Fluoride induces oxidative damage and SIRT1/autophagy through ROS-mediated JNK signaling

Abstract

Fluoride is an effective caries prophylactic, but at high doses can also be an environmental health hazard. Acute or chronic exposure to high fluoride doses can result in dental enamel and skeletal and soft tissue fluorosis. Dental fluorosis is manifested as mottled, discolored, porous enamel that is susceptible to dental caries. Fluoride induces cell stress, including endoplasmic reticulum stress and oxidative stress, which leads to impairment of ameloblasts responsible for dental enamel formation. Recently we reported that fluoride activates SIRT1 and autophagy as an adaptive response to protect cells from stress. However, it still remains unclear how SIRT1/autophagy is regulated in dental fluorosis. In this study, we demonstrate that fluoride exposure generates reactive oxygen species (ROS) and the resulting oxidative damage is counteracted by SIRT1/autophagy induction through c-Jun N-terminal kinase (JNK) signaling in ameloblasts. In the mouse-ameloblast-derived cell line LS8, fluoride induced ROS, mitochondrial damage including cytochrome-c release, up-regulation of UCP2, attenuation of ATP synthesis, and H2AX phosphorylation (γH2AX), which is a marker of DNA damage. We evaluated the effects of the ROS inhibitor N-acetylcysteine (NAC) and the JNK inhibitor SP600125 on fluoride-induced SIRT1/autophagy activation. NAC decreased fluoride-induced ROS generation and attenuated JNK and c-Jun phosphorylation. NAC decreased SIRT1 phosphorylation and formation of the autophagy marker LC3II, which resulted in an increase in the apoptosis mediators γH2AX and cleaved/activated caspase-3. SP600125 attenuated fluoride-induced SIRT1 phosphorylation, indicating that fluoride activates SIRT1/autophagy via the ROS-mediated JNK pathway. In enamel organs from rats or mice treated with 50, 100, or 125 ppm fluoride for 6 weeks, cytochrome-c release and the DNA damage markers 8-oxoguanine, p-ATM, and γH2AX were increased compared to those in controls (0 ppm fluoride). These results suggest that fluoride-induced ROS generation causes mitochondrial damage and DNA damage, which may lead to impairment of ameloblast function. To counteract this impairment, SIRT1/autophagy is induced via JNK signaling to protect cells/ameloblasts from fluoride-induced oxidative damage that may cause dental fluorosis.

Cardiac‐specific Mst1 deficiency inhibits ROS‐mediated JNK signalling to alleviate Ang II‐induced cardiomyocyte apoptosis

“…However, Maiko Suzuki and colleagues reported that ROS also promotes antioxidant defense and plays a protective role in the posttranslational modification of proteins; NAC also induced apoptosis via mitochondrial‐dependent means . Therefore, the roles and underlying mechanisms of intracellular ROS and NAC in various cell types remain elusive . In this study, Ang II administration triggered the production of intracellular ROS, leading to cardiomyocyte apoptosis and cardiac dysfunction; this effect was abrogated by NAC and by cardiomyocyte‐specific Mst1 knockout, which resulted in the downregulation of p47 phox , an NAD(P)H oxidase subunit.…”

“…Some previous studies suggested that ROS induced cell apoptosis and NAC restored the ROS‐mediated apoptosis . However, Maiko Suzuki and colleagues reported that ROS also promotes antioxidant defense and plays a protective role in the posttranslational modification of proteins; NAC also induced apoptosis via mitochondrial‐dependent means . Therefore, the roles and underlying mechanisms of intracellular ROS and NAC in various cell types remain elusive .…”

Reactive Oxygen Species-Mediated c-Jun NH 2 -Terminal Kinase Activation Contributes to Hepatitis B Virus X Protein-Induced Autophagy via Regulation of the Beclin-1/Bcl-2 Interaction

“…The reason for this discrepancy is unclear, but it may be due to the different cell lines or experimental conditions used. Recent evidence highlights the role of ROS in JNK-activated autophagosome formation (46,47), and ROS has also been reported to be closely associated with HBx-and HBV-mediated biological processes (50,51,61). We therefore further investigated the role of ROS in HBx-activated JNK signaling and autophagosome formation.…”

“…Recent work has revealed that ROS may play an important role in JNK-dependent autophagy regulation (46)(47)(48)(49), while HBx treatment has been reported to cause ROS generation in hepatoma cells (50,51). We thus investigated the role of ROS in HBx-induced JNK activation and the ensuing autophagosome formation.…”

Formation and repair of oxidatively generated damage in cellular DNA

“…Similarly, immunological detection of 5-formylcytosine (5-FoCyt) and 5-carboxycytosine (5-CaCyt) generated through iterative oxidation is not accurate because the modifications are present at least 100-fold less than 5-HmCyt. Antibodies against 8-oxoGua are still widely used for sensitive histochemical immunofluorescence detection in human and animal tissues with various clinical applications and basic research involving DNA repair [104–109]. However the specificity of these antibodies remains to be established under in vivo conditions.…”