As one of the sources of Tibetan medicine “Dali”, Rhododendron has the effects of relieving cough and asthma, clearing heat and detumescence and tonifying the kidney(23). It is clinically used to treat respiratory diseases such as cough, asthma, phlegm and so on(24). Its chemical constituents mainly include flavonoids, sesquiterpenoids, triterpenoids, chromogen ketones, phenolic acids and etc.(22). In this study, the active components of RAM were obtained to the greatest extent through multiple literature search databases, after screening the components in TCMSP and PubChem database, the related targets of each component were integrated, and the active targets and involved pathways of ALI/ARDS were studied by using those databases. The mechanism of RAM in the treatment of ALI/ARDS was explained from the perspective of targets and signal pathways, the “Component-Target-Protein” interaction network diagram of RAM in the treatment of ALI/ARDS was constructed, 92 active components and 46 common action targets of RAM were screened, and 13 significant and effective action targets were screened by STRING and Cytoscape software. At the same time, GO and KEGG enrichment showed that they involved 26 biological processes, 4 cellular components, 8 molecular functions and 18 signal pathways. Through the network pharmacological analysis of the components and targets of RAM, the targets and pathways of RAM in the treatment of ALI/ARDS had a scientific basis.
By searching literatures and online pharmacological research methods, most of the screening criteria for drug active ingredients included integrated model ADME information screening to identify the potential bioactive, including the evaluation of Caco2 permeability, oral bioavailability (OB), and drug-likeness (DL), and half-life (HL) were used to apply ADME-related models(25–27). In this study, because most Tibetan drugs were natural drugs, with diverse sources of drugs, complex chemical components, and the active components that exert the efficacy were not clear. Meanwhile, there were few studies on the mechanism of drug action, and many factors such as cultivation, harvesting, processing and storage might have a certain impact on the components of drugs(28). Combined with various literature databases, it was found that there were few studies on RAM with messy data information, and its active components and action targets were less recorded in TCMSP and PubChem databases. Based on the above reasons, in the current study, we did not screen the active components of RAM at the ADME level. Therefore, the chemical components of RAM that might play a role but were not retrieved in TCMSP and PubChem need to be studied.
Through the screening of active components and target prediction of RAM, it could be seen that RAM mainly contains 92 chemical components such as naringenin, isorhamnetin, kaempferol and avicularin. A Study had reported that naringenin can regulate T cell subsets and reduce the serum concentration of proinflammatory cytokines such as TNF-α, INF-γ, IL-6, and the globular inflammatory reaction, so as to achieve the effect of treating inflammation(29). Isorhamnetin inhibited the activity of myeloperoxidase (MPO), the levels of TNF-α and IL-6, to abolish inflammation and support the mRNA expression of pro-inflammatory mediators(30). Kaempferol reduced the infiltration of CD3 + T cell and gene expression of major pro-inflammatory cytokines including IL-6, IL-17A and TNF-α, at the same time, it also reduced the signal of regulated pro-inflammatory NF-κB(31). Avicularin prevented the activation of MEK/NF-κB pathway, inhibited inflammatory response, prevented cell viability, and induced TNF-α treatment of apoptosis of human RA synovial cells(32). It can be seen that the active ingredients represented by naringenin, isorhamnetin, kaempferol and avicularin play an important role RAM in the treatment of ALI/ARDS, especially in terms of anti-inflammatory.
Based on the“components-target-protein” network and PPI network, 13 core targets were screened for RAM treatment of ALI/ARDS, including IL-6, TNF, CXCL8, NOS2, RELA, NF-κB1, NF-κBIA, etc, which were closely related to inflammation, immune and inflammatory response, cell growth, differentiation and apoptosis. As an inflammatory factor, IL-6 is an indicator of inflammatory response, and TNF can promote the expression of IL-6 and mediate inflammatory response(33). The abnormal regulation of CXCL8 and its receptors was related to many inflammatory mediated diseases. In response to external stimuli, lung macrophages secrete CXCL8, which coordinated the migration of PMN to the lungs and led to airway inflammatory diseases(34). Through enhancing macrophage migration and survival, circulating inflammatory signals, and impairing lipid metabolism, the lung SCC was promoted by NOS2. Also, inflammatory signals between macrophages and epithelial cells could be circulated by NOS2(35). As macrophages, RELA was significant in inflammatory response, excessive RELA activation in macrophages during inflammation could lead to serious tissue damage and endanger health(36). A group of transcription factors composed of NF-κB family contained NF-κB1, NF-κBIA were crucial mediators of inflammatory response. In addition to the inflammatory response, there were various signal transduction processes gathered on the NF-κB pathway, including cell proliferation, apoptosis, angiogenesis and many other processes(37). From the above description, it was not difficult to see that the above targets were closely related to the subsequent pathways
Go and KEGG enrichment analysis showed that the biological processes of RAM in the treatment of ALI/ARDS were mainly related to the response of cellular response to lipopolysaccharide, inflammatory response, positive regulation of nitric oxide biosynthetic process, etc. Molecular functions were mainly focused on identical protein binding, transcription factor binding, etc. Cellular components were mainly involved in I-κB/NF-κB complex, cytosol and so on. At the same time, combined with KEGG pathway enrichment analysis, RAM treatment of ALI/ARDS involved Toll-like receptor signaling pathway, TNF signaling pathway, etc. By querying the role and mechanism of each pathway in PubChem database, it could be concluded that cAMP signaling pathway, MAPK signaling pathway, Sphingolipid signaling pathway,PI3K-Akt signaling pathway, FoxO signaling pathway can regulate many physiological processes such as cell growth, differentiation, apoptosis and death, while other 8 pathways such as Toll-like receptor signaling pathway, NOD-like receptor signaling pathway, RIG-I-like receptor signaling pathway, NF-κB signaling pathway were related to cellular inflammatory response, immune response and the NF‐κB dependent expression of inflammatory cytokines. And Jak-STAT signaling pathway, TNF signaling pathway, Adipocytokine signaling pathway can participate in many important biological processes such as cell proliferation, differentiation, apoptosis and immune regulation at the same time. In addition, when screening all KEGG pathways provided by David website, we retained small cell lung cancer, and as the only lung related disease in the results, we believe that it might be related to ALI/ARDS to some extent.
In conclusion, this study shown that RAM might act on 46 targets such as TNF, TP53, IL6, PTGS2, CCL2 and VEGFA through 92 active components to regulate Toll-like receptor signaling pathway, NOD-like receptor signaling pathway, RIG-I-like receptor signaling pathway and NF-κB signaling pathway, to involve in anti-inflammatory, immune response and promoting inflammatory cytokines NF‐κB dependent expression and other biological processes, which play a therapeutic role in the treatment of ALI/ARDS. This study made an exploratory study on the action pathway and mechanism of RAM on ALI/ARDS. It was found that RAM played an effective role in the treatment of ALI/ARDS through the synergistic action of multi-component, multi-target and multi-pathway, which provided a theoretical basis for the follow-up in-depth research, in order to provide new ideas and references for the development and utilization of RAM. However, further experimental research and demonstration are still needed to lay a foundation for further exploring the mechanism of RAM’s drug-target interaction