ALI is a syndrome with various causes, where oxidative stress and inflammation are key factors affecting lung function[55,56].Despite extensive research in determining the influencing factors and repair mechanisms[57], current treatments have not significantly reduced the high mortality rate of ALI[58].
Research has demonstrated that TCM is effective in treating coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2[59].TCM targets inflammation, oxidative stress, and organ injuries associated with the virus, providing unique clinical advantages[60]. TCM is now a crucial component in preventing and treating ALI, classified as "lung heat syndrome" and "asthma syndrome"in TCM.The pathogenesis incloud evil toxin trapped in the lung causes heat to consume body fluid, leading to lung dysfunction and accumulation of phlegm and heat[61]. This can progress to ALI and potentially acute respiratory distress syndrome (ARDS). DYY has been used in China for centuries for epidemic diseases.It has the effect of eliminating turbid, clearing heat and detoxification. It is usually used to treat influenza, cold, fever and other upper respiratory diseases, showing a good effect.
ALI is characterized by uncontrolled inflammation and redox imbalance[62], with a focus on oxidative stress, inflammatory response and apoptosis in treatment research[63]. Network pharmacology is commonly used to uncover the complex pharmacological mechanisms of traditional Chinese medicine for treating complex diseases.This study used network pharmacology and vivo experiments to investigate how DYY works as an anti-ALI treatment. We confirmed 15 main targets out of 111 potential targets identified, including TP53, TNF, IL6, MAPK1,CASP3, and BCL2, associating with oxidative stress, inflammation, and apoptosis.
Oxidative stress causes the buildup of ROS in lung cells, impacting their function and triggering inflammation. In acute lung injury, excessive NO production due to oxidative stress can lead to the formation of reactive nitrogen species (RNS)[64],disrupting pulmonary function[65],which leads to induced NOS overexpression / activity and the release of pro-inflammatory cytokines [66]. In this study, we created an ALI rat model using LPS and treated it with DYY. Results showed that DYY effectively treated ALI by restoring normal W/D ratio, reducing inflammatory cell infiltration, and reconstructing alveolar structure.After LPS stimulation, pro-inflammatory cytokines like IL-6 and TNF-α were secreted in BALF, indicating pulmonary inflammation. DYY reversed this by reducing cytokines, indicating inhibition of the inflammatory response.
It is well known that oxidative stress is caused by an imbalance between oxidants and antioxidants, leading to damage from ROS and depletion of antioxidants. This can worsen inflammation and harm mitochondria due to the production of oxides exceeding antioxidant defense[67].MDA is a common biomarker for oxidative stress[68],inducer NO synthase (iNOS) and endothelial nitric oxide synthase (eNOS)are enzymes involved in oxidative stress and can contribute to lung injury in ALI by increasing superoxide production such as NO and MDA[69].New research suggests that oxidative stress is a key factor in the development of ALI, with ROS playing a crucial role in the process. Mitochondrial channels like mPTP and inner membrane anion channel (IMAC),their activation may be involved in intra- and intermitochondrial redox-environment changes leading to ROS release[70]. It should be noted that while ROS can protect cells from oxidative stress, high levels can damage endothelial barriers and cause inflammation. Excessive ROS can deplete NO levels, and NO reacts with ROS to form excessive peroxynitrite, leading to oxidative damage and cell death through reacting with lipids, DNA and proteins[67].UCP2, an anion transporter, helps maintain mitochondrial function, immune response and regulate oxidative stress, and protect against cell apoptosis[71,45].Here, it may play a crucial role in protecting against LPS-induced ALI by stabilizing mitochondrial structure and reducing inflammation and oxidative stress induced by ROS, MDA and so on . To confirm DYY's inhibitory effect on ALI in rats, we observed changes in ROS, MDA, and NO levels. ROS levels increased significantly in ALI rats, indicating oxidative damage in lung tissue. MDA and NO levels also increased, indirectly reflecting tissue injury degree. DYY intervention effectively reduced intracellular ROS, inhibited ROS accumulation, decreased MDA and NO levels, tending to the Con group,and reversed the decrease in UCP2 mRNA expression induced by LPS. This suggests that DYY can alleviate lung tissue inflammation and improve ALI by regulating oxidative stress processes.
We analyzed the mechanism of DYY in ALI through KEGG and GO enrichment , identifying involvement of mitochondrial membrane, cell response to nitrogen compounds and pathways like PI3K-AKT and MAPK signaling in ALI development. By studying the signaling cascade, we aim to understand the key regulatory roles of upstream and downstream targets in inflammation and oxidative stress pathways in ALI. Numerous studies have demonstrated the significance of Nrf2 in controlling oxidative stress[72] and its role in iron apoptosis by regulating iron homeostasis and lipid peroxidation [73,74]. Furthermore,the Nrf2/HO-1 pathway helps regulate anti-inflammatory and antioxidant responses, providing multi-organ protection[75,76]. Recent reports suggest that natural drug ingredients protect against ALI through the Nrf2 signaling pathway[77]. Our study shows that DYY activates the Nrf2/HO-1 pathway in ALI rats, reducing oxidative stress and inflammation in the lungs,such as the reduction of TLR4 expression and TNF-α,IL-6 levels,suggesting that DYY can prevent ALI by reducing inflammation and increasing antioxidant response. Nrf2 protects lung cells by influencing TLR4 signaling[78], confirming its role in inflammatory lung injury,consistenting with previous studies [79].
Molecular docking techniques were used to predict the binding activity of core components to key targets. Results showed Honokiol had good binding activity to HO-1 and TLR4, while other components bound well to Nrf2, HO-1, and TLR4. Quercetin, a flavonoid compound, regulates oxidative stress and inflammation by inhibiting inflammatory factors and excessive release of ROS, proving effective in treating various diseases[80].Wogonin is also flavonoids that reduce inflammation and oxidative stress by enhancing antioxidant capacity and counteracting inflammatory signals[81].Honokiol is a natural polyphenol that has been shown to counteract oxidative stress and inflammatory signals in a variety of ways, including reversing elevated levels of inflammatory factors, increasing the production of antioxidants GSH and SOD in the body, and alleviating LPS-induced apoptosis[82,83].Prior research has demonstrated that Licochalcone A has antimicrobial and anti-inflammatory properties, protects against oxidative stress, and activates nuclear translocation of Nrf2 and enhancing HO-1 expression in cells[84]. We speculate DYY improves ALI in rats by utilizing multiple compounds.
Our study shows that DYY protects against ALI in rats by reducing oxidative stress and inflammation through the Nrf2/HO-1 pathway, inhibiting TLR4 signaling. However, further research is needed to identify the main bioactive ingredients and fully understand the complex signaling cascade mechanisms involved in its anti-ALI effects.Nevertheless, these findings provide a further pharmacological basis for DYY as a new treatment for ALI.