pathways including Nrf2. Furthermore, carnosol derivatives were stable in a water solution and induced HO-1 up to 1 week after preparation.
Conclusion: Although additional investigation is required, carnosol and its derivatives can significantly reduce organ and cell damage by inducing HO-1, and may have potential as novel organ protective agents.
