Watanabe Yoshifumi

Body is always exposed to both endogenous and exogenous oxidative stress, and neutralizes them with antioxidant defense system. Imbalance between oxidant and antioxidant produces reactive oxygen and nitrogen species, causing oxidation of protein, lipid, and DNA damage that lead to aging and various types of diseases such as neurodegenerative, pulmonary, or cardiovascular disease. Therefore, antioxidative agents are important for prevention and treatment of these diseases. In this study, we screened Okinawan plant extract library of 80 species to find nuclear factor erythroid-derived two-like-2 (Nrf2)-dependent antioxidant activity by antioxidant response element (ARE)-lucifearase reporter assay. The three species extracts of Garcinia subelliptica, Ocimum gratissimum, and Plectranthus Ornatus dose-dependently induced mRNA and protein expression of HO1, a downstream antioxidant gene of Nrf2. Furthermore, these extracts induced nuclear translocation of Nrf2 and the reduction of reactive oxygen species (ROS). These results suggested that these plants are natural antioxidant sources of Nrf2 activators.

DNA methylation is an epigenetic modification that regulates gene transcription. DNA methyltransferase 1 (DNMT1) plays an important role in DNA methylation. However, the involvement of DNMT1 and DNA methylation in the pathogenesis of atopic dermatitis (AD) remains unclear. In this study, microarray analysis revealed that peripheral blood mononuclear cells of AD patients with low DNMT1 expression (DNMT1-low) highly expressed dendritic cell (DC) activation-related genes. Also, DNMT1-low AD patients exhibited a higher itch score compared to AD patients with high DNMT1 expression (DNMT1-high). By using an AD-like mouse model induced by the application of Dermatophagoides farinae body ointment, we found that Dnmt1 expression was decreased, while the expression of CeC chemokine receptor type 7 (Ccr7) was upregulated in mouse skin DCs. Furthermore, mice exposed to social defeat stress exhibited Dnmt1 downregulation and Ccr7 upregulation in skin DCs. Additionally, dermatitis and itch-related scratching behavior were exacerbated in AD mice exposed to stress. The relationship between low DNMT1 and itch induction was found in both human AD patients and AD mice. In mouse bone marrowderived DCs, Ccr7 expression was inhibited by 5-aza-2-deoxycytidine, a methylation inhibitor. Furthermore, in mouse skin DCs, methylation of CpG sites in Ccr7 was modified by either AD induction or social defeat stress. Collectively, these findings suggest that social defeat stress exacerbates AD pathology through Dnmt1 downregulation and Ccr7 upregulation in mouse skin DCs. The data also suggest a role of DNMT1 downregulation in the exacerbation of AD pathology.

Liver resident macrophages designated Kupffer cells (KCs) form the largest subpopulation of tissue macrophages. KCs are involved in the pathogenesis of liver inflammation. However, the role of KCs in the systemic inflammation is still elusive. In this study, we examined whether KCs are involved in not only intrahepatic inflammation but also extrahepatic systemic inflammation. Administration of clodronate liposomes resulted in the KC deletion and in the suppression of liver injury in T cell-mediated hepatitis by ConA as a local acute inflammation model, while the treatment did not influence dextran sulfate sodium-(DSS-) induced colitis featured by weight loss, intestinal shrink, and pathological observation as an ectopic local acute inflammation model. In contrast, KC deletion inhibited collagen-induced arthritis as a model of extrahepatic, systemic chronical inflammation. KC deleted mice showed weaker arthritic scores, less joint swelling, and more joint space compared to arthritis-induced control mice. These results strongly suggest that KCs are involved in not only intrahepatic inflammatory response but also systemic (especially) chronic inflammation.

Hepatocyte apoptosis plays a key role in the pathogenesis of immune-mediated hepatitis. However, the detailed mechanisms of apoptosis signaling are still unclear and effective therapeutic drugs for hepatitis have been explored. Here, we show that tryptophan (Trp) suppressed IFN-gamma-mediated hepatic apoptosis in vitro. Trp inhibited the downstream apoptotic events of mitochondria disruption, such as cell death and caspase-3 activation, while it did not influence upstream signaling including STAT1 activation and IRF1 expression. Trp suppressed reactive oxygen species (ROS) generation at the mitochondria. IFN-gamma induced ROS in mitochondria by inhibiting complex I and III, but not II. This ROS generation by IFN-gamma required de novo protein synthesis. Trp showed relatively weak direct scavenging activity but antagonized IFN-gamma against the suppression of complex I. In addition, Trp increased the expression of the Nrf2-dependent antioxidant genes NQO1, HO-1 and GCS in hepatocytes both in vitro and in vivo. Finally, the administration of Trp in an acetaminophen-induced ROS-dependent hepatitis model suppressed the liver injury in vivo. Thus, Trp protects hepatocytes from ROS-dependent cell injury via multiple pathways. This study suggests Trp as a therapeutic antioxidant drug for hepatitis and a regulator for Nrf2-dependent genes.