Dendrobine alleviates CCl(4)-induced acute liver injury by inhibiting necroptosis through activation of the Nrf2/PPARγ/SOD2 signaling pathways.

BACKGROUND: Acute liver injury (ALI) represents a critical clinical challenge characterized by rapid degradation of hepatic function, necessitating prompt intervention for improved patient outcomes. Dendrobine (DDB) is the main bioactive component of Dendrobium nobile Lindl. (DNL), a traditional Chinese herb renowned for its protective effects against liver injury. However, the specific mechanisms underlying its hepatoprotective effects have not yet been fully elucidated. OBJECTIVE: This study aims to elucidate the potential mechanism underlying the protective effects of DDB against ALI, particularly through the Nrf2/PPARγ/SOD2 pathways, to provide a scientific basis for its application in ALI treatment. METHODS: CCl(4)-induced ALI models in animals were used to evaluate DDB's therapeutic effects via biochemical and pathological analyses. The regulation of the Nrf2/PPARγ/SOD2 axis and mitochondrial ROS (mtROS) by DDB was observed through WB, RT-qPCR, and Mito-SOX assays, which was confirmed both in vitro and in vivo. JASPAR predictions and ChIP assays validated PPARγ's regulation of SOD2. DDB's interaction with Keap1 was assessed by molecular docking, cellular thermal shift assay (CETSA), and drug affinity responsive target stability (DARTS). RESULTS: DDB effectively suppressed MLKL activation and significantly alleviated ALI. DDB also upregulated Nrf2/PPARγ/SOD2 expression and reduced mtROS production. Further studies using pharmacological approaches showed that PPARγ activation increased SOD2 expression, reduced p-MLKL, and lowered mtROS levels. Conversely, PPARγ inhibition reversed these effects and diminished DDB's efficacy. Silencing Nrf2 in vivo decreased PPARγ/SOD2 expression and activated MLKL, counteracting DDB's protective effects. Overexpression of Nrf2 prevented the decrease in PPARγ/SOD2 protein expression induced by CCl(4), and inhibited mtROS release and MLKL activation, indicating that Nrf2 regulates p-MLKL via the PPARγ/SOD2/mtROS axis to suppress necroptosis. Analysis using JASPAR revealed that the SOD2 promoter contains a PPARγ response element. ChIP assays showed that Nrf2 activated SOD2 through PPARγ, not directly transcriptionally regulated SOD2. Further studies demonstrated that DDB interacted with Keap1 to promote Nrf2 nuclear translocation, thereby protecting the liver from damage. CONCLUSION: This study demonstrates that DDB inhibits mtROS and p-MLKL through the Nrf2/PPARγ/SOD2 signaling axis, thereby suppressing necroptosis and ameliorating ALI.