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BACKGROUND: Intervertebral disc degeneration is one of the most common underlying factors causing low back pain. Recent studies have shown that melatonin has a positive effect on alleviating intervertebral disc degeneration. However, the underlying mechanism of melatonin remains to be elucidated. OBJECTIVE: To explore the biological effect and potential mechanism of melatonin in inhibiting hydrogen peroxide (H2O2)-induced injury of human nucleus pulposus cells. METHODS: Human nucleus pulposus cells insolated from degenerative intervertebral disc were cultured in vitro. Cell proliferation and the optimal intervention concentration of melatonin and H2O2 were detected by cell counting kit-8. The Human nucleus pulposus cells treated with H2O2 were used as a model group; the cells treated with H2O2 and intervened with melatonin were used as a melatonin group; the cells cultured in simple medium were used as a control group. The reactive oxygen species levels were detected by 2',7'-dichlorofluorescin diacetate (DCFH-DA), the expression levels of BAX and Caspase3 were detected by immunofluorescence, and the mRNA expression levels of BAX, BCL-2, Casepase3, PI3K and AKT were detected using the real-time fluorescent quantitative reverse transcription PCR. RESULTS AND CONCLUSION: The results of cell counting kit-8 experiment showed that the optimal intervention concentration of H2O2 was 400 µmol/L and the optimal intervention concentration of melatonin was 5 µmol/L. The reactive oxygen species level in the melatonin group was significantly lower than that in the model group. The average fluorescence intensity of BAX and Caspase3 in the melatonin group was significantly lower than that in the model group. The mRNA expressions of BAX and Caspase3 in the melatonin group were lower than those in the model group, while the mRNA expression of Bcl-2 was increased. In addition, the mRNA expressions of PI3K and AKT were also higher in the melatonin group compared with the model group. To conclude, melatonin may protect human nucleus pulposus cells from H2O2-induced oxidative damage through the PI3K/AKT signaling pathway. [ABSTRACT FROM AUTHOR]

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To elucidate the mechanism of Euodiae Fructus stir-fried with water decoction of Coptidis Rhizoma in the treatment of chronic colitis, this study employed ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS), network pharmacology, and experimental verification to predict the involved targets and signaling pathways. The chronic colitis mouse model was constructed to verify the core targets. A total of 48 compounds in the herbal medicine were identified by UPLC-Q-TOF-MS. SwissTargetPrediction was used to screen the potential active components and drug targets. GeneCards, OMIM, PharmGKB, and TDD were used to search for the disease targets. A total of 31 active ingredients, 453 targets of the herbal medicine, and 3 960 targets of chronic colitis were obtained. The common targets shared by the herbal medicine and chronic colitis were introduced into STRING to construct the protein-protein interaction(PPI) network, and CytoNCA plug-in was used to screen the key targets. A total of 90 key targets were obtained, and the key active components included isorhamnetin, quercetin, limonin, and oxyberberine. GO annotation and KEGG pathway enrichment for the key targets were carried out via DAVID. The targets were mainly involved in the positive regulation of protein phosphorylation, positive regulation of nitric oxide biosynthetic process, and negative regulation of apoptotic process. The medicine may treat chronic colitis through PI3 K-Akt, VEGF, HIF-1, and TNF signaling pathways. A mouse model of chronic colitis was established and then treated with Euodiae Fructus stir-fried with the water decoction of Coptidis Rhizoma. The experimental results demonstrated that the medicine can alleviate the pathological damage of colon, significantly reduce the levels of IL-1β, IL-6, and TNF-α, inhibit the activation of PI3 K/Akt pathway, and down-regulate the expression of VEGFA in the treatment of chronic colitis.