RESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Qizhuyanggan Decoction (QZD), a traditional Chinese medicine formula, is frequently utilized in clinical practice for managing hepatic fibrosis. However, the specific target and mechanism of action of QZD for hepatic fibrosis treatment remain unknown. AIM OF THE STUDY: By combining network pharmacology, serum medicinal chemistry, and experimental validation methods, our study aimed to investigate the therapeutic effects of QZD on hepatic fibrosis, the anti-hepatic fibrosis active ingredients, and the possible mechanism of anti-hepatic fibrosis action. MATERIALS AND METHODS: The study aimed to investigate the therapeutic effect of QZD on hepatic fibrosis induced by CCl4 in SD rats, as well as its mechanism of action. The rats were anesthetized intraperitoneally using 3% pentobarbital and were executed after asphyxiation with high concentrations of carbon dioxide. Several techniques were employed to evaluate the efficacy of QZD, including ELISA, Western blot, HYP reagent assay, and various pathological examinations such as HE, Masson, Sirius Red staining, and immunohistochemistry (IHC). Additionally, serum biochemical assays were conducted to assess the effect of QZD on liver injury. Network pharmacology, UPLC, molecular docking, and molecular dynamics simulation were utilized to explore the mechanism of QZD in treating hepatic fibrosis. Finally, experimental validation was performed through ELISA, IHC, RT-qPCR, and Western blot analysis. RESULT: Liver histopathology showed that QZD reduced inflammation and inhibited collagen production, and QZD significantly reduced HA and LN content to treat hepatic fibrosis. Serum biochemical analysis showed that QZD improved liver injury. Network pharmacology combined with UPLC screened six active ingredients and obtained 87 targets for the intersection of active ingredients and diseases. The enrichment analysis results indicated that the PI3K/AKT pathway might be the mechanism of action of QZD in the treatment of hepatic fibrosis, and counteracting the inflammatory response might be one of the pathways of action of QZD. Molecular docking and molecular dynamics simulations showed that the active ingredient had good binding properties with PI3K, AKT, and mTOR proteins. Western blot, ELISA, PCR, and IHC results indicated that QZD may treat hepatic fibrosis by inhibiting the PI3K/AKT/mTOR pathway and suppressing M1 macrophage polarization, while also promoting M2 macrophage polarization. CONCLUSIONS: QZD may be effective in the treatment of hepatic fibrosis by inhibiting the PI3K/AKT/mTOR signaling pathway and M1 macrophage polarization, while promoting M2 macrophage polarization. This provides a strong basis for the clinical application of QZD.
Asunto(s)
Química Farmacéutica , Medicamentos Herbarios Chinos , Animales , Ratas , Ratas Sprague-Dawley , Simulación del Acoplamiento Molecular , Farmacología en Red , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Cirrosis Hepática/tratamiento farmacológico , Serina-Treonina Quinasas TOR , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/uso terapéuticoRESUMEN
Purpose: Our study aims to reveal the pharmacological mechanism of Astragaloside IV in the treatment of pulmonary fibrosis(PF) through network pharmacology and experimental validation. Methods: We first determined the in vivo anti-pulmonary fibrosis effect of Astragaloside IV by HE, MASSON staining, and lung coefficients, then used network pharmacology to predict the signaling pathways and molecularly docked key pathway proteins, and finally validated the results by in vivo and in vitro experiments. Results: In in vivo experiments, we found that Astragaloside IV improved body weight (P < 0.05), increased lung coefficients (P < 0.05), and reduced lung inflammation and collagen deposition in mice with pulmonary fibrosis. The network pharmacology results showed that Astragaloside IV had 104 cross-targets with idiopathic pulmonary fibrosis, and the results of KEGG enrichment analysis indicated that cellular senescence could be an important pathway for Astragaloside IV in the treatment of pulmonary fibrosis. Astragaloside IV also bound well to senescence-associated proteins, according to molecular docking results. The results of both in vivo and in vitro experiments showed that Astragaloside IV significantly inhibited senescence protein markers such as P53, P21, and P16 and delayed cellular senescence (P < 0.05). In in vivo experiments, we also found that Astragaloside IV reduced the production of SASPs (P < 0.05), and in in vitro experiments, Astragaloside IV also reduced the production of ROS. In addition, by detecting epithelial-mesenchymal transition(EMT)-related marker protein expression, we also found that Astragaloside IV significantly inhibited the development of EMT in both in vivo and in vitro experiments (P < 0.05). Conclusion: Our research found that Astragaloside IV could alleviate bleomycin-induced PF by preventing cellular senescence and EMT.
Asunto(s)
Bleomicina , Fibrosis Pulmonar Idiopática , Ratones , Animales , Simulación del Acoplamiento Molecular , Farmacología en Red , Transición Epitelial-MesenquimalRESUMEN
Adenosine triphosphate (ATP) is mainly produced in the mitochondrion and used as a universal energy source for various cellular events. Various fluorescent probes for ATP have been established successfully, but most of them are not appropriate for monitoring the fluctuation of the mitochondrial ATP level. Herein, a fluorescent probe named Mito-Rh is first synthesized and used to recognize ATP in mitochondrion. In the probe, rhodamine, diethylenetriamine, and triphenylphosphonium are selected as fluorophore, reaction site, and mitochondrion-targeting group, respectively. Probe Mito-Rh shows high sensitivity to ATP with 81-fold fluorescence enhancement, and the detection range (0.1-10 mM) can match the concentration level of ATP in the mitochondrion. Moreover, Mito-Rh provides excellent selectivity toward ATP over other biological anions (ADP, AMP, GTP, CTP, UTP) owing to a concurrent effect of dual recognition sites (hydrogen bond and π-π stacking). In particular, the probe can localize in mitochondrion specifically and demonstrates utility in the real-time detection of mitochondrial ATP concentration changes.