Inhibiting NLRP3 inflammasome activation prevents copper-induced neuropathology in a murine model of Wilson's disease.

Wilson's disease (WD) is an inherited disorder characterized by excessive accumulation of copper in the body, particularly in the liver and brain. In the central nervous system (CNS), extracellular copper accumulation triggers pathological microglial activation and subsequent neurotoxicity. Growing evidence suggests that levels of inflammatory cytokines are elevated in the brain of murine WD models. However, the mechanisms associated with copper deposition to neuroinflammation have not been completely elucidated. In this study, we investigated how the activation of NLR family pyrin domain containing 3 (NLRP3) inflammasome contributes to copper-mediated neuroinflammation in an animal model of WD. Elevated levels of interleukin-1ß, interleukin-18, interleukin-6, and tumor necrosis factor-α were observed in the sera of WD patients and toxic milk (TX) mice. The protein levels of inflammasome adaptor molecule apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC), cleaved caspase-1, and interleukin-1ß were upregulated in the brain regions of the TX mice. The NLRP3 inflammasome was activated in the TX mice brains. Furthermore, the activation of NLRP3 inflammasome was noted in primary microglia treated with CuCl2, accompanied by the increased levels of cleaved caspase-1, ASC, and interleukin-1ß. Blocking NLRP3 inflammasome activation with siNlrp3 or MCC950 reduced interleukin-1ß and interleukin-18 production, thereby effectively mitigating cognitive decline, locomotor behavior impairment, and neurodegeneration in TX mice. Overall, our study demonstrates the contribution of copper overload-mediated activation of NLRP3 inflammasome to progressive neuropathology in the CNS of a murine model of WD. Therefore, blockade of the NLRP3 inflammasome activation could be a potential therapeutic strategy for WD.

Gandou decoction, a Chinese medicinal formula, in the treatment of hepatic injury by Wnt/ß-catenin pathway regulation in models of Wilson disease.

BACKGROUND: Gandou decoction (GDD) has been widely used in the treatment of Wilson disease (WD) for decades. It is optimized from the Dahuanghuanglianxiexin decoction, Yinchenhao decoction, and Huanglianjiedu decoction. It was first reported in the Treatise on Febrile and Miscellaneous Diseases and A Handbook of Prescriptions for Emergencies published in the Eastern Han Dynasty and the Eastern Jin Dynasty respectively. Hepatic injury is one of the most severe complications of WD. The current study aimed to explore the hepatic-protection effects of GDD and its exact therapeutic target, with a particular focus on the expression of oxidative stress and the Wnt/ß-catenin pathway. METHODS: Hepatic injury was induced in a copper-loaded rat model using the intragastric administration of copper(II) sulfate pentahydrate (CuSO4·5H2O). The water extract of GDD (0.4 g/kg/d) was administered twice a day for 4 weeks. Copper content and alanine aminotransferase (ALT) level, structural observation under the microscope, and immunohistochemical analysis of liver tissue were performed after the final administration. Moreover, the expression of ß-catenin, GSK3ß, Dishevelled-3, c-Myc, and p-GSK3ß of liver tissue were detected to explore the relationship between the hepatic protection of GDD and the Wnt/ ß-catenin signal pathway of GDD. We also stimulated the rat hepatic cell line BRL-3A with CuSO4·5H2O to establish a hepatic injury cytomodel. GDD serum at a concentration of 20% was administered into the model cell for 24 h. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry were performed to detect cell viability, mitochondrial membrane potential (MMP), and the expression of reactive oxygen species (ROS). Meanwhile, the expression of the Wnt/ß-catenin signal pathway-related proteins was evaluated. RESULTS: GDD reduced copper and ALT while inhibiting oxidative stress and the degeneration and necrosis of liver tissue and hepatocytes. Treatment with GDD improved cell viability and MMP while suppressing the ROS level. Furthermore, GDD rectified the expression of Wnt/ß-catenin signal pathway-related proteins in both livers of the copper-loaded and copper-stimulated BRL-3A cell lines. CONCLUSIONS: GDD had apparent therapeutic effects on the hepatic injury of copper-loaded rats and copper-stimulated BRL-3A cells. Its mechanism is related to its regulatory effect on the Wnt/ß-catenin pathway rectification and oxidative stress antagonism.

Thymoquinone Prevents Dopaminergic Neurodegeneration by Attenuating Oxidative Stress Via the Nrf2/ARE Pathway.

Studies have indicated that oxidative stress plays a crucial role in the development of Parkinson's disease (PD) and other neurodegenerative conditions. Research has also revealed that nuclear factor erythroid 2-related factor 2 (Nrf2) triggers the expression of antioxidant genes via a series of antioxidant response elements (AREs), thus preventing oxidative stress. Thymoquinone (TQ) is the bioactive component of Nigella sativa, a medicinal plant that exhibits antioxidant and neuroprotective effects. In the present study we examined whether TQ alleviates in vivo and in vitro neurodegeneration induced by 1-methyl-4-phenylpyridinium (MPP+) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) by acting as an activator of the Nrf2/ARE cascade. We showed that TQ significantly reduced MPP+-mediated cell death and apoptosis. Moreover, TQ significantly elevated the nuclear translocation of Nrf2 and significantly increased the subsequent expression of antioxidative genes such as Heme oxygenase 1 (HO-1), quinone oxidoreductase (NQO1) and Glutathione-S-Transferase (GST). The application of siRNA to silence Nrf2 led to an abolishment in the protective effects of TQ. We also found that the intraperitoneal injection of TQ into a rodent model of PD ameliorated oxidative stress and effectively mitigated nigrostriatal dopaminergic degeneration by activating the Nrf2-ARE pathway. However, these effects were inhibited by the injection of a lentivirus wrapped Nrf2 siRNA (siNrf2). Collectively, these findings suggest that TQ alleviates progressive dopaminergic neuropathology by activating the Nrf2/ARE signaling cascade and by attenuating oxidative stress, thus demonstrating that TQ is a potential novel drug candidate for the treatment of PD.