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Various components of Lonicera japonica Thunb. (LJT) exhibit pharmacological activities, including anti-inflammatory and antioxidant effects. Nevertheless, the relationship between LJT and ferroptosis remains largely unexplored. The purpose of this research was to look into the role of LJT in regulating LPS-induced ferroptosis in ALI and to compare the effects of different parts of LJT. We established a mice ALI model by treating with LPS. Administered mice with different doses of Lonicerae Japonicae Flos (LJF), Lonicera Japonica Leaves (LJL) and Lonicerae Caulis (LRC) extracts, respectively. The levels of IL-6, IL-1β, TNF-α, IL-4, IL-10, and PGE2 in bronchoalveolar lavage fluid (BALF) were measured using enzyme-linked immunosorbent assay. Furthermore, the concentrations of superoxide dismutase (SOD), malondialdehyde (MDA), glutathione (GSH), reactive oxygen species (ROS), and total ferrous ions (Fe2+) in lung tissues were evaluated. Hematoxylin and eosin staining was conducted to examine the morphological structure of lung tissues. Transmission electron microscopy was used to investigate the ultrastructural morphology of mitochondria. Furthermore, the effects of LJT were evaluated via immunohistochemical staining, western blotting, and quantitative real-time polymerase chain reaction analyses. Finally, employing molecular docking and molecular dynamics research techniques, we aimed to identify crucial components in LJT that might inhibit ferroptosis by targeting nuclear factor erythroid 2-related factor 2 (Nrf2) and glutathione peroxidase 4 (GPX4). We observed that pretreatment with LJT significantly mitigated LPS-induced lung injury and suppressed ferroptosis. This was supported by reduced accumulation of pro-inflammatory cytokines, ROS, MDA, and Fe2+, along with increased levels of anti-inflammatory cytokines, SOD, GSH, Nrf2, and GPX4 in the lung tissues of ALI mice. Luteolin-7- O -rutinoside, apigenin-7- O -rutinoside, and amentoflavone in LJT exhibit excellent docking effects with key targets of ferroptosis, Nrf2 and GPX4. Pretreatment with LJT may alleviate LPS-induced ALI, possibly by suppressing ferroptosis. Our initial results indicate that LJT activates the Nrf2/GPX4 axis, providing protection against ferroptosis in ALI. This finding offers a promising therapeutic candidate for ALI treatment. [Display omitted] • The different parts of LJT exhibit pharmacological activities in anti-inflammation, antioxidative stress, and inhibition of ferroptosis. Luteolin-7- O -rutinoside, apigenin-7- O -rutinoside, and amentoflavone are potential key components in LJT that exert inhibitory effects on ferroptosis. • Pretreatment with LJT can alleviate LPS-induced ALI by suppressing ferroptosis. • LJT acts on TFRC, SLC7A11, and FHC, activating the Nrf2/GPX4 axis to inhibit ALI-induced ferropotosis. [ABSTRACT FROM AUTHOR]

Ginseng (Panax ginseng C. A. Meyer) is a precious traditional Chinese medicine with multiple pharmacological effects. Ginsenoside Rg1 is a main active ingredient extracted from ginseng, which is known for its age-delaying and antioxidant effects. Increasing evidence indicates that Rg1 exhibits anti-inflammatory properties in numerous diseases and may ameliorate oxidative damage and inflammation in many chronic liver diseases. Chronic inflammatory injury in liver cells is an important pathological basis of many liver diseases. However, its mechanism remains unclear and therapeutic strategies to prevent its development need to be further explored. Thus, our study is to delve the protective effect and mechanism of Rg1 against chronic hepatic inflammatory injuries induced by lipopolysaccharide (LPS). The chronic liver damage model in mice was build up by injecting intraperitoneally with LPS (200 μg/kg) for 21 days. Serum liver function indicators and levels of IL-1β, IL-6 and TNF-α were examined by using corresponding Kits. Hematoxylin and Eosin (H&E), Periodic acid-Schiff (PAS), and Masson stains were utilized to visualize hepatic histopathological damage, glycogen deposition, and liver fibrosis. The nuclear import of p-Nrf2 and the generation of Col4 in the liver were detected by IF, while IHC was employed to detect the expressions of NLRP3 and AIM2 in the hepatic. The Western blot and q-PCR were used to survey the expressions of proteins and mRNAs of fibrosis and apoptosis, and the expressions of Keap1, p-Nrf2 and NLRP3, NLRP1, AIM2 inflammasome-related proteins in mouse liver. The cell viability of human hepatocellular carcinoma cells (HepG2) was detected by Cell Counting Kit-8 to select the action concentration of LPS, and intracellular ROS generation was detected using a kit. The expressions of Nuclear Nrf2, HO-1, NQO1 and NLRP3, NLRP1, and AIM2 inflammasome-related proteins in HepG2 cells were detected by Western blot. Finally, the feasibility of the molecular interlinking between Rg1 and Nrf2 was demonstrated by molecular docking. Rg1 treatment for 21 days decreased the levels of ALT, AST, and inflammatory factors of serum IL-1β, IL-6 and TNF-α in mice induced by LPS. Pathological results indicated that Rg1 treatment obviously alleviated hepatocellular injury and apoptosis, inflammatory cell infiltration and liver fibrosis in LPS stimulated mice. Rg1 promoted Keap1 degradation and enhanced the expressions of p-Nrf2, HO-1 and decreased the levels of NLRP1, NLRP3, AIM2, cleaved caspase-1, IL-1β and IL-6 in livers caused by LPS. Furthermore, Rg1 effectively suppressed the rise of ROS in HepG2 cells induced by LPS, whereas inhibition of Nrf2 reversed the role of Rg1 in reducing the production of ROS and NLRP3, NLRP1, and AIM2 expressions in LPS-stimulated HepG2 cells. Finally, the molecular docking illustrated that Rg1 exhibits a strong affinity towards Nrf2. The findings indicate that Rg1 significantly ameliorates chronic liver damage and fibrosis induced by LPS. The mechanism may be mediated through promoting the dissociation of Nrf2 from Keap1 and then activating Nrf2 signaling and further inhibiting NLRP3, NLRP1, and AIM2 inflammasomes in liver cells. [Display omitted] • Rg1 treatment attenuates LPS-induced chronic liver injury and alleviates fibrosis. • Rg1 treatment attenuates LPS-induced ROS accumulation and hepatocyte inflammation. • Rg1 treatment alleviates LPS-induced chronic liver injury by activating Nrf2 signaling. • Rg1 treatment inhibits LPS-induced hepatocyte NLRP3, NLRP1, and AIM2 inflammasomes. [ABSTRACT FROM AUTHOR]

Xiebai San (XBS), a classic Chinese prescription, has been used for the clinical treatment of pneumonia-related diseases for thousands of years. However, the anti-pneumonia pharmacodynamic material basis of XBS and its underlying mechanisms remain unclear. This study aimed to comprehensively investigate and verify the anti-pneumonia pharmacodynamic material basis and mechanisms of XBS. This study explored the anti-pneumonia activity and key pneumonia targets of XBS in lipopolysaccharide (LPS)-induced zebrafish and RAW264.7 cells in vivo and in vitro through transcriptomics, western blotting, and reverse transcription–quantitative polymerase chain reaction (RT–qPCR). The chemical fingerprint of XBS was established using high-performance liquid chromatography, and the similarities and areas of characteristic peaks of 15 batches of XBS were analyzed. Based on the spectrum–efficacy relationship, the potential anti-inflammatory components were screened according to their peak areas and efficacy using principal component analysis (PCA), bivariate correlation, and partial least squares regression analysis. Active components that bind to core targets were further screened based on surface plasmon resonance (SPR). The binding mode of proteins and components was simulated via molecular docking, which enabled the identification of the primary active components of XBS, thereby elucidating its anti-pneumonia properties. Finally, the anti-inflammatory activities of these components were verified in vitro. XBS decreased neutrophil aggregation in zebrafish and nitric oxide (NO) secretion in RAW264.7 cells as well as suppressed the release of downstream inflammatory cytokines such as iNOS, TNF-α, IL-1β, IL-18, and CXCL10 related to TNF and JAK–STAT signaling pathways. The phosphorylation of IκBα, Akt, and Stat3 was alleviated after XBS in cells. The fingerprint similarities of 15 batches of XBS ranged from 0.381 to 0.994, with a large difference. A total of 15 characteristic peaks were identified, and the relative standard deviation of their peak areas ranged from 24.1% to 70.7%. The results of in vitro anti-inflammatory activities of 15 batches of XBS showed that all samples inhibited the expression levels of NO and nine inflammatory markers. The anti-inflammatory index of 15 batches of XBS was determined to be 0.69–0.96 based on transformation of the anti-inflammatory rate and composite index method via PCA. The spectrum–efficacy relationship model of 15 characteristic peak areas and the anti-inflammatory index showed that 7 main potential active components were related to the anti-inflammatory activity of XBS. Moreover, four components (mulberroside A, isoquercitrin, liquiritigenin, and glycyrrhizic acid) screened based on SPR had different affinities toward TNFR1, Akt1, and Stat3 proteins, and the binding modes were elucidated via molecular docking. Finally, in LPS-induced RAW264.7 cells, all four active components (at a concentration of 60 μM) significantly inhibited the expression levels of NO and inflammatory markers. Based on the comprehensive strategy of spectrum–efficacy relationship and SPR, mulberroside A, isoquercitrin, liquiritigenin, and glycyrrhizic acid were identified as the primary pharmacodynamic active components involved in the anti-pneumonia activity of XBS and were found to intervene in TNF and JAK–STAT signaling pathways. [Display omitted] • Xiebai San (XBS) relieved inflammation by regulating TNF and JAK-STAT pathways. • Potential active components were explored by spectrum–efficacy relationship. • Target and component interactions were evaluated by surface plasmon resonance. • Four anti-inflammatory components of XBS for three targets were revealed and verified. [ABSTRACT FROM AUTHOR]

Aristolochic acids (AAs) are naturally occurring nitro phenanthrene carboxylic acids primarily found in plants of the Aristolochiaceae family. Aristolochic acid D (AAD) is a major constituent in the roots and rhizomes of the Chinese herb Xixin (the roots and rhizomes of Asarum heterotropoides F. Schmidt), which is a key material for preparing a suite of marketed Chinese medicines. Structurally, AAD is nearly identical to the nephrotoxic aristolochic acid I (AAI), with an additional phenolic group at the C-6 site. Although the nephrotoxicity and metabolic pathways of AAI have been well-investigated, the metabolic pathway(s) of AAD in humans and the influence of AAD metabolism on its nephrotoxicity has not been investigated yet. To identify the major metabolites of AAD in human tissues and to characterize AAD O -glucuronidation kinetics in different enzyme sources, as well as to explore the influence of AAD O -glucuronidation on its nephrotoxicity. The O -glucuronide of AAD was biosynthesized and its chemical structure was fully characterized by both 1H-NMR and 13C-NMR. Reaction phenotyping assays, chemical inhibition assays, and enzyme kinetics analyses were conducted to assess the crucial enzymes involved in AAD O -glucuronidation in humans. Docking simulations were performed to mimic the catalytic conformations of AAD in human UDP-glucuronosyltransferases (UGTs), while the predicted binding energies and distances between the deprotonated C-6 phenolic group of AAD and the glucuronyl moiety of UDPGA in each tested human UGT isoenzyme were measured. The mitochondrial membrane potentials (MMP) and reactive oxygen species (ROS) levels in HK-2 cells treated with either AAI, or AAD, or AAD O -glucuronide were tested, to elucidate the impact of O -glucuronidation on the nephrotoxicity of AAD. AAD could be rapidly metabolized in human liver and intestinal microsomes (HLM and HIM, respectively) to form a mono-glucuronide, which was purified and fully characterized as AAD-6- O -β-D-glucuronide (AADG) by NMR. UGT1A1 was the predominant enzyme responsible for AAD-6- O -glucuronidation, while UGT1A9 contributed to a lesser extent. AAD-6- O -glucuronidation in HLM, HIM, UGT1A1 and UGT1A9 followed Michaelis-Menten kinetics, with the K m values of 4.27 μM, 9.05 μM, 3.87 μM, and 7.00 μM, respectively. Docking simulations suggested that AAD was accessible to the catalytic cavity of UGT1A1 or UGT1A9 and formed catalytic conformations. Further investigations showed that both AAI and AAD could trigger the elevated intracellular ROS levels and induce mitochondrial dysfunction and in HK-2 cells, but AADG was hardly to trigger ROS accumulation and mitochondrial dysfunction. Collectively, UGT1A-catalyzed AAD 6- O -glucuronidation represents a crucial detoxification pathway of this naturally occurring AAI analogs in humans, which is very different from that of AAI. Aristolochic acid D (AAD), an abundant naturally occurring aristolochic acid derivative isolated from the Chinese herb Xixin (the roots and rhizomes of Asarum heterotropoides F. Schmidt), can be rapidly converted into AAD-6- O -β-D-glucuronide (AADG) in human tissue preparations. UGT1A-catalyzed AAD 6- O -glucuronidation offers a crucial detoxification pathway of this aristolochic acid derivative in humans, which is very different from that of the nephrotoxin aristolochic acid I (AAI). [Display omitted] • AAD- O -glucuronidation was a major metabolic pathway of AAD in humans. • The glucuronide was biosynthesized and characterized as AAD-6- O -β-D-glucuronide. • UGT1A1 was the predominant enzyme responsible for AAD-6- O -glucuronidation. • AAD-6- O -glucuronidation offers a crucial detoxification pathway of AAD in humans. [ABSTRACT FROM AUTHOR]

The root of Croton crassifolius has been used as a traditional Chinese medicine (TCM), called Radix Croton Crassifolius, and commonly known as "Ji Gu Xiang" in Chinese. Its medicinal value has been recorded in several medical books or handbooks, such as "Sheng Cao Yao Xing Bei Yao", "Ben Cao Qiu Yuan" and "Zhong Hua Ben Cao". It has been traditional employed for treating sore throat, stomach-ache, rheumatism and cancer. At present, there are limited studies on the evaluation of low-polarity extracts of roots in C. crassifolius. Consequently, the aim of this study was to evaluate the antitumor effect of the low-polarity extract of C. crassifolius root. Extracts were obtained by supercritical fluid extraction. The extracts were tested for antitumor effects in vitro on several cancer cell lines. A CCK-8 kit was used for further analysis of cell viability. A flow cytometer and propidium iodide staining were used to evaluate the cell cycle and apoptosis. Hoechst staining, JC-1 staining and the fluorescence probe DCFH-DA were used to evaluate apoptotic cells. Molecular mechanisms of action were analyzed by quantitative RT‒PCR and Western blotting. Immunohistochemistry was used for the evaluation of xenograft tumors in male BALB/c mice. Finally, molecular docking was employed to predict the bond between the desired bioactive compound and molecular targets. Eleven diterpenoids were isolated from low-polarity C. crassifolius root extracts. Among the compounds, chettaphanin II showed the strongest activity (IC 50 = 8.58 μM) against A549 cells. Evaluation of cell viability and the cell cycle showed that Chettaphanin II reduced A549 cell proliferation and induced G2/M-phase arrest. Chttaphanin II significantly induced apoptosis in A549 cells, which was related to the level of apoptosis-related proteins. The growth of tumor tissue was significantly inhibited by chettaphanin II in experiments performed on naked mice. The antitumor mechanism of chettaphanin II is that it can obstruct the mTOR/PI3K/Akt signaling pathway in A549 cells. Molecular docking established that chettaphanin II could bind to the active sites of Bcl-2 and Bax. Taken together, the natural diterpenoid chettaphanin II was identified as the major antitumor active component, and its potential for developing anticancer therapies was demonstrated for the first time by antiproliferation evaluation in vitro and in vivo. [Display omitted] • Eleven diterpenoids were obtained from the SFE extract of C. crassifolius roots. • Chettaphanin II showed the strongest antitumor activity against A549 cells. • Chettaphanin II inhibited A549 cell proliferation and arrested cells in G2/M-phase. • Chettaphanin II induced A549 cell apoptosis by inhibiting PI3K/AKT/mTOR pathway. • Chettaphanin II significantly inhibited the growth of tumor tissue in nude mice. [ABSTRACT FROM AUTHOR]

Polyrhachis vicina Roger (P. vicina), a traditional Chinese medicinal animal, has been used to treat rheumatoid arthritis, hepatitis, cancer, and other conditions. Due to its anti-inflammatory properties, our previous pharmacological investigations have demonstrated that it is effective against cancer, depression, and hyperuricemia. Nevertheless, the key active components and targets of P. vicina in cancers are still unexplored. The study aimed to evaluate the pharmacological treatment mechanism of the active fraction of P. vicina (AFPR) in treating colorectal cancer (CRC) and to further reveal its active ingredients and key targets. To examine the inhibitory impact of AFPR on CRC growth, tumorigenesis assays, cck-8 assays, colony formation assays, and MMP detection were utilized. The primary components of AFPR were identified by GC-MS analysis. The network pharmacology, molecular docking, qRT-PCR, western blotting, CCK-8 assays, colony formation assay, Hoechst staining, Annexin V-FITC/PI double staining, and MMP detection were performed to pick out the active ingredients and potential key targets of AFPR. The function of Elaidic acid on necroptosis was investigated through siRNA interference and the utilization of inhibitors. Elaidic acid's effectiveness to suppress CRC growth in vivo was assessed using a tumorigenesis experiment. Studies confirmed that AFPR prevented CRC from growing and evoked cell death. Elaidic acid was the main bioactive ingredient in AFPR that targeted ERK. Elaidic acid greatly affected the ability of SW116 cells to form colonies, produce MMP, and undergo necroptosis. Additionally, Elaidic acid promoted necroptosis predominantly by activating ERK/RIPK1/RIPK3/MLKL. According to our findings, Elaidic acid is the main active component of AFPR, which induced necroptosis in CRC through the activation of ERK. It represents a promising alternative therapeutic option for CRC. This work provided experimental support for the therapeutic application of P. vicina Roger in the treatment of CRC. [Display omitted] • The active fraction of Polyrhachis vicina Roger (AFPR) inhibited CRC growth in vivo and in vitro. • Network pharmacology and molecular docking predicted ERK might be the key target of AFPR in antagonizing CRC. • Elaidic acid was a key active component of AFPR and induced necroptosis. • Elaidic acid induced necroptosis via ERK activation. [ABSTRACT FROM AUTHOR]