Neuropathic pain is a difficult-to-treat pain condition that can affect patients for years. (2R,6R)-hydroxynorketamine (R-HNK) is a ketamine metabolite without dissociative effects and has been evaluated as an alternative to ketamine in chronic pain management. The mechanism of action remains elusive. Here we report that repeated systemic or contra-prelimbic cortex (PrL) infusion of R-HNK in the acute stage of nerve injury produces sustained pain relief for at least 14 days in the mouse spared nerve injury (SNI) model of neuropathic pain. Transcriptomic analysis suggests that SNI is associated with increased Brain-derived neurotrophic factor (Bdnf) signaling, abnormal dendritic spine organization, and reduced α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) activity in PrL. Activity-regulated cytoskeleton-associated protein (Arc) is identified as the top gene in the leading-edge analysis of the gene set. R-HNK administration abolishes these transcriptomic changes. Further studies confirm the transcriptome findings. Finally, we show that enhancing PrL activity by R-HNK increases PrL-periaqueductal gray (PAG) connectivity, which is essential for R-HNK-mediated pain relief. Our study highlights AMPAR suppression due to continuous Bdnf/Arc elevation in PrL as a mechanism of central sensitization after SNI. R-HNK can recalibrate Bdnf/Arc/AMPAR axis and restore PrL-PAG connectivity to induce sustained alleviation of neuropathic pain.
Background Anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis is a severe autoimmune neuropsychiatric disease. Brain access of anti-NMDAR autoantibody through the blood–brain barrier (BBB) is essential for pathogenesis. Most previous animal models limit the investigation of etiologies of BBB damage in patients. Methods In this study, we established a novel humanized mouse model of anti-NMDAR encephalitis by intraperitoneal injection of patients’ peripheral blood mononuclear cells (PBMCs) into BALB/c Rag2−/−Il2rg−/−SirpαNODFlk2−/− mice. Results We found that engraftment of patients’ PBMCs not only produced potent anti-GluN1 autoantibodies, but also disrupted BBB integrity to allow brain access of autoantibodies, resulting in a hyperactive locomotor phenotype, anxiety- and depressive-like behaviors, cognitive deficits, as well as functional changes in corresponding brain regions. Transcriptome analysis suggested an exaggerated immune response and impaired neurotransmission in the mouse model and highlighted Il-1β as a hub gene implicated in pathological changes. We further demonstrated that Il-1β was produced by endothelial cells and disrupted BBB by repressing tight junction proteins. Treatment with Anakinra, an Il-1 receptor antagonist, ameliorated BBB damage and neuropsychiatric behaviors. Conclusions Our study provided a novel and clinically more relevant humanized mouse model of anti-NMDAR encephalitis and revealed an intrinsic pathogenic property of the patient’s lymphocytes.
Background Clinically, although chemotherapy is one of the most commonly used methods of treating tumors, chemotherapeutic drugs can induce autophagic flux and increase tumor cell resistance, leading to drug tolerance. Therefore,theoretically, inhibiting autophagy may improve the efficacy of chemotherapy. The discovery of autophagy regulators and their potential application as adjuvant anti-cancer drugs is of substantial importance. In this study, we clarified that Fangjihuangqi Decoction (FJHQ, traditional Chinese medicine) is an autophagy inhibitor, which can synergistically enhance the effect of cisplatin and paclitaxel. Methods We observed the changes of autophagy level in lung cancer cells under the effect of FJHQ, and verified the level of the autophagy marker protein and cathepsin. Apoptosis was detected after the combination of FJHQ with cisplatin or paclitaxel, and NAC (ROS scavenger) was further used to verify the activation of ROS-MAPK pathway by FJHQ. Results We observed that FJHQ induced autophagosomes in lung cancer cells and increased the levels of P62 and LC3-II protein expression in a concentration- and time-gradient-dependent manner, indicating that autophagic flux was inhibited. Co-localization experiments further showed that while FJHQ did not inhibit autophagosome and lysosome fusion, it affected the maturation of cathepsin and thus inhibited the autophagic pathway. Finally, we found that the combination of FJHQ with cisplatin or paclitaxel increased the apoptosis rate of lung cancer cells, due to increased ROS accumulation and further activation of the ROS-MAPK pathway. This synergistic effect could be reversed by NAC. Conclusion Collectively, these results demonstrate that FJHQ is a novel late-stage autophagy inhibitor that can amplify the anti-tumor effect of cisplatin and paclitaxel against non-small cell lung cancer.
Celery seeds have been used as an effective dietary supplement to manage hyperuricemia and diminish gout recurrence. Xanthine oxidase (XOD), the critical enzyme responsible for uric acid production, represents the most promising target for anti-hyperuricemia in clinical practice. In this study, we aimed to establish a method based on affinity ultrafiltration–liquid chromatography–mass spectrometry (UF–LC–MS) to directly and rapidly identify the bioactive compounds contributing to the XOD-inhibitory effects of celery seed crude extracts. Chemical profiling of celery seed extracts was performed using UPLC-TOF/MS. The structure was elucidated by matching the multistage fragment ion data to the database and publications of high-resolution natural product mass spectrometry. Thirty-two compounds, including fourteen flavonoids and six phenylpeptides, were identified from celery seed extracts. UF–LC–MS showed that luteolin-7-O-apinosyl glucoside, luteolin-7-O-glucoside, luteolin-7-O-malonyl apinoside, luteolin-7-O-6′-malonyl glucoside, luteolin, apigenin, and chrysoeriol were potential binding compounds of XOD. A further enzyme activity assay demonstrated that celery seed extract (IC50 = 1.98 mg/mL), luteolin-7-O-apinosyl glucoside (IC50 = 3140.51 μmol/L), luteolin-7-O-glucoside (IC50 = 975.83 μmol/L), luteolin-7-O-6′-malonyl glucoside (IC50 = 2018.37 μmol/L), luteolin (IC50 = 69.23 μmol/L), apigenin (IC50 = 92.56 μmol/L), and chrysoeriol (IC50 = 40.52 μmol/L) could dose-dependently inhibit XOD activities. This study highlighted UF–LC–MS as a useful platform for screening novel XOD inhibitors and revealed the chemical basis of celery seed as an anti-gout dietary supplement.
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