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As a zoonotic pathogen, S. suis serotype 2 (SS2) can cause severe diseases in both pigs and humans, and develop resistance to antibiotics. Plant natural compounds are regarded as promising alternatives to conventional antibiotics. Phillyrin is the major bioactive components of Chinese herbal medicine Forsythia suspensa. In this study, we explored the activity and action mechanism of phillyrin against SS2. The results showed that phillyrin could disrupt membrane integrity, destroy intracellular structures, and increase the exosmosis of DNA. Results of PCR revealed that phillyrin affected bacterial-virulence-related genes' expression levels. Meanwhile, phillyrin significantly decreased the adhesion activity, inhibited lactate dehydrogenase (LDH) secretion, and reduced biofilm formation of SS2 in Newborn pig trachea epithelial (NPTr) cells. Furthermore, phillyrin protected tight junction protein of NPTr cells from SS2. We reported that phillyrin (0.1 mg/kg) treatment after bacterial challenge significantly improved the survival rate, ameliorated pulmonary inflammation, and inhibited the accumulation of multiple cytokines (IL-1, IL-6, IL-8, and TNF-α). Molecular docking showed that phillyrin had a good binding activity with the Ala88 and Asp111 of suilysin (SLY), one of the most important virulence factors of SS2. Collectively, phillyrin possesses antibacterial and anti-inflammatory activities, and is a promising candidate for preventing SS2 infection. [ABSTRACT FROM AUTHOR]

Gouty arthritis (GA) is an inflammatory disorder characterized by deposition of monosodium urate (MSU) crystal in joints. Phillyrin, a natural compound with anti-inflammatory properties, shows promise in mitigating inflammatory responses. This study investigates the therapeutic potential of phillyrin in GA and explores its mechanisms of action. GA was induced in mice via intraarticular MSU injection, and joint inflammation, inflammatory cell infiltration, and their level in serum/tissue were assessed. Key proteins in the NF-κB and NLRP3 pathways were examined using western blot analysis. The impact of phillyrin on oxidative stress, neutrophil extracellular trap (NET) formation, and neutrophil accumulation was evaluated by measuring CD11b + Ly6G + cells, MPO, CitH3, extracellular DNA ratio, and oxidative stress markers. In vitro studies assessed the effects of phillyrin on oxidative stress, cell viability, cytokine production, and NET formation in MSU-treated neutrophils. The KEAP1/NRF2 pathway's role was analyzed using ML385, an NRF2 inhibitor. Phillyrin significantly reversed MSU-induced ankle swelling and inflammatory cell infiltration in joint tissues. It suppressed pro-inflammatory cytokines and proteins in the NF-κB and NLRP3 pathways. Phillyrin reduced neutrophil infiltration, evidenced by lower MPO activity and NET formation, marked by reduced CitH3 expression. In vitro, phillyrin inhibited inflammatory marker expression and NET formation without affecting cell viability. It also restored antioxidant enzyme levels and reduced ROS production, regulating the KEAP1/NRF2 pathway, enhancing NRF2 expression and stability. These effects were reversed by NRF2 inhibition with ML385. Phillyrin alleviates GA by reducing joint inflammation, inhibiting NET formation, and suppressing oxidative stress through NRF2 modulation. [ABSTRACT FROM AUTHOR]

ABSTRACT Phillyrin (PHY), also known as forsythin, is an active constituent isolated from the fruit of Forsythia suspensa (Thunb.) Vahl (Oleaceae). It exhibits anti-inflammatory, anti-viral, and antioxidant properties. However, the precise impact of PHY on colitis induced by dextran sodium sulfate (DSS) and its mechanism remain elusive. The present investigation revealed that PHY (12.5, 25.0, and 50.0 mg/kg) exhibited significant therapeutic efficacy in protecting mice against DSS-induced colitis. This effect was manifested as reduced weight loss, a shortened colon, increased secretion of inflammatory factors, increased intestinal permeability, and an enhanced disease activity index in mice with ulcerative colitis (UC). Molecular investigations have determined that PHY mitigates the nuclear translocation of nuclear factor kappa B, thereby downregulating myosin light-chain kinase-driven myosin light-chain phosphorylation. This mechanism results in the preservation of the integrity of the intestinal barrier. The outcomes of 16S rRNA sequencing suggest that PHY (50 mg/kg) augmented the relative abundance of certain probiotic strains, including Lactobacillaceae and Lachnospiraceae. Additionally, PHY supplementation elevated the short-chain fatty acid contents within the intestinal contents of mice with UC. In conclusion, pre-treatment with PHY may ameliorate the DSS-induced UC in mice by lowering the expression of inflammatory factors, protecting intestinal barrier function, and enhancing the structure of the intestinal flora.IMPORTANCEThe protective effect of phillyrin on DSS-induced colitis was explained for the first time, and the anti-inflammatory effect of phillyrin was demonstrated by fecal microbiota transplantation experiments mainly through intestinal flora.

Background: Phillyrin, the major lignin compound of Forsythia suspense (Thunb.) Vahl, has been shown the effects of anti-inflammatory and antioxidant. Our study was aimed to explore the protective effect of phillyrin on glomerular mesangial cells (HBZY-1) and the potential mechanism. Methods: Cell viability, cytokine production, levels of reactive oxygen radicals (ROS), glutathione (GSH), malondialdehyde (MDA), and superoxide dismutase (SOD), as well as autophagy and apoptosis levels were determined to verify the mechanism of phillyrin on HBZY-1 cells. Results: Our result indicated that phillyrin significantly inhibited HG-induced HBZY-1 proliferation by inhibiting Bcl-2 expression and upregulating Bad, cleaved caspase-3, and -9 expression. Also, phillyrin suppressed HG-induced mesangial extracellular matrix accumulation by inhibiting the expression of fibronectin and transforming growth factor-β1. Further, phillyrin inhibited oxidative stress and inflammation by decreasing ROS, MDA, TNF-α, IL-1β, and IL-6 contents and increasing SOD and GSH expression. Phillyrin also promoted autophagy by increasing LC3-II/LC3-I ratio and down-regulating p62 expression. Furthermore, WB assay showed that phillyrin inhibited oxidative stress caused by HG via activating Nrf2 signaling pathway, while attenuated proliferation and inflammation in HBZY-1 cells through inactivating PI3K/Akt/mTOR and NF-κB pathways. Conclusion: All results showed that phillyrin might be a promising therapeutic agent for the treatment of DN. Graphical Abstract [ABSTRACT FROM AUTHOR]

Background: Nasopharyngeal carcinoma (NPC) is one of the malignant cancers in southern China. Phillyrin is a major active constituent extracted from Forsythia suspensa (Thunb.) and possesses anti-inflammatory and anti-tumor properties. Objective: This study was conducted to clarify the functional role and mechanism of autophagy in the anti-tumor effects induced by phillyrin, as well as the combined effects of phillyrin and the autophagy inhibitor chloroquine (CQ). Results: Phillyrin dose-dependently suppressed NPC cell viability, whereas promoted cell apoptosis. In addition, phillyrin induced autophagy in NPC cells by increasing Beclin-1 protein expression and LC3-II/I ratio as well as decreasing p62 protein level. Pathway analysis showed that phillyrin enhanced the phosphorylation levels of AMPK but inhibited the phosphorylation of mTOR and p70s6k. Furthermore, the combined use of phillyrin and CQ further suppressed CNE-1 cell viability and promoted cell apoptosis. Conclusion: These findings indicate that inhibition of autophagy by CQ enhances phillyrin-mediated anti-tumor activity in vitro, which may provide a novel strategy to improving the anti-cancer efficacy of drug treatment. [ABSTRACT FROM AUTHOR]

Objectives: Prolonged exposure to chronic hypertension places the heart under excessive strain, resulting in myocardial remodeling. Phillyrin, derived from the natural plant Forsythia suspensa, has been found to possess cardioprotective properties. The objective of this study is to investigate the role and mechanism of phillyrin in hypertension-induced myocardial remodeling in mice. Methods: We constructed a mouse model of salt-sensitive hypertension. The mice were treated with varying doses of phillyrin, and their blood pressure, cardiac function, cardiac hypertrophy, fibrosis, inflammation, and other conditions were assessed. Key findings: Our research findings demonstrated that phillyrin has the potential to lower blood pressure, enhance cardiac function, and mitigate cardiac hypertrophy, fibrosis, and inflammatory responses in deoxycorticosterone acetate-salt hypertension mice. In hypertensive mice, there was an elevated expression of endothelin1 (ET-1) in heart tissue, which can be reduced by phillyrin. Additionally, phillyrin effectively reduced the hypertrophy of H9c2 cells induced by ET-1 stimulation. Conclusions: Our research highlights the therapeutic capabilities of phillyrin in the treatment of myocardial remodeling through the reduction of ET-1 signaling. These results contribute to the advancement of novel applications for phillyrin and establish a solid conceptual basis for future investigations in this area. [ABSTRACT FROM AUTHOR]

Abstract Background Intervertebral disc degeneration (IDD) is an increasingly important cause of low back pain (LBP) that results in substantial health and economic burdens. Inflammatory pathway activation and the production of reactive oxygen species (ROS) play vital roles in the progression of IDD. Several studies have suggested that phillyrin has a protective role and inhibits inflammation and the production of ROS. However, the role of phillyrin in IDD has not been confirmed. Purpose The purpose of this study was to investigate the role of phillyrin in IDD and its mechanisms. Study design To establish IDD models in vivo, ex-vivo, and in vitro to verify the function of phillyrin in IDD. Method The effects of phillyrin on extracellular matrix (ECM) degeneration, inflammation, and oxidation in nucleus pulposus (NP) cells were assessed using immunoblotting and immunofluorescence analysis. Additionally, the impact of phillyrin administration on acupuncture-mediated intervertebral disc degeneration (IDD) in rats was evaluated using various techniques such as MRI, HE staining, S-O staining, and immunohistochemistry (IHC). Result Pretreatment with phillyrin significantly inhibited the IL-1β-mediated reduction in the degeneration of ECM and apoptosis by alleviating activation of the NF-κB inflammatory pathway and the generation of ROS. In addition, in vivo and ex-vivo experiments verified the protective effect of phillyrin against IDD. Conclusion Phillyrin can attenuate the progression of IDD by reducing ROS production and activating inflammatory pathways.

Background: Intervertebral disc degeneration (IDD) is an increasingly important cause of low back pain (LBP) that results in substantial health and economic burdens. Inflammatory pathway activation and the production of reactive oxygen species (ROS) play vital roles in the progression of IDD. Several studies have suggested that phillyrin has a protective role and inhibits inflammation and the production of ROS. However, the role of phillyrin in IDD has not been confirmed. Purpose: The purpose of this study was to investigate the role of phillyrin in IDD and its mechanisms. Study design: To establish IDD models in vivo, ex-vivo, and in vitro to verify the function of phillyrin in IDD. Method: The effects of phillyrin on extracellular matrix (ECM) degeneration, inflammation, and oxidation in nucleus pulposus (NP) cells were assessed using immunoblotting and immunofluorescence analysis. Additionally, the impact of phillyrin administration on acupuncture-mediated intervertebral disc degeneration (IDD) in rats was evaluated using various techniques such as MRI, HE staining, S-O staining, and immunohistochemistry (IHC). Result: Pretreatment with phillyrin significantly inhibited the IL-1β-mediated reduction in the degeneration of ECM and apoptosis by alleviating activation of the NF-κB inflammatory pathway and the generation of ROS. In addition, in vivo and ex-vivo experiments verified the protective effect of phillyrin against IDD. Conclusion: Phillyrin can attenuate the progression of IDD by reducing ROS production and activating inflammatory pathways. [ABSTRACT FROM AUTHOR]

【Objective】This work aims to investigate the effects and mechanisms of crude extract of F. suspensa leaves tea and its main components, phillyrin and Forsythia glycoside A, on the proliferation and migration function of hepatocellular carcinoma LM3.【Method】 Water extraction method was used to extract green tea and black tea from Forsythia suspensa leaves tea, HPLC was used to detect the content of effective components such as phillyrin and Forsythia glycoside A in F. suspensa green tea and F. suspensa black tea. CCK8 experiment was used to explore the effects of crude extracts of two types of F. suspensa leaves tea, phillyrin and Forsythia glycoside A on the proliferation of hepatocellular carcinoma LM3. Two crude extracts of Forsythia suspensa leaves tea, phillyrin and Forsythia glycoside A, were used to culture hepatocellular carcinoma LM3. The migration of cells was observed at 24, 48, 72, and 96 h, and the effects of the two crude extracts of F. suspensa leaves tea, phillyrin and Forsythia glycoside A on the migration of hepatocellular carcinoma LM3 were detected through the cell scratch test. To reveal the molecular mechanism of crude extract of F. suspensa leaves tea and its main components phillyrin and Forsythia glycoside A affect the proliferation and migration of hepatocellular carcinoma. The expressions of proliferation and migration-related gene Ki-67, Erk, PI3K and mTOR were detected using RT-PCR technology.【Result】Both the green tea and black tea extracts of F. suspensa leaves tea, as well as phillyrin and Forsythia glycoside A, significantly inhibited the proliferation and migration of hepatocellular carcinoma LM3, while Forsythia glycoside A significantly inhibited the migration of hepatocellular carcinoma line LM3. Among them, the crude extract of F. suspensa green tea significantly inhibited the proliferation and migration of HCC at low, medium, and high concentrations when 24, 48, and 72 h. The inhibitory effect of the crude extract of F. suspensa black tea on the proliferation and migration of LM3 cells continued to increase with increasing concentration, and the best inhibitory effect was achieved at high concentration for 72 h. The analysis of RT-PCR results suggests that its main mechanism of action may be achieved by reducing the expression levels of proliferation and migration related genes such as Ki-67.【Conclusion】 In summary, the crude extracts of F. suspensa green tea and F. suspensa black tea, as well as phillyrin and Forsythia glycoside A, can inhibit the proliferation or migration of HCC LM3 by reducing the expression of Ki-67. [ABSTRACT FROM AUTHOR]

Context: Adipose triglyceride lipase (ATGL), a key enzyme responsible for lipolysis, catalyzes the first step of lipolysis and converts triglycerides to diacylglycerols and free fatty acids (FFA). Our previous work suggested that phillyrin treatment improves insulin resistance in HFD-fed mice, which was associated with ATGL inhibition. In this study, using docking simulation, we explored the binding pose of phillyrin and atglistatin (a mouse ATGL inhibitor) to ATGL in mouse. From the docking results, the interactions with Ser47 and Asp166 were speculated to have caused phillyrin to inhibit ATGL in mice. Further, molecular dynamics simulation of 100 ns and MM-GBSA were conducted for the protein–ligand complex, which indicated that the system was stable and that phillyrin displayed a better affinity to ATGL than did atglistatin throughout the simulation period. Moreover, the results of pharmacological validation were consistent with those of the in silico simulations. In summary, our study illustrates the potential of molecular docking to accurately predict the binding protein produced by AlphaFold and suggests that phillyrin is a potential small molecule that targets and inhibits ATGL enzymatic activity. Methods: The ATGL-predicted protein structure, verified by PROCHECK, was determined using AlphaFold. Molecular docking, molecular dynamics simulation, and prime molecular mechanic–generalized born surface area were performed using LigPrep, Desmond, and prime MM-GBSA modules of Schrödinger software release 2021-2, respectively. For pharmacological validation, immunoblotting was performed to assess ATGL protein expression. The fluorescence intensity and glycerol concentration were quantified to evaluate the efficiency of phillyrin in inhibiting ATGL. [ABSTRACT FROM AUTHOR]

Abstract Influenza is an acute viral respiratory illness with high morbidity rates worldwide. Excessive pulmonary inflammation is the main characteristic of lethal influenza A virus (IAV) infections. Therapeutic options for managing influenza are limited to vaccines and some antiviral medications. Phillyrin is one of the major bioactive components of the Chinese herbal medicine Forsythia suspensa, which has the functions of sterilization, heat clearing and detoxification. In this work, the effect and mechanism of phillyrin on H1N1 influenza (PR8)-induced pneumonia were investigated. We reported that phillyrin (15 mg/kg) treatment after viral challenge significantly improved the weight loss, ameliorated pulmonary inflammation and inhibited the accumulation of multiple cytokines and chemokines in bronchoalveolar lavage fluid on 7 days post infection (dpi). In vitro, phillyrin suppressed influenza viral replication (Matrixprotein and nucleoprotein messenger RNA level) and reduced influenza virus-induced cytopathic effect (CPE). Furthermore,chemokine receptor CXCR2 was confirmed to be markedly inhibited by phillyrin. Surface plasmon resonance results reveal that phillyrin exhibits binding affinity to CXCR2, having a binding affinity constant (KD) value of 1.858e-5 M, suggesting that CXCR2 is a potential therapeutic target for phillyrin. Moreover, phillyrin inhibited the mRNA and protein expression levels of Caspase1, ASC and NLRP3 in the lungs of mice with H1N1-induced pneumonia.This study reveals that phillyrin ameliorates IAV-induced pulmonary inflammation by antagonizing CXCR2 and inhibiting NLRP3 inflammasome activation partly.

Objective To investigate the impacts of phillyrin on exudates and lung injury in rats with acute pleurisy by regulating the NLRP3 inflammatory pathway. Methods Ninety rats were randomly divided into the control group, the model group, the low-dose phillyrin (PH-L, 5 mg/kg) group, the medium-dose phillyrin (PH-M, 10 mg/kg) group, the high-dose phillyrin (PH-H, 20 mg/kg) group and the NLRP3 pathway inhibitor (PJ34, 10 mg/kg) group. FVC, FEV 0.1 and FEV 0.3 were detected by lung function analyzer. Electronic balance was used to weigh the mass of chest exudate. The number of white blood cells in exudate was detected by Wright staining. Contents of prostaglandin E2 (PGE2), monocyte chemoattractant protein-1 (MCP-1), interleukin (IL) -6 and tumor necrosis factor-α (TNF-α) in exudate were detected by ELISA. Automatic blood gas analyzer was used to detect p (CO2) and p (O2) of rats. HE staining was used to observe pathological changes of lung tissue. The expression levels of NLRP3 and Caspase-1 protein were detected by immunohistochemistry. Western blot assay was used to detect the expression of NLRP3 pathway protein. Results Compared with the control group, the quality of pleural exudate and the number of white blood cells, the contents of PGE2, MCP-1, IL-6, TNF-α, the expression of p (CO2) and NLRP3 pathway proteins in exudate of the model group increased obviously, FVC, FEV 0.1, FEV 0.3 and p (O2) decreased obviously, and the lung tissue showed obvious pathological damage (P＜0.05). Compared with the model group, the quality of pleural exudate and the number of white blood cells, the contents of PGE2, MCP-1, IL-6, TNF- α, the expression of p (CO2), NLRP3 pathway proteins in the exudate of rats decreased obviously in the PH group and the PJ34 group, FVC, FEV 0.1, FEV 0.3 and p (O2) increased obviously, the pathological injury of lung tissue was obviously improved (P＜0.05). Compared with the PH-H group, there were no significant differences in the above indexes in the PJ34 group (P＞0.05). Conclusion PH can improve lung injury induced by acute pleurisy in rats by inhibiting the activation of NLRP3 pathway and inhibiting inflammatory reaction. [ABSTRACT FROM AUTHOR]

Phillyrin is isolated from the fruit of Forsythia suspensa, and exhibits multiple pharmacological effects, including anti-tumor, antiinflammation, and anti-oxidation activities. Here, we investigated whether phillyrin could alleviate airway hyperresponsiveness (AHR) and eosinophil infiltration in the lungs of asthmatic mice, and mitigate inflammatory responses in tracheal epithelial BEAS-2B cells. IL-4/TNF-a-stimulated BEAS-2B cells were treated with various phillyrin doses. Female BALB/c mice were sensitised and challenged with ovalbumin (OVA), and then treated with intraperitoneal injection different phillyrin doses. In IL-4/TNF-a-stimulated BEAS-2B cells, phillyrin effectively reduced proinflammatory cytokines, chemokines, and eotaxin (CCL11) levels. In the lungs of asthmatic mice, phillyrin treatment relieved AHR, airway inflammation, eosinophil infiltration, and goblet cell hyperplasia. Phillyrin also reduced serum OVA-IgE, and Th2-associated cytokine levels in splenocyte culture medium, and in bronchoalveolar lavage fluid of asthmatic mice. Our results indicate that phillyrin attenuated airway inflammation and eosinophil infiltration in asthmatic mice by suppressing Th2 cytokine production. [ABSTRACT FROM AUTHOR]

Influenza is an acute viral respiratory illness with high morbidity rates worldwide. Excessive pulmonary inflammation is the main characteristic of lethal influenza A virus (IAV) infections. Therapeutic options for managing influenza are limited to vaccines and some antiviral medications. Phillyrin is one of the major bioactive components of the Chinese herbal medicine Forsythia suspensa, which has the functions of sterilization, heat clearing and detoxification. In this work, the effect and mechanism of phillyrin on H1N1 influenza (PR8)-induced pneumonia were investigated. We reported that phillyrin (15 mg/kg) treatment after viral challenge significantly improved the weight loss, ameliorated pulmonary inflammation and inhibited the accumulation of multiple cytokines and chemokines in bronchoalveolar lavage fluid on 7 days post infection (dpi). In vitro, phillyrin suppressed influenza viral replication (Matrixprotein and nucleoprotein messenger RNA level) and reduced influenza virus-induced cytopathic effect (CPE). Furthermore,chemokine receptor CXCR2 was confirmed to be markedly inhibited by phillyrin. Surface plasmon resonance results reveal that phillyrin exhibits binding affinity to CXCR2, having a binding affinity constant (KD) value of 1.858e-5 M, suggesting that CXCR2 is a potential therapeutic target for phillyrin. Moreover, phillyrin inhibited the mRNA and protein expression levels of Caspase1, ASC and NLRP3 in the lungs of mice with H1N1-induced pneumonia.This study reveals that phillyrin ameliorates IAV-induced pulmonary inflammation by antagonizing CXCR2 and inhibiting NLRP3 inflammasome activation partly. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of phillyrin (PHN) on dexamethasone (DEX)-induced autophagy and apoptosis of osteoblasts by regulating phosphatidylinositol 3-kinase (PI3K)/serine threonine kinase (AKT)/mammalian target of rapamycin (mTOR) signaling pathway. Methods MC3T3-E1 cells were divided into control group (NOR group), DEX group (10 μmol/L DEX treated MC3T3-E1 cells), L-PHN group (5 μmol/L PHN treated MC3T3-E1 cells), M-PHN group (10 μmol/L PHN treated MC3T3-E1 cells), H-PHN group (20 μmol/L PHN treated MC3T3-E1 cells), and ZSTK474 group (treated with 20 μmol/L PHN and 2 μmol/L ZSTK474, an inhibitor of PI3K/AKT/mTOR signaling pathway). MTT assay was used to detect cytotoxicity and cell viability. The apoptosis of MC3T3-E1 cells was detected with flow cytometry. The autophagosomes were observed with transmission electron microscopy (TEM). Western blotting was used to detect the expression of autophagy, apoptosis, and PI3K/AKT/mTOR pathway related proteins in MC3T3-E1 cells. ALP activity and ALP staining were used to detect the differentiation ability of MC3T3-E1 cells. Results 0-80 μmol/L PHN had no obvious toxic effect on MC3T3-E1 cells. Compared to those in NOR group, the OD570 value, Bcl-2 protein level, p-PI3K/PI3K, p-AKT/AKT, mTOR protein levels, ALP activity, Beclin1, LC3-II/I protein levels, and the number of autophagosomes in DEX group decreased significantly (P<0.05), and the apoptosis rate, Bax, and cleaved Casase-3 increased significantly (P<0.05). Compared to that in DEX group, the level of apoptosis in L-PHN group, M-PHN group, and H-PHN group decreased significantly (P<0.05), and the proliferation activity, autophagy level, osteogenic differentiation ability and pathway protein levels increased significantly (P<0.05). ZSTK474 eliminated the beneficial effects of PHN on MC3T3-E1 cells. Conclusion PHN promotes autophagy of osteoblasts by activating PI3K/AKT/mTOR signaling pathway, and then inhibits apoptosis of osteoblasts. [ABSTRACT FROM AUTHOR]

The discovery of alternative medicines with less adverse effects is extremely urgent for rheumatoid arthritis (RA). Phillyrin (Phil), the predominant lignan glycoside of Forsythia suspensa, has been reported to exert several pharmacological effects, such as antivirus, anti-inflammation, anti-oxidation, anti-obesity, and antipyretic activity. However, the effect of Phil on RA remains unknown. In this study, We utilized both in vivo collagen-induced arthritis (CIA) rat models and in vitro TNFα-induced fibroblast-like synoviocytes (FLSs) to study the inhibitory effects of Phil on RA. The in vivo studies revealed that Phil treatment effectively ameliorated synovial inflammation and bone erosion in CIA rats. The in vitro studies demonstrated that Phil could significantly suppress the proliferation and migration of arthritic FLSs. In addition, treatment with Phil resulted in decreased mRNA expression of proinflammatory cytokines including TNFα, IL-1β, IL-6, IL-8 and MMP9. Molecular mechanistic investigations revealed that the suppressive effects of Phil were mediated by blockade of the MAPK (ERK, p38, and JNK) and NF-κB pathways. Taken together, our findings suggest that Phil has an anti-arthritic effect in CIA rats by inhibiting the pathogenic characteristic of arthritic FLSs throught suppression of NF-κB and MAPKs signaling pathways. These results demonstrate the potential of Phil as a novel therapeutic agent for the treatment of RA.

Objective: To investigate the effects of phillyrin on D-galactose (D-gal)-induced aging in the brain of mice and mechanism. Methods: Institute of Cancer Research mice were intraperitoneally injected with D-gal 150 mg/kg to induce the aging model, and the mice were treated with phillyrin 5, 15, and 45 mg/kg once a day for 8 weeks. Results: The memory impairment induced by D-gal in brain aging was ameliorated by phillyrin treatment, and the oxidative stress in the aging brain and serum was inhibited, as evidenced by increased superoxide dismutase, glutathione peroxidase enzyme activities, and decreased malondialdehyde levels. Additionally, the hippocampus damage has also been well improved by phillyrin. Moreover, the protein expressions of nuclear factor erythroid-2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NADPH quinone oxidoreductase 1 (NQO1) were markedly up-regulated and the expression of Keap-1 was down-regulated in the phillyrin group than that in the aging model group. Conclusion: These findings suggest that phillyrin is effective in the prevention of age-related memory impairment, and the attenuation oxidative stress by activating the Nrf2/HO-1 signaling pathway in the hippocampus may be one of the mechanisms. [ABSTRACT FROM AUTHOR]

Background: Influenza virus causes significant morbidity and mortality with pandemic threat. Oleaceae Fructus Forsythiae is a medicinal herb. This study aimed to investigate antiviral effect of Phillyrin, a purified bioactive compound from this herb, and its reformulated preparation FS21 against influenza and its mechanism. Methods: Madin–Darby Canine Kidney (MDCK) cells were infected by one of six influenza viruses: five influenza A viruses (IAVs: three H1N1 and two H3N2) and one influenza B virus (IBV). Virus‐induced cytopathic effects were observed and recorded under microscope. Viral replication and mRNA transcription were evaluated by quantitative polymerase chain reaction (qPCR) and protein expression by Western blot. Infectious virus production was assessed using TCID50 assay, and IC50 was calculated accordingly. Pretreatment and time‐of‐addition experiments with Phillyrin or FS21 added 1 h before or in early (0–3 h), mid (3–6 h), or late (6–9 h) stages of viral infection were performed to assess their antiviral effects. Mechanistic studies included hemagglutination and neuraminidase inhibition, viral binding and entry, endosomal acidification, and plasmid‐based influenza RNA polymerase activity. Results: Phillyrin and FS21 had potent antiviral effects against all six IAV and IBV in a dose‐dependent manner. Mechanistic studies showed that both suppressed influenza viral RNA polymerase with no effect on virus‐mediated hemagglutination inhibition, viral binding or entry, endosomal acidification, or neuraminidase activity. Conclusions: Phillyrin and FS21 have broad and potent antiviral effects against influenza viruses with inhibition of viral RNA polymerase as the distinct antiviral mechanism. [ABSTRACT FROM AUTHOR]

Cardiac hypertrophy (CH) is the main compensatory response to chronic heart stress and often progresses to a decompensation state potentially leading to heart failure. Phillyrin (PHI) is a novel compound derived from Forsythia, which has shown anti-inflammatory and anti-virus activities as well as renal protective effects on diabetic nephropathy. Therefore, we investigated the effects of PHI on CH induced by isoproterenol (ISO). Cardiac hypertrophy was induced by ISO in vivo, and the H9C2 cells were treated with ISO. PHI treatment alleviated CH in isoproterenol-induced mice in 7 and 14 days. Echocardiography showed that the PHI improved ISO-induced CH heart function and structure. PHI significantly decreased heart weight/body weight (HW/BW) and heart weight/tibia length (HW/TL) ratios and improved left ventricular (LV) function in ISO-treated mice. Hematoxylin and eosin staining revealed cardiomyocyte areas of the ISO group were significantly increased, and PHI was significantly reduced at 7 and 14 days, PHI-100 groups showed significantly better improvements than PHI-50. Sirius red staining indicated PHI significantly decreased collagen deposition in heart cross-sections induced by ISO, and PHI repressed ISO-induced cTn-I and NT-proBNP expression in mouse serum. In vitro data from H9C2 cells showed that PHI decreased cell areas and total cell protein levels in cells induced by ISO, whereas ANP, BNP, IL-6, and IL-1β expression was significantly inhibited by PHI. Also, PHI simultaneously inhibited P65 and P38 phosphorylation in vivo and in vitro. In conclusion, this study demonstrated the protective effect of PHI on CH in in vivo and in vitro, and this effect was related to the suppression of inflammation through the activation of the P38/NF-κB pathway. [ABSTRACT FROM AUTHOR]

Taste masking of traditional Chinese medicines (TCMs) containing multiple bitter components remains an important challenge. In this study, berberine (BER) in alkaloids and phillyrin (PHI) in flavonoid glycosides, which are common bitter components in traditional Chinese medicines, were selected as model drugs. Chitosan (CS) was used to mask their unfriendly taste. Firstly, from the molecular level, we explained the taste-masking mechanism of CS on those two bitter components in detail. Based on those taste-masking mechanisms, the bitter taste of a mixture of BER and PHI was easily masked by CS in this work. The physicochemical characterization results showed the taste-masking compounds formed by CS with BER (named as BER/CS) and PHI (named as PHI/CS) were uneven in appearance. The drug binding efficiency of BER/CS and PHI/CS was 50.15 ± 2.63% and 67.10 ± 2.52%, respectively. The results of DSC, XRD, FTIR and molecular simulation further indicated that CS mainly masks the bitter taste by disturbing the binding site of bitter drugs and bitter receptors in the oral cavity via forming hydrogen bonds between its hydroxyl or amine groups and the nucleophilic groups of BER and PHI. The taste-masking evaluation results by the electronic tongue test confirmed the excellent taste-masking effects on alkaloids, flavonoid glycosides or a mixture of the two kinds of bitter components. The in vitro release as well as in vivo pharmacokinetic results suggested that the taste-masked compounds in this work could achieve rapid drug release in the gastric acid environment and did not influence the in vivo pharmacokinetic results of the drug. The taste-masking method in this work may have potential for the taste masking of traditional Chinese medicine compounds containing multiple bitter components. [ABSTRACT FROM AUTHOR]

Forsythia suspensa (Thunb.) has been widely used in traditional medicines in Asia. According to the 2020 edition of Chinese Pharmacopoeia, phillyrin is the main active ingredient in F. suspensa, which is effective in clearing heat, reducing swelling, and dispersing nodules. F. suspensa leaf is a non-toxic substance and it can be used to make a health tea. Here, we combine elicitors and transcriptomics to investigate the inducible biosynthesis of the phillyrin from the F. suspensa. After the fruits and leaves of F. suspensa were treated with different concentrations of methyl jasmonate (MeJA), the content of phillyrin in the fruits reached a peak at 200 µM MeJA for 12 h, but which was decreased in leaves. To analyze the differences in key enzyme genes involved in the phillyrin biosynthesis, we sequenced the transcriptome of F. suspensa leaves and fruits treated with 200 µM MeJA for 12 h. We hypothesized that nine genes related to coniferin synthesis including: F. suspensa UDP-glycosyltransferase (FsUGT); F. suspensa 4-coumarate coenzyme CoA ligase (Fs4CL); and F. suspensa Caffeoyl-CoA O-methyltransferase (FsCCoAOMT) etc. The qRT-PCR analysis of genes related to phillyrin biosynthesis was consistent with RNA-seq analysis. We also investigated the dynamic changes of genes in F. suspensa leaves and fruits at different time points after 200 µM MeJA treatment, which laid the foundation for further study of the molecular mechanisms regulating the biosynthesis of phillyrin. [ABSTRACT FROM AUTHOR]

The function of white adipose tissue as an energy reservoir is impaired in obesity, leading to lipid spillover and ectopic lipid deposition. Adipose tissue inflammation can reduce the efficacy of lipid storage in adipocytes by augmenting basal lipolysis through producing interleukin-6 (IL-6). Therefore, pharmacological compounds targeting adipose tissue inflammation or IL-6 signaling might have the potential to combat obesity. This study aims to investigate the impact of Phillyrin, which is frequently used for treating respiratory infections in clinics in China, on obesity-related metabolic dysfunctions. Firstly, a mouse model of diet-induced obesity is used to assess the pharmacological applications of Phillyrin on obesity in vivo. Secondly, ex vivo culture of adipose tissue explants is utilized to investigate actions of Phillyrin on IL-6-linked basal lipolysis. Thirdly, a mouse model of IL-6 injection into visceral adipose tissue is explored to confirm the anti-basal lipolytic effect of Phillyrin against IL-6 in vivo. The results show that Phillyrin treatment reduces circulating level of glycerol, decreases hepatic steatosis and improves insulin sensitivity in obese mice. Meanwhile, Phillyrin attenuates obesity-related inflammation and IL-6 production in adipose tissue in obese mice. Furthermore, Phillyrin treatment results in resistance to IL-6-induced basal lipolysis in adipose tissue through suppressing expression of adipose triglyceride lipase (ATGL) both in vivo and in vitro. Collectively, these findings suggest that Phillyrin can restrain lipid efflux from inflamed adipose tissue in obesity by inhibiting IL-6-initiated basal lipolysis and ATGL expression, and thus is a potential candidate in the treatment of obesity-associated complications.

• The treatment of osteoarthritis is mainly based on palliative care. • Before the experiment, network pharmacology and molecular docking were used to predict the pathway and target of phillyrin. • Phillyrin inhibits IL-1β-induced extracellular matrix degradation. • Phillyrin inhibits osteoarthritis by regulating nrf-2/NF-κB. • Phillyrin can be used as a potential treatment for osteoarthritis. Osteoarthritis (OA) is the predominant cause of disability among elderly people worldwide and is characterized by cartilage degeneration and excessive bone formation. Phillyrin, derived from forsythia, is a key extract renowned for its pronounced antibacterial and anti-inflammatory effects. Forsythia, deeply integrated into traditional Oriental medicine, has historically been utilized for its various pharmacological effects, including antibacterial, anti-inflammatory, and hepato-protective properties. Nevertheless, the anti-inflammatory impact of phillyrin on the progression of osteoarthritis remains enigmatic. The objective of this research was to assess the anti-inflammatory and anti-aging properties of phillyrin in mouse chondrocytes induced by IL-1β, as well as to elucidate the fundamental mechanisms underlying the phenomenon at play. Additionally, the investigation extends to observing the impact of phillyrin by establishing a murine osteoarthritic model. The ultimate goal was to identify phillyrin as a potential antiosteoarthritic agent. This investigation employs a multifaceted approach. Initially, key action targets of phillyrin, along with its probable action pathways, were identified by molecular docking and network pharmacological techniques. These findings were subsequently confirmed through both in vivo and in vitro studies. Network pharmacological analysis revealed NFE2L2 (NRF2), NFKB1, TLR4, and SERPING1 as pivotal candidate targets for the treatment of osteoarthritis with phillyrin. Molecular docking revealed hydrogen bond interactions between phillyrin and Arg415, Arg483, Ser508, and Asn387 on the Nrf2 receptor, while electrostatic interactions occurred with residues Arg415 and Arg380. Experiments conducted in vitro indicated that phillyrin preconditioning hindered the IL-1β-induced expression of proinflammatory factors which included TNF-α, COX-2, IL-6, and iNOS. Furthermore, phillyrin counteracts the IL-1β-induced degradation of aggrecan and collagen II within the extracellular matrix (ECM). This protective action is caused by the inhibition of the NF-κB pathway by phillyrin. Additionally, the mitigation of chondrocyte aging by phillyrin was observed. Our investigation revealed that phillyrin mitigates inflammation and counteracts cartilage degeneration in osteoarthritis (OA) patients by suppressing inflammation in chondrocytes and impeding aging through suppression of the NF-κB pathway. [ABSTRACT FROM AUTHOR]

• Regulating GAPDH, MDH2 and IDH2, the key proteins in energy metabolism, can exert antipyretic effects by reducing energy supply and regulating febrile related inflammatory factors. • Phr have a significant antipyretic effect. • The antipyretic effect of Phr is related to its metabolites Phg and Phg-S targeting the NAD+ binding domain of GAPDH, IDH2 and MDH2 and inhibiting the enzyme activity of them. Fever is one of the main pathophysiological reactions that occurs during the acute phase of various diseases. Excessive body temperature can lead to various adverse consequences such as brain tissue damage and abnormal immune responses. Phillyrin (Phr) is the main active ingredient in Forsythia suspensa (Thunb.) Vahl (Lian Qiao) and has antipyretic effects; however, its antipyretic mechanism of action remains unclear. This study aimed to explore the antipyretic mechanisms of Phr and provide a new treatment plan for fever. The antipyretic effects of Phr were evaluated using a mouse model of pneumonia fever. The main metabolites of Phr involved in its antipyretic function were identified using a mitochondrial temperature-sensitive probe. Further synthesis of the main metabolite, phillygenin (Phg), an alkynylated probe, was performed, and chemical proteomics was used to capture and analyze its direct target for antipyretic effects. The mechanism of action of Phg and its antipyretic targets was explored using metabolomics and various molecular biology methods. Phr showed significant antipyretic and anti-inflammatory effects in a mouse model of lipopolysaccharide-induced fever. Phg reversibly targeted the nicotinamide adenine dinucleotide (NAD+) binding domain of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), malate dehydrogenase 2 (MDH2), and isocitrate dehydrogenase 2 (IDH2) to inhibit their enzymatic activity. In-depth analysis of cellular metabolomics and mitochondrial stress testing indicated that inhibition of GAPDH, MDH2, and IDH2 enzyme activity by Phg led to a decrease in cellular energy supply and heat production regulated by glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation signaling pathways. Phg specifically targeted macrophages and inhibited LPS-induced macrophage activation by downregulating GAPDH enzyme activity, thereby exerting anti-inflammatory effects. In vivo experiments also confirmed that the antipyretic effect of Phr in LPS-induced fever model mice was related to its main metabolites, Phg and Phg-sulfonate (Phg-S), which directly targeted the NAD+ binding domain of GAPDH, IDH2, and MDH2, inhibiting the activity of these enzymes, thereby reducing energy supply and regulating febrile-related inflammatory factors. This study reported for the first time that the antipyretic effect of Phr is produced by targeting GAPDH, IDH2, and MDH2 to regulate energy supply and febrile-related inflammatory factors through its main metabolites Phg and Phg-S. This study not only provides potential drugs for fever treatment but also provides new ideas for improving clinical fever treatment plans. The antipyretic mechanism of Phillyrin (Phr) is related to its main metabolite Phillygenin (Phg) targeting the NAD+ binding domain of GAPDH, MDH2 and IDH2, reducing energy supply and regulating fever related inflammatory factors. [Display omitted] [ABSTRACT FROM AUTHOR]

The preservation of autophagosome formation presents a promising strategy for tackling neurological disorders, such as Parkinson's disease (PD). Mitochondria-associated endoplasmic reticulum (ER) membranes (MAM) serve not only as a focal point linked to various neurological disorders but also play a crucial role in supporting the biogenesis of autophagosomes. This investigation aimed to elucidate the neuroprotective properties of phillyrin against PD and its underlying mechanisms in promoting autophagosome formation. ER and mitochondria co-localization was assessed via fluorescent staining. Annexin V-fluorescein isothiocyanate (FITC) fluorescence was employed to quantify accessible cardiolipin (CL) on mitochondrial surfaces. The levels of CL within the MAM fraction of SH-SY5Y cells were evaluated using a CL probe assay kit. Monodansylcadaverine staining was utilized to detect autophagosome formation in SH-SY5Y cells. In an A53T-alpha-synuclein (αSyn)-induced PD mouse model, the anti-PD properties of phillyrin were assessed using open field, pole climbing, and rotarod tests, as well as immunohistochemistry staining of TH+ neurons in the brain sections. In A53T-αSyn-treated SH-SY5Y cells, phillyrin facilitated autophagosome formation by suppressing CL externalization and restoring MAM integrity. Phillyrin enhanced the localization of receptor expression-enhancing protein 1 (REEP1) within MAM and mitochondria, bolstering MAM formation. Increased REEP1 levels in mitochondria, attributed to phillyrin, enhanced the interaction between REEP1 and NDPK-D, thereby reducing CL externalization. Furthermore, phillyrin exhibited a dose-dependent enhancement of motor function in mice, accompanied by an increase in the abundance of dopaminergic neurons within the substantia nigra. These findings illuminate phillyrin's ability to enhance MAM formation through upregulation of REEP1 expression within MAM, while concurrently attenuating CL externalization via the REEP1-NDPK-D interaction. These mechanisms bolster autophagosome biogenesis, offering resilience against A53T-αSyn-induced PD. Thus, our study advances the understanding of phillyrin's complex mechanisms and underscores its potential as a therapeutic approach for PD, opening new avenues in natural product pharmacology. [Display omitted] [ABSTRACT FROM AUTHOR]

Lian Qiao (LQ), the dried fruit of Forsythia suspensa (Thunb.) Vahl, is a well-documented traditional Chinese medicine known for its detoxifying and heat-clearing properties. Clinically, compounds containing LQ are widely used to treat thrombotic diseases, indicating that it may have antithrombotic effects. However, its exact mechanism of action remains unknown. This study aimed to verify the antithrombotic effect of LQ and further explore the material basis and target mechanism of its antithrombotic effect using various biological methods. An epinephrine-collagen-thrombin-induced mouse model of acute pulmonary embolism (APE) was established to study the effects of LQ on thrombus development. A UPLC/Q/TOF-MS screening and identification system based on the inhibition of platelet aggregation and Ca2+ antagonism was established to determine the pharmacodynamic components of LQ that inhibit platelet activation. The inhibitory effect of active ingredients on platelet activation, and the determination of the target of their inhibitory effect on platelet activation have been studied using chemical proteomics. Furthermore, based on the structure and function of the target protein, a multidisciplinary approach was adopted to analyze the molecular mechanism of active ingredient binding to target proteins and to evaluate the effects of active ingredients on the downstream signaling pathways of target proteins. LQ showed significant anticoagulant effects in APE model mice. Phillyrin and phillygenin were the antiplatelet-activating components of LQ. PLCβ3 was identified as a target for inhibiting platelet activation by phillyrin and its metabolites. The mechanism underlying the effect involves phillyrin and its metabolites inhibiting PLCβ3 activity by blocking the binding of PLCβ3 to Gαq through non-covalently targeting the ASN260 of PLCβ3, thus inhibiting the downstream Gαq-PLCβ3-Ca2+ signaling pathway, effectively hindering platelet activation and therefore playing an anticoagulant role. This study not only proposes and validates the antithrombotic effect of LQ for the first time but also finds that phillyrin and phillygenin are the main pharmacological substances through which LQ exerts antithrombotic activity and reveals a novel mechanism by which they exert antiplatelet activity by directly targeting and inhibiting PLCβ3 activity. These findings significantly contribute to our understanding of the therapeutic potential of phillyrin and provide important clues for the discovery and development of new antiplatelet drugs. Phillyrin (Phr) and its metabolites (Phg, Phg-G and Phg-S) block the binding of PLCβ3 to Gαq through non-covalently targeting the ASN260 of PLCβ3, thus inhibiting the downstream Gαq-PLCβ3-Ca2+ signaling pathway, thereby exerting antiplatelet and antithrombotic effect. [Display omitted] • Phillyrin and its metabolites deprives Gαq-PLCβ3 interaction by non-covalently and selectively targeting the ASN260 of PLCβ3. • Phillyrin and its metabolites exerts antiplatelet and antithrombotic effects by inhibiting Gαq-PLCβ3-Ca2+ signaling pathway. • Pharmacologically targeting PLCβ3 is a promising therapeutic strategy for thrombosis. [ABSTRACT FROM AUTHOR]

Phillyrin is extracted from Forsythia suspensa (Thunb.) Vahl, is significantly higher in (unripe Forsythiae Fructus) Qing qiao than in (ripe Forsythiae Fructus) Lao qiao fruits of the plant. However, the toxicity of phillyrin has not been adequately investigated. The study investigates the genetic and teratogenic effects of phillyrin to determine its safety profile. Assessing the genotoxicity and teratogenicity of phillyrin involved various tests, such as the bacterial reverse mutation assay, mammalian erythrocyte micronucleus assay, spermatocyte chromosome aberration assay, and teratogenicity assay. The results demonstrated that phillyrin exhibited no discernible impact on the following: number of colonies that spontaneously revert for Salmonella typhimurium TA 97, TA98, TA100, TA102, and TA1535, frequency of bone marrow polychromatic erythrocytes, and the rate of chromosomal aberrations. In the teratogenicity test, the pregnant rats exhibited no signs of toxicity or abnormal changes, and the growth, embryonic development, and visual anatomy of each pup were normal. In comparison with the negative control group, there were no significant differences in fetal body weight, mortality, deformity rate, malformed nest rate, gravid uterus weight, average number of fetuses per litter, fetal body length, or visceral and skeletal development in each dose group. In conclusion, these findings provide evidence that phillyrin does not exhibit genotoxic or teratogenic effects, supporting its potential safety for pharmacological applications. [Display omitted] • This study conducts the safety assessment of the key compound phillyrin in Forsythia. • Employed rigorous toxicological protocols to establish safety profile of Phillyrin. • Phillyrin is not genotoxic or teratogenic at the doses indicated. [ABSTRACT FROM AUTHOR]

Phillyrin is an effective lignan glycoside extracted from a traditional Chinese medicine Forsythia suspensa (Thunb.) Vahl (Oleaceae). It mainly exists in the roots, stems, leaves and fruits of the plant, with the highest content in the leaves. In terms of its medicinal application, there are a large number of experimental data proving its pharmacological effects in vitro and in animal models, such as anti-inflammatory, anti-obesity, anti-tumor, etc. Furthermore, pharmacokinetic experiments have also shown phillyrin's high effectiveness and low toxicity. Despite more than one thousand studies in the literature on phillyrin retrievable from Web of Science, PubMed, and CNKI, few reviews on its pharmacological activities have been presented conclusively. In this paper, we aimed to summarize the pharmacological and pharmacokinetic characteristics of phillyrin from the current literature, focusing on its anti-inflammatory, anti-aging, antiviral, antibacterial, hepatoprotective and anti-cancer effects, hoping to come up with new insights for its application as well as future studies. [ABSTRACT FROM AUTHOR]

Copyright of Modern Food Science & Technology is the property of Editorial Office of Modern Food Science & Technology and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Despite advancements and refinements in the therapeutic approaches for hepatic malignancies, liver cancer remains a prevalent and deadly form of cancer, with its grim outlook posing as a significant clinical challenge. Phillyrin (PHN) has been reported to have anticancer effects, but the anticancer mechanism in liver cancer is ominous. By searching the potential target of PHN in the online database and liver cancer disease database, it was found that there is only one overlap gene, and DNA topoisomerase II alpha (TOP2A) is abnormally expressed in liver cancer tissues. TOP2A overexpression and downregulated hepatocellular carcinoma cell lines were then constructed in vitro , and it was examined whether PHN treatment induced ferroptosis in hepatocellular carcinoma by regulating TOP2A's inhibition of Janus kinase 2/Signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway through phenotypic assay, western blot assay, reverse transcription‑quantitative PCR assay and electron microscopy. The results showed that PHN could inhibit the expression of TOP2A protein and JAK2/STAT3 signaling pathway in hepatoma cells. PHN could also downregulate glutathione peroxidase 4 by suppressing the expression of TOP2A protein. PHN impeded the activity of factor inhibiting hypoxia‑inducible factor 1 alpha, thereby augmenting the synthesis of iron‑dependent apoptosis‑related proteins including cytochrome c oxidase subunit II, long‑chain acyl‑CoA synthetase family member 4 and NADPH oxidase 1, thus facilitating an increase in Fe 2+ concentration and accelerating oxidative harm within hepatocellular carcinoma cells, culminating in the induction of ferroptotic cell death in these liver malignancy cells.

Background: Fever is one of the main pathophysiological reactions that occurs during the acute phase of various diseases. Excessive body temperature can lead to various adverse consequences such as brain tissue damage and abnormal immune responses. Phillyrin (Phr) is the main active ingredient in Forsythia suspensa (Thunb.) Vahl (Lian Qiao) and has antipyretic effects; however, its antipyretic mechanism of action remains unclear., Purpose: This study aimed to explore the antipyretic mechanisms of Phr and provide a new treatment plan for fever., Methods: The antipyretic effects of Phr were evaluated using a mouse model of pneumonia fever. The main metabolites of Phr involved in its antipyretic function were identified using a mitochondrial temperature-sensitive probe. Further synthesis of the main metabolite, phillygenin (Phg), an alkynylated probe, was performed, and chemical proteomics was used to capture and analyze its direct target for antipyretic effects. The mechanism of action of Phg and its antipyretic targets was explored using metabolomics and various molecular biology methods., Results: Phr showed significant antipyretic and anti-inflammatory effects in a mouse model of lipopolysaccharide-induced fever. Phg reversibly targeted the nicotinamide adenine dinucleotide (NAD + ) binding domain of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), malate dehydrogenase 2 (MDH2), and isocitrate dehydrogenase 2 (IDH2) to inhibit their enzymatic activity. In-depth analysis of cellular metabolomics and mitochondrial stress testing indicated that inhibition of GAPDH, MDH2, and IDH2 enzyme activity by Phg led to a decrease in cellular energy supply and heat production regulated by glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation signaling pathways. Phg specifically targeted macrophages and inhibited LPS-induced macrophage activation by downregulating GAPDH enzyme activity, thereby exerting anti-inflammatory effects. In vivo experiments also confirmed that the antipyretic effect of Phr in LPS-induced fever model mice was related to its main metabolites, Phg and Phg-sulfonate (Phg-S), which directly targeted the NAD + binding domain of GAPDH, IDH2, and MDH2, inhibiting the activity of these enzymes, thereby reducing energy supply and regulating febrile-related inflammatory factors., Conclusion: This study reported for the first time that the antipyretic effect of Phr is produced by targeting GAPDH, IDH2, and MDH2 to regulate energy supply and febrile-related inflammatory factors through its main metabolites Phg and Phg-S. This study not only provides potential drugs for fever treatment but also provides new ideas for improving clinical fever treatment plans., Competing Interests: Declaration of competing interest The authors declared that no conflicts of interest., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Acute lung injury (ALI) is a severe pulmonary disorder of sepsis with high clinical incidence and mortality. Nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3)-cysteinyl aspartate specific proteinase 1-gasdermin D (GSDMD)-dependent pyroptosis of alveolar epithelial cells (AECs) has emerged as a crucial contributor to ALI during sepsis. Phillyrin (PHI), a natural lignan isolated from the traditional Chinese herbal medicine Forsythia suspensa , has been shown to have anti-inflammatory, antioxidant and antiviral properties. However, little is known about the protective role and potential mechanism of PHI in sepsis-induced ALI, and it is uncertain whether the protective effect of PHI in sepsis-induced ALI is connected to pyroptosis. This study aims to examine the preventive effects of PHI on sepsis-induced ALI via the inhibition of NLRP3/caspase-1/GSDMD-mediated pyroptosis in AECs. Our findings demonstrate that preadministration of PHI successfully reduces sepsis-induced pulmonary edema, systemic/pulmonary inflammation, and pulmonary histological damage in lung tissues, bronchoalveolar lavage fluid, and the serum of septic mice. Intriguingly, PHI preadministration suppresses sepsis-induced protein expressions of pyroptosis-specific markers, especially their active forms. In vitro assays show that PHI pretreatment also protects type II AECs (MLE-12) from lipopolysaccharide-induced pyroptosis by preventing the activation of the pyroptosis signaling pathway. The results from molecular docking and surface plasmon resonance reveal that PHI has a significant affinity for direct binding to the GSDMD protein, suggesting that GSDMD is a potential pharmacological target for PHI. In conclusion, PHI can prevent sepsis-triggered ALI by effectively suppressing the activation of the canonical pyroptosis signaling pathway and pyroptosis of AECs.

Diabetic nephropathy is a progressive kidney disease resulting from long-term hyperglycaemia in diabetic patients, and the underlying mechanism is complex and lacks effective treatments. Various active ingredients in Chinese herbs have been shown to alleviate renal injury and improve DN in recent years. Phillyrin, a natural medicinal active compound extracted from the Oleaceae family, has various pharmacological effects, including antioxidative, antiapoptotic and antiobesity effects. However, the role of phillyrin and its underlying mechanism in DN have not yet been explored. To investigate the effects of phillyrin on DN and its potential mechanisms of action, we performed experiments using streptozotocin (STZ)-induced DN mice as models. Phillyrin significantly reduced the levels of fasting blood glucose (FBG) and glycosylated haemoglobin A1c (HbA1c), downregulated the levels of serum blood urea nitrogen (BUN), serum creatinine (Scr), serum and urine β2-microglobulins (β2-MG) and improved the pathological changes of the kidney in a DN mouse model. Phillyrin also increased the level of antioxidants and attenuated oxidative damage in DN model mice. In addition, phillyrin inhibited Glycogen synthase kinase-3β (GSK-3β) activity by activating the PI3K/Akt signalling pathway, increased the Bcl-2/Bax ratio, reduced the release of cytochrome c from the mitochondria to the cytoplasm, subsequently inhibited the activation of caspase-3 and ultimately suppressed renal cell apoptosis. These findings suggested that phillyrin could be a new promising therapeutic strategy for DN, and this protective effect might be related to suppressing oxidative stress and apoptosis via the PI3K/Akt/GSK-3β pathway. [ABSTRACT FROM AUTHOR]

Background: The risk of co-epidemic between COVID-19 and influenza is very high, so it is urgent to find a treatment strategy for the co-infection. Previous studies have shown that phillyrin can not only inhibit the replication of the two viruses, but also has a good anti-inflammatory effect, which is expected to become a candidate compound against COVID-19 and influenza.Objective: To explore the possibility of phillyrin as a candidate compound for the treatment of COVID-19 and influenza co-infection and to speculate its potential regulatory mechanism.Methods: We used a series of bioinformatics network pharmacology methods to understand and characterize the pharmacological targets, biological functions, and therapeutic mechanisms of phillyrin in COVID-19 and influenza co-infection and discover its therapeutic potential.Results: We revealed potential targets, biological processes, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and upstream pathway activity of phillyrin against COVID-19 and influenza co-infection. We constructed protein–protein interaction (PPI) network and identified 50 hub genes, such as MMP9, IL-2, VEGFA, AKT, and HIF-1A. Furthermore, our findings indicated that the treatment of phillyrin for COVID-19 and influenza co-infection was associated with immune balance and regulation of hypoxia-cytokine storm, including HIF-1 signaling pathway, PI3K-Akt signaling pathway, Ras signaling pathway, and T cell receptor signaling pathway.Conclusion: For the first time, we uncovered the potential targets and biological pathways of phillyrin for COVID-19 and influenza co-infection. These findings should solve the urgent problem of co-infection of COVID-19 and influenza that the world will face in the future, but clinical drug trials are needed for verification in the future.

Background: Phillyrin (Phi) is the main polyphenolic compound found in Forsythia suspensa. Recent studies have revealed that Phi has potent antioxidative and anti-inflammatory effects. However, whether Phi could relieve blood–brain barrier (BBB) damage following traumatic brain injury (TBI) remains unknown.Materials and Methods: Lipopolysaccharide (LPS) was used to activate primary microglia, which were then treated with different doses of Phi or the peroxisome proliferator–activated receptor-gamma (PPARγ) antagonist (GW9662). CCK-8 assay was used for evaluating cell viability, and the cytokines (including IL-1β, IL-6, TNFα, IL-4, IL-10, and TGFβ), microglial phenotypic markers (iNOS, COX2, and CD86 for “M1” polarization; Arg1, Ym1, and CD206 for “M2” polarization), PPARγ, and NF-κB were determined by RT-PCR, Western blot, or cellular immunofluorescence. Primary cultured mouse brain microvascular endothelial cells (BMECs) were stimulated by the condition medium (CM) from microglia. The cell viability, angiogenesis, and tight junction of BMECs were determined via CCK-8 assay, tube formation assay, and Western blot (for detecting MMP3, MMP9, ZO1, claudin-5, and occludin). Furthermore, the mouse TBI model was constructed and treated with Phi and/or GW9662. The BBB integrity was evaluated by H&E staining, Evans blue staining, and tissue immunofluorescence.Results: Phi markedly restrained the pro-inflammatory (“M1” state) cytokines and promoted anti-inflammatory (“M2” polarization) cytokines in LPS-mediated microglia. Phi mitigated “M1” polarization and promoted “M2” polarization of microglia via enhancing PPARγ and inhibiting the NF-κB pathway. The PPARγ antagonist GW9662 significantly repressed Phi-mediated anti-inflammatory effects. Meanwhile, Phi enhanced the viability, tube formation ability, and cell junction of BMECs. In the TBI mouse model, Phi promoted “M2” polarization, whereas it repressed the “M1” polarization of microglia. In addition, Phi reduced TBI-mediated BBB damage. However, the protective effects of Phi were reversed mainly by GW9662 treatment.Conclusion: Phi prevents BBB damage via inhibiting the neuroinflammation of microglia through the PPARγ/NF-κB pathway, which provides a potential therapeutic drug against TBI.

Background: The risk of co-epidemic between COVID-19 and influenza is very high, so it is urgent to find a treatment strategy for the co-infection. Previous studies have shown that phillyrin can not only inhibit the replication of the two viruses, but also has a good anti-inflammatory effect, which is expected to become a candidate compound against COVID-19 and influenza. Objective: To explore the possibility of phillyrin as a candidate compound for the treatment of COVID-19 and influenza co-infection and to speculate its potential regulatory mechanism. Methods: We used a series of bioinformatics network pharmacology methods to understand and characterize the pharmacological targets, biological functions, and therapeutic mechanisms of phillyrin in COVID-19 and influenza co-infection and discover its therapeutic potential. Results: We revealed potential targets, biological processes, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and upstream pathway activity of phillyrin against COVID-19 and influenza co-infection. We constructed protein–protein interaction (PPI) network and identified 50 hub genes, such as MMP9, IL-2, VEGFA, AKT, and HIF-1A. Furthermore, our findings indicated that the treatment of phillyrin for COVID-19 and influenza co-infection was associated with immune balance and regulation of hypoxia-cytokine storm, including HIF-1 signaling pathway, PI3K-Akt signaling pathway, Ras signaling pathway, and T cell receptor signaling pathway. Conclusion: For the first time, we uncovered the potential targets and biological pathways of phillyrin for COVID-19 and influenza co-infection. These findings should solve the urgent problem of co-infection of COVID-19 and influenza that the world will face in the future, but clinical drug trials are needed for verification in the future. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Chinese Institute of Food Science & Technology / Zhongguo Shipin Xuebao is the property of Journal of Chinese Institute of Food Science & Technology Periodical Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: Phillyrin (Phi) is the main polyphenolic compound found in Forsythia suspensa. Recent studies have revealed that Phi has potent antioxidative and anti-inflammatory effects. However, whether Phi could relieve blood–brain barrier (BBB) damage following traumatic brain injury (TBI) remains unknown. Materials and Methods: Lipopolysaccharide (LPS) was used to activate primary microglia, which were then treated with different doses of Phi or the peroxisome proliferator–activated receptor-gamma (PPARγ) antagonist (GW9662). CCK-8 assay was used for evaluating cell viability, and the cytokines (including IL-1β, IL-6, TNFα, IL-4, IL-10, and TGFβ), microglial phenotypic markers (iNOS, COX2, and CD86 for "M1" polarization; Arg1, Ym1, and CD206 for "M2" polarization), PPARγ, and NF-κB were determined by RT-PCR, Western blot, or cellular immunofluorescence. Primary cultured mouse brain microvascular endothelial cells (BMECs) were stimulated by the condition medium (CM) from microglia. The cell viability, angiogenesis, and tight junction of BMECs were determined via CCK-8 assay, tube formation assay, and Western blot (for detecting MMP3, MMP9, ZO1, claudin-5, and occludin). Furthermore, the mouse TBI model was constructed and treated with Phi and/or GW9662. The BBB integrity was evaluated by H&E staining, Evans blue staining, and tissue immunofluorescence. Results: Phi markedly restrained the pro-inflammatory ("M1" state) cytokines and promoted anti-inflammatory ("M2" polarization) cytokines in LPS-mediated microglia. Phi mitigated "M1" polarization and promoted "M2" polarization of microglia via enhancing PPARγ and inhibiting the NF-κB pathway. The PPARγ antagonist GW9662 significantly repressed Phi-mediated anti-inflammatory effects. Meanwhile, Phi enhanced the viability, tube formation ability, and cell junction of BMECs. In the TBI mouse model, Phi promoted "M2" polarization, whereas it repressed the "M1" polarization of microglia. In addition, Phi reduced TBI-mediated BBB damage. However, the protective effects of Phi were reversed mainly by GW9662 treatment. Conclusion: Phi prevents BBB damage via inhibiting the neuroinflammation of microglia through the PPARγ/NF-κB pathway, which provides a potential therapeutic drug against TBI. [ABSTRACT FROM AUTHOR]

Objective To establish a HPLC fingerprints of Biyuanjing capsules. Methods The column was Agilent SB-C18( 4.6mm×250 mm, 5 μm). The mobile phase was acetonitrile-water with gradient elution at a flow rate of 1.0 ml/min. The detection wavelength was 210 nm. The detection time was 80 min. Results The HPLC fingerprints of Biyuanjing capsules were established. Twenty common peaks were confirmed, of which, 15 peaks were belonging to each crude drug and 5 peaks were identified as chemical components. The overall similarity of the fingerprints of 10 batches of samples was above 90% comparing with the control. Conclusion　This method can be used for the quality control of Biyuanjing capsules. [ABSTRACT FROM AUTHOR]

Objective To investigate the inhibitory effect of phillyrin (PHI) on lipopolysaccharide (LPS)-induced hepatic stellate cell (HSC) activation and explore the possible mechanisms. Methods Cultured rat hepatic stellate cells (HSC-T6) were divided into blank control group, LPS (1 μg/mL) group, LPS (1 μg/mL) + PHI (1 μg/mL) group, and LPS (1 μg/mL) + PHI (10 μg/mL) group. We assessed the changes in the proliferation and migration of the cells (HSC-T6) after the treatments using MTT assay and Transwell migration assay, respectively. The intracellular reactive oxygen species (ROS) level in the cells was determined with flow cytometry, and the concentrations of proinflammatory cytokines in the supernatant were tested using enzyme-linked immunosorbant assay (ELISA). The expression levels of α-SMA and phosphorylated nuclear factor-κB (NF-κB) p65 in the cells were analyzed with Western blotting. Results PHI from the concentration of 1 to 100 μmol/L did not significantly affect the proliferation of HSC-T6 cells. LPS stimaulation significantly promoted the proliferation and migration of the cells, and such effects were obviously suppressed by PHI treatment at both 1 (16.13% and 17.14%) and 10 μmol/L(48.99% and 58.86%). PHI at 10 μmol/L significantly reduced the expression of α-SMA (27.33%) in HSC-T6 cells exposed to LPS; at both 1 and 10 μmol/L, PHI strongly suppressed LPS-induced elevation of intracellular ROS level(30.03% and 51.26%), lowered the levels of proinflammatory cytokines TNF-α(23.16% and 39.51%), IL-6(36.57% and 43.75%) and IL-1β(58.99% and 73.01%) and decreased the expression of phosphorylated NF-κB p65(13.90% and 24.96%) in LPS-stimulated cells. The decreased expression level of phosphorylated NF-κB p65 protein in HSC-T6 cells was positively correlated with intracellular ROS level, concentrations of TNF-α and IL-1β, and the expression level of HSC activation marker α-SMA protein. Conclusion PHI can inhibit HSC activation induced by LPS possibly by down-regulating inflammatory responses mediated by NF-κB signaling pathway.

Forsythia suspensa (Thunb.) has been widely used in traditional medicines in Asia. According to the 2020 edition of Chinese Pharmacopoeia, phillyrin is the main active ingredient in F. suspensa, which is effective in clearing heat, reducing swelling, and dispersing nodules. F. suspensa leaf is a non-toxic substance and it can be used to make a health tea. Here, we combine elicitors and transcriptomics to investigate the inducible biosynthesis of the phillyrin from the F. suspensa. After the fruits and leaves of F. suspensa were treated with different concentrations of methyl jasmonate (MeJA), the content of phillyrin in the fruits reached a peak at 200 µM MeJA for 12 h, but which was decreased in leaves. To analyze the differences in key enzyme genes involved in the phillyrin biosynthesis, we sequenced the transcriptome of F. suspensa leaves and fruits treated with 200 µM MeJA for 12 h. We hypothesized that nine genes related to coniferin synthesis including: F. suspensa UDP-glycosyltransferase (FsUGT); F. suspensa 4-coumarate coenzyme CoA ligase (Fs4CL); and F. suspensa Caffeoyl-CoA O-methyltransferase (FsCCoAOMT) etc. The qRT-PCR analysis of genes related to phillyrin biosynthesis was consistent with RNA-seq analysis. We also investigated the dynamic changes of genes in F. suspensa leaves and fruits at different time points after 200 µM MeJA treatment, which laid the foundation for further study of the molecular mechanisms regulating the biosynthesis of phillyrin.

Objective: This article intends to use molecular docking technology to find potential inhibitors that can respond to COVID-19 from active compounds in Mongolian medicine. Methods: Mongolian medicine with anti-inflammatory and antiviral effects is selected from Mongolian medicine prescription preparations. TCMSP, ETCM database and document mining methods were used to collect active compounds. Swiss TargetPrediction and SuperPred server were used to find targets of compounds with smiles number. Drugbank and Genecard database were used to collect antiviral drug targets. Then the above targets were compared and analyzed to screen out antiviral targets of Mongolia medicine. Metascape database platform was used to enrich and analyze the GO (Gene ontology) annotation and KEGG pathway of the targets. In view of the high homology of gene sequences between SARS-CoV-2 S-protein RBD domain and SARS virus, as well as their similarities in pathogenesis and clinical manifestations, we established SARS-CoV-2 S-protein model using Swiss-Model. The ZDOCK protein docking software was applied to dock the S-protein with the human angiotensin ACE2 protein to find out the key amino acids of the binding site. Taking ACE2 as the receptor, the molecular docking between the active ingredients and the target protein was studied by AutoDock molecular docking software. The interaction between ligand and receptor is applied to provide a choice for screening anti-COVID-19 drugs. Results: A total of 253 active components were predicted. Metascape analysis showed that key candidate targets were significantly enriched in multiple pathways related to different toxins. These key candidate targets were mainly derived from phillyrin and chlorogenic acid. Through the protein docking between S-protein and ACE2, it is found that Glu329/Gln325 and Gln42/Asp38 in ACE2 play an important role in the binding process of the two. The results of molecular docking virtual calculation showed that phillyrin and chlorogenic acid could stably combine with Gln325 and Gln42/Asp38 in ACE2, respectively, which hindered the combination between S- protein and ACE2. Conclusion: Phillyrin and chlorogenic acid can effectively prevent the combination of SARS-CoV-2 S-protein and ACE2 at the molecular level. Phillyrin and chlorogenic acid can be used as potential inhibitors of COVID-19 for further research and development.

Cardiac fibrosis (CF) greatly influences the therapeutic effects of heart diseases and remains an urgent challenge in clinical therapy. Till now, only a few methods are used to find potential anti-CF drugs effectively. This study aimed to construct a gene functional module to represent the core pathological process of CF and screen antifibrotic agents capable of decreasing the expression of the gene functional module. First, three CF marker genes Postn, Ddr2, and Pdgfra were selected to identify the corresponding highest coexpressed genes in the genome-based transcriptional profiles of human hearts. Both the marker genes and the coexpressed genes formed the CF-related gene functional module. Second, the correlation of the module with the CF process was measured in a collection of gene expression profiles of heart diseases to evaluate the participation of the functional module in heart diseases. Third, the anti-CF effects of phillyrin were predicted by the enrichment analysis of the module in the phillyrin-induced transcriptional profile. Finally, the myocardial infarction animal model was used to validate the cardioprotective and anti-CF effects of phillyrin experimentally. The results showed that phillyrin was a novel antifibrotic agent in heart diseases.

Cardiac fibrosis (CF) greatly influences the therapeutic effects of heart diseases and remains an urgent challenge in clinical therapy. Till now, only a few methods are used to find potential anti-CF drugs effectively. This study aimed to construct a gene functional module to represent the core pathological process of CF and screen antifibrotic agents capable of decreasing the expression of the gene functional module. First, three CF marker genes Postn , Ddr2 , and Pdgfra were selected to identify the corresponding highest coexpressed genes in the genome-based transcriptional profiles of human hearts. Both the marker genes and the coexpressed genes formed the CF-related gene functional module. Second, the correlation of the module with the CF process was measured in a collection of gene expression profiles of heart diseases to evaluate the participation of the functional module in heart diseases. Third, the anti-CF effects of phillyrin were predicted by the enrichment analysis of the module in the phillyrin-induced transcriptional profile. Finally, the myocardial infarction animal model was used to validate the cardioprotective and anti-CF effects of phillyrin experimentally. The results showed that phillyrin was a novel antifibrotic agent in heart diseases. [ABSTRACT FROM AUTHOR]

Damage to the integrity of heparin sulfate (HS) in the endothelial glycocalyx is an important factor of glomerular filtration barrier dysfunction, which is the basic pathological feature of acute kidney injury (AKI). AKI is a common clinical critical illness with few drugs options offering effective treatment. Phillyrin (Phil), the main pharmacological component of Forsythia suspensa, possesses a wide range of pharmacological activities. However, the effects of Phil on lipopolysaccharide (LPS)-induced AKI have yet to be reported. The aim of the present study is to analyze the effects of Phil on HS damage and inflammatory signaling pathways in LPS-induced AKI. Results revealed that Phil reduces pathological changes and improves renal function in LPS-induced AKI. Further analysis indicated that Phil effectively protects against glycocalyx HS degradation in LPS-stimulated EA.hy926 cells in vitro and LPS-induced AKI mice in vivo. The protective effect of Phil on HS damage may be associated with the isolate's ability to suppress the production of reactive oxygen species, and decrease expression levels of cathepsin L and heparanase in vitro and in vivo. In addition, ELISA and Western blot results revealed that Phil inhibits the activation of the NF-κB and MAPK signaling pathways and decreases the levels of inflammatory cytokines (IL-1β, IL-6, and TNF-α) in LPS-induced ARDS mice. In general, protection against endothelial glycocalyx HS damage and inhibition of inflammatory responses by Phil may be used as treatment targets for LPS-induced AKI. [ABSTRACT FROM AUTHOR]

• Phillyrin and phillygenol trigger an untypical static fluorescence quench of lysozyme. • Hydrophobic interaction play major role in forming complex. • The higher affinity and more structural changes of lysozyme induced by phillygenol. Phillyrin (PR) and its derivative phillygenol (PG) are major active extracts of the Chinese medicine Forsythiae Fructus. Lysozyme, an important carrier protein, has a variety of physiological along with pharmaceutical functions. Herein, the interactions of PR and PG with hen egg white lysozyme (HEWL) are investigated in details by spectroscopic analyses, differential scanning calorimetry (DSC) tests, and molecular modeling approaches. Fluorescence measurement and UV–visible absorption difference spectroscopy reveal that an "untypical static quenching" takes place pointing towards the formation of ground state complex. The binding constants of HEWL-PG are greater than that of HEWL-PR. Moreover, the major driving force for the binding process is elucidated as hydrophobic interactions for both cases. Circular dichroism spectroscopy and three-dimensional fluorescence indicate that PR and PG decrease the helical structure of the HEWL and hydrophobicity surrounding Trp and Tyr residues. The thermal stability of HEWL decreases in the presence of PR and PG by DSC. Docking study infers that both PR and PG could bind to the hydrophobic cavity of HEWL and supports the higher binding of PG than PR. Molecular dynamics study illustrates that PR has higher relative position change, which may attribute to its lower binding ability. In terms of the radius of gyration (Rg), PG causes the protein structure to swell even more. The average hydrogen bonds number of HEWL-PG is less than that of HEWL-PR and more changes of the residual fluctuation are observed in HEWL-PG system. Furthermore, MM-PBSA analysis exhibits that HEWL-PG has lower binding energy. Collectively, these results support that both PR and PG could bind to HEWL, but PG possesses higher affinity which leads to stronger secondary structural changes of HEWL than PR. These findings are beneficial to comprehensively understand the binding characteristic of PR and PG with HEWL, providing experimental basis for pharmacokenitics of PR and PG. [ABSTRACT FROM AUTHOR]

Impaired wound healing in traumatic skin injuries remains a severe clinical challenge due to impaired re-vascularization, harmful bacteria infection, and inflammation dysregulation. Macrophages are recognized as prominent immune cells in tissue regeneration and wound healing. Consequently, the modulation of macrophages provides a promising therapeutic target for wound healing disorders. Here, we aimed to explore whether a novel constructed combination of thermosensitive hydrogel Pluronic F-127 (PF-127) and phillyrin (PH, the main active compound of forsythia suspensa) could improve skin wound healing. Firstly, the biological effects of pH on the phenotype and inflammation of macrophages were assessed by flow cytometry and ELISA. The biocompatibility of the PF-127 plus PH combination was investigated on keratinocytes and red blood cells. The biological effect of PF-127/PH hydrogel on the migratory ability of keratinocytes in vitro was evaluated using the scratch and transwell migration assays. In addition, S. aureus and E. coli were employed to test the antibacterial properties of the PF-127 plus PH combination. Finally, PF-127 plus PH scaffold was applied to the full-thickness skin defect in mice. Histomorphological evaluation and immunochemistry were performed to explore the wound-healing activity of PF-127/PH hydrogel. PH can promote the polarization of macrophages from the M1 (pro-inflammatory) phenotype to the M2 (anti-inflammatory) phenotype. The PF-127/PH hydrogel was highly biocompatible and showed a potent stimulative effect on the migration of keratinocytes in vitro. The combination of PF-127 and PH exerted a pronounced antibacterial activity on S. aureus and E. coli in vitro. PF-127/PH hydrogel potently accelerates the healing of full-thickness skin defects by promoting skin cell proliferation, accelerating angiogenesis, and inhibiting inflammation. Our study suggests that PF-127/PH hydrogel has excellent potential for treating traumatic skin defects. [ABSTRACT FROM AUTHOR]

Background/Aims: Forsythia suspensa Vahl. (Oleaceae) fruits are widely used in traditional Chinese medicine to treat pneumonia, typhoid, dysentery, ulcers and oedema. Antibacterial and anti-inflammatory activities have been reported for phillyrin (PHN), the main ingredient in Forsythia suspensa Vahl fruits, in vitro. However, the underlying mechanisms in vivo remain poorly defined. In this study, we discovered that PHN exerted potent anti-inflammatory effects in lethal LPS-induced neutrophil inflammation by suppressing the MyD88-dependent signalling pathway in zebrafish. Methods: LPS-yolk microinjection was used to induce a lethal LPS-infected zebrafish model. The effect of PHN on the survival of zebrafish challenged with lethal LPS was evaluated using survival analysis. The effect of PHN on neutrophil inflammation grading in vivo was assessed by tracking neutrophils with a transgenic line. The effects of PHN on neutrophil production and migration were analysed by SB+ cell counts during consecutive hours after modelling. Additionally, key cytokines and members of the MyD88 signalling pathway that are involved in inflammatory response were detected using quantitative RT-PCR. To assess gene expression changes during consecutive hours after modelling, the IL-1β, IL-6, TNF-α, MyD88, TRIF, ERK1/2, JNK, IκBa and NF-κB expression levels were measured. Results: PHN could protect zebrafish against a lethal LPS challenge in a dose-dependent manner, as indicated by decreased neutrophil infltration, reduced tissue necrosis and increased survival rates. Up-regulated IL-1β, IL-6 and TNF-α expression also showed the same tendencies of depression by PHN. Critically, PHN significantly inhibited the LPS-induced activation of MyD88, IκBa, and NF-κB but did not affect the expression of ERK1/2 MAPKs or JNK MAPKs in LPS-stimulated zebrafish. Additionally, PHN regulated the MyD88/IκBα/NF-κB signalling pathway by controlling IκBα, IL-1β, IL-6, and TNF-α expression. Conclusion: This study provides a rationale for the clinical application of PHN as an anti-inflammatory agent.