Daphnetin alleviates silica-induced pulmonary inflammation and fibrosis by regulating the PI3K/AKT1 signaling pathway in mice.

Silicosis is a hazardous occupational disease caused by inhalation of silica, characterized by persistent lung inflammation that leads to fibrosis and subsequent lung dysfunction. Moreover, the complex pathophysiology of silicosis, the challenges associated with early detection, and the unfavorable prognosis contribute to the limited availability of treatment options. Daphnetin (DAP), a natural lactone, has demonstrated various pharmacological properties, including anti-inflammatory, anti-fibrotic, and pulmonary protective effects. However, the effects of DAP on silicosis and its molecular mechanisms remain uncover. This study aimed to evaluate the therapeutic effects of DAP against pulmonary inflammation and fibrosis using a silica-induced silicosis mouse model, and investigate the potential mechanisms and targets through network pharmacology, proteomics, molecular docking, and cellular thermal shift assay (CETSA). Here, we found that DAP significantly alleviated silica-induced lung injury in mice with silicosis. The results of H&E staining, Masson staining, and Sirius red staining indicated that DAP effectively reduced the inflammatory response and collagen deposition over a 28-day period following lung exposure to silica. Furthermore, DAP reduced the number of TUNEL-positive cells, increased the expression levels of Bcl-2, and decreased the expression of Bax and cleaved caspase-3 in the mice with silicosis. More importantly, DAP suppressed the expression levels of NLRP3 signaling pathway-related proteins, including NLRP3, ASC, and cleaved caspase-1, thereby inhibiting silica-induced lung inflammation. Further studies demonstrated that DAP possesses the ability to inhibit the epithelial mesenchymal transition (EMT) induced by silica through the inhibition of the TGF-ß1/Smad2/3 signaling pathway. The experimental results of proteomic analysis found that the PI3K/AKT1 signaling pathway was the key targets of DAP to alleviate lung injury induced by silica. DAP significantly inhibited the activation of the PI3K/AKT1 signaling pathway induced by silica in lung tissues. The conclusion was also verified by the results of molecular and CETSA. To further verify this conclusion, the activity of PI3K/AKT1 signaling pathway was inhibited in A549 cells using LY294002. When the A549 cells were pretreated with LY294002, the protective effect of DAP on silica-induced injury was lost. In conclusion, the results of this study suggest that DAP alleviates pulmonary inflammation and fibrosis induced by silica by modulating the PI3K/AKT1 signaling pathway, and holds promise as a potentially effective treatment for silicosis.

[Intervention effect of Jingfang Mixture on urticaria mice based on NF-κB/NLRP3/IL-1ß signaling pathway].

This study explored the curative effect of Jingfang Mixture on urticaria mice induced by aluminum hydroxide/ovalbumin, and discussed its mechanism. Sixty male Kunming mice were randomly divided into a normal group, a model group, three Jingfang Mixture(low-dose, medium-dose, and high-dose) groups, and a positive drug(cetirizine hydrochloride) group. The urticarial model in mice was induced by the intraperitoneal injection of the mixed solution of ovalbumin and aluminum hydroxide. The degrees of pruritus were observed after the second immunization. Pathological changes were detected by hematoxylin and eosin(HE) staining. Levels of interleukin 1ß(IL-1ß) and tumor necrosis factor α(TNF-α) in the serum were detected by enzyme linked immunosorbent assay(ELISA). Expressions of NOD-like receptor protein 3(NLRP3) and IL-1ß were detected by immunohistochemistry(IHC). Expressions of nuclear factor kappa-B(NF-κB p65), NLRP3, apoptosis-associated speck-like protein containing a CARD(ASC), cysteinyl aspartate-specific proteases 1(caspase-1), and IL-1ß proteins were detected by Western blot. The results showed that, except for the normal group, the mice in all groups had different degrees of pruritus. Compared with the model group, the Jingfang Mixture groups and the positive drug group prolonged the scratching latency of mice(P<0.05), and significantly reduced the number of scratching(P<0.05). In addition, the Jingfang Mixture groups and the positive drug group improved the pathological morphology of skin tissue. The expression levels of IL-1ß and TNF-α in serum were significantly reduced(P<0.05), and the number of NLRP3 and IL-1ß positive cells was decreased(P<0.01). The expressions of p-NF-κB p65, NLRP3, ASC, cleaved caspase-1, and IL-1ß protein were significantly down-regulated(P<0.05). The results of the above study indicate that Jingfang Mixture inhibit the inflammatory response in urticaria mice, and the mechanism may be related to the inhibition of activating NF-κB/NLRP3/IL-1ß signaling pathway.

Pretreatment with Tilianin improves mitochondrial energy metabolism and oxidative stress in rats with myocardial ischemia/reperfusion injury via AMPK/SIRT1/PGC-1 alpha signaling pathway.

Mitochondrial energy metabolism and oxidative stress play a crucial role in ameliorating myocardial ischemia/reperfusion injury (MIRI). Tilianin has been reported to have a significant protection for mitochondrion in MIRI. However, the underlying mechanisms remain unknown. This study investigated whether Tilianin regulates mitochondrial energy metabolism and oxidative stress in MIRI via AMPK/SIRT1/PGC-1 alpha signaling pathway. The MIRI model was established by 30 min of coronary occlusion followed by 2 h of reperfusion in rats. The results revealed that Tilianin significantly reduced myocardial infarction, improved the pathological morphology of myocardium, markedly increased the contents of ATP and NAD+, decreased ADP and AMP contents and the ratio of AMP/ATP, reduced the level of ROS and MDA, enhanced SOD activity, evidently increased the levels of AMPK, SIRT1 and PGC-1 alpha mRNA, up-regulated the expressions of AMPK, pAMPK, SIRT1, PGC-1alpha, NRF1, TFAM and FOXO1 proteins. However, these effects were respectively abolished by Compound C (a specific AMPK inhibitor) and EX-527 (a specific SIRT1 inhibitor). Taken together, this study found that Tilianin could attenuate MIRI by improving mitochondrial energy metabolism and reducing oxidative stress via AMPK/SIRT1/PGC-1 alpha signaling pathway.