The Efficacy & Molecular Mechanisms of a Terpenoid Compound Ganoderic Acid C1 on Corticosteroid-Resistant Neutrophilic Airway Inflammation: In vivo and in vitro Validation.

Introduction: Neutrophil predominant airway inflammation is associated with severe and steroid-resistant asthma clusters. Previously, we reported efficacy of ASHMI, a three-herb TCM asthma formula in a steroid-resistant neutrophil-dominant murine asthma model and further identified Ganoderic Acid C1 (GAC1) as a key ASHMI active compound in vitro. The objective of this study is to investigate GAC1 effect on neutrophil-dominant, steroid-resistant asthma in a murine model. Methods: In this study, Balb/c mice were systematically sensitized with ragweed (RW) and alum and intranasally challenged with ragweed. Unsensitized/PBS challenged mice served as normal controls. Post sensitization, mice were given 4 weeks of oral treatment with GAC1 or acute dexamethasone (Dex) treatment at 48 hours prior to challenge. Pulmonary cytokines were measured by ELISA, and lung sections were processed for histology by H&E staining. Furthermore, GAC1 effect on MUC5AC expression and on reactive oxygen species (ROS) production in human lung epithelial cell line (NCI-H292) was determined by qRT-PCR and ROS assay kit, respectively. Computational analysis was applied to select potential targets of GAC1 in steroid-resistant neutrophil-dominant asthma. Molecular docking was performed to predict binding modes between GAC1 and Dex with TNF-α. Results: The result of the study showed that chronic GAC1 treatment, significantly reduced pulmonary inflammation (P < 0.01-0.001 vs Sham) and airway neutrophilia (P < 0.01 vs Sham), inhibited TNF-α, IL-4 and IL-5 levels (P < 0.05-0.001 vs Sham). Acute Dex treatment reduced eosinophilic inflammation and IL-4, IL-5 levels, but had no effect on neutrophilia and TNF-α production. GAC1 treated H292 cells showed decreased MUC5AC gene expression and production of ROS (P < 0.001 vs stimulated/untreated cells). Molecular docking results showed binding energy of complex GAC1-TNF was -10.8 kcal/mol. Discussion: GAC1 may be a promising anti-asthma botanical drug for treatment of steroid-resistant asthma.

Time-dependent dual beneficial modulation of interferon-γ, interleukin 5, and Treg cytokines in asthma patient peripheral blood mononuclear cells by ganoderic acid B.

Th2 cytokines play a dominant role in the pathogenesis of allergic asthma. Interferon gamma (IFN-γ), a Th1 cytokine, links to therapeutic mechanisms of allergic asthma. Interleukin (IL)-10, a regulatory cytokine, is involved in the induction of immune tolerance. We previously demonstrated that Anti-Asthma Simplified Herbal Medicine Intervention (ASHMI) suppressed Th2 and increased IFN-γ in patients with asthma and in animal models, but its bioactive compound is unknown. Ganoderic acid beta (GAB) was isolated from Ganoderma lucidum (one herb in ASHMI). Human peripheral blood mononuclear cells (PBMCs) from adult patients with asthma were cultured with GAB or dexamethasone (Dex) in the presence of environmental allergens. The cytokine levels of IL-10, IFN-γ, IL-5, transcription factors T-bet, Foxp-3, and GATA3 were measured. Following 3-day culture, GAB, but not Dex, significantly increased IL-10 and IFN-γ levels by allergic patients' PBMCs. Following 6-day treatment, GAB inhibited IL-5 production, but IL-10 and IFN-γ remained high. Dex suppressed production of all three cytokines. GAB suppressed GATA3 and maintained Foxp-3 and T-bet gene expression, while Dex significantly suppressed GATA3 and T-bet expression. GAB simultaneously increased IL-10, IFN-γ associated with induction of T-bet and Foxp3, while suppressing IL-5, which was associated with suppression of GATA3, demonstrating unique beneficial cytokine modulatory effect, which distinguishes from Dex's overall suppression.

Anti-IgE effect of small-molecule-compound arctigenin on food allergy in association with a distinct transcriptome profile.

BACKGROUND: Excessive production of IgE plays a major role in the pathology of food allergy. In an attempt to identify anti-IgE natural products, Arctium Lappa was one of the most effective herbs among approximately 300 screened medicinal herbs. However, little is known about its anti-IgE compounds. OBJECTIVE: To identify compounds from Arctium Lappa for targeted therapy on IgE production and explore their underlying mechanisms. METHODS: Liquid-liquid extraction and column chromatographic methods were used to purify the compounds. IgE inhibitory effects were determined on IgE-producing human myeloma U266 cells, peanut-allergic murine model and PBMCs from food-allergic patients. Genes involved in IgE inhibition in PBMCs were studied by RNA sequencing. RESULTS: The main compounds isolated were identified as arctiin and arctigenin. Both compounds significantly inhibited IgE production in U266 cells, with arctigenin the most potent (IC50=5.09µg/mL). Arctigenin (at a dose of 13 mg/kg) markedly reduced peanut-specific IgE levels, blocked hypothermia and histamine release in a peanut-allergic mouse model. Arctigenin also significantly reduced IgE production and Th2 cytokines (IL-5, IL-13) by PBMCs. We found 479 differentially expressed genes in PBMCs with arctigenin treatment (p < .001 and fold-change ≥1.5), involving 24 gene ontology terms (p < .001, FDR <0.05); cell division was the most significant. Eleven genes including UBE2C and BCL6 were validated by qPCR. CONCLUSION: Arctigenin markedly inhibited IgE production in U266 cells, peanut-allergic murine model and PBMCs from allergic patients by down-regulating cell division, cell cycle-related genes and up-regulating anti-inflammatory factors.

Sophora flavescens Alkaloids and Corticosteroid Synergistically Augment IL-10/IL-5 Ratio with Foxp3-Gene-Epigenetic Modification in Asthma PBMCs.

BACKGROUND: It has been demonstrated that ASHMI (antiasthma-simplified herbal medicine intervention) can improve airway function and reduce inflammation in human asthmatic patients with high safety and tolerability. In addition, ASHMI significantly suppresses Th2 cytokine production and increases Th1 cytokine production in treating asthma. OBJECTIVE: Allergic asthma is associated with dysregulation of cytokines. We focused on IL-5 and IL-10 as signature Th2 and Treg cytokines to characterize ASHMI immunomodulatory components. METHODS: The effects of ASHMI and individual herbal constituents on IL-5 and IL-10 production by PBMCs from asthmatic subjects were determined ex vivo. Sophora flavescens (SF)-F2, containing alkaloid compounds, effects on PBMC IL-10 and IL-5 production in the presence or absence of dexamethasone (Dex), and on DNA methylation levels at the foxp3 gene promoter were determined. RESULTS: The ratio of anti-CD3/CD28 stimulated IL-10/IL-5 production by PBMCs from asthmatic subjects was significantly reduced compared to healthy subjects. In PBMCs from asthmatic subjects, ASHMI significantly reduced IL-5 production and increased IL-10 secretion in a dose-dependent manner (p < 0.05-0.01). SF-F2 was most effective in increasing IL-10, whereas SF-F4 (flavonoid compounds) was most effective in suppressing IL-5 production. Dex-treated PBMCs from asthma subjects showed a trend of increasing ratio of IL-10/IL-5 while demonstrating reduced levels in both IL-5 and IL-10 (p < 0.05). Co-culture with Dex and SF-F2 significantly prevented Dex suppression of IL-10, while retained Dex-suppression of IL-5 production, and increased IL-10/IL-5 ratio by Dex. Co-culture with SF-F2 and Dex significantly reduced DNA methylation levels at the foxp3 gene promoter at CpG-126. CONCLUSION: The SF alkaloid-rich fraction may be responsible for ASHMI induction of IL-10 production by PBMCs and plays a synergistic effect with Dex for augmenting IL-10/IL-5 ratio.

Optically enriched and guided dynamics of active skyrmions.

Light provides a powerful means of controlling physical behavior of materials but is rarely used to power and guide active matter systems. We demonstrate optical control of liquid crystalline topological solitons dubbed "skyrmions", which recently emerged as highly reconfigurable inanimate active particles capable of exhibiting emergent collective behaviors like schooling. Because of a chiral nematic liquid crystal's natural tendency to twist and its facile response to electric fields and light, it serves as a testbed for dynamic control of skyrmions and other active particles. Using ambient-intensity unstructured light, we demonstrate large-scale multifaceted reconfigurations and unjamming of collective skyrmion motions powered by oscillating electric fields and guided by optically-induced obstacles and patterned illumination.

Publisher Correction: Schools of skyrmions with electrically tunable elastic interactions.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Light-controlled skyrmions and torons as reconfigurable particles.

Topological solitons, such as skyrmions, arise in field theories of systems ranging from Bose-Einstein condensates to optics, particle physics, and cosmology, but they are rarely accessible experimentally. Chiral nematic liquid crystals provide a platform to study skyrmions because of their natural tendency to form twisted structures arising from the lack of mirror symmetry at the molecular level. However, large-scale dynamic control and technological utility of skyrmions remain limited. Combining experiments and numerical modeling of chiral liquid crystals with optically controlled helical pitch, we demonstrate that low-intensity, unstructured light can control stability, dimensions, interactions, spatial patterning, self-assembly, and dynamics of these topological solitons.

Schools of skyrmions with electrically tunable elastic interactions.

Coexistence of order and fluidity in soft matter often mimics that in biology, allowing for complex dynamics and applications-like displays. In active soft matter, emergent order can arise because of such dynamics. Powered by local energy conversion, this behavior resembles motions in living systems, like schooling of fish. Similar dynamics at cellular levels drive biological processes and generate macroscopic work. Inanimate particles capable of such emergent behavior could power nanomachines, but most active systems have biological origins. Here we show that thousands-to-millions of topological solitons, dubbed "skyrmions", while each converting macroscopically-supplied electric energy, exhibit collective motions along spontaneously-chosen directions uncorrelated with the direction of electric field. Within these "schools" of skyrmions, we uncover polar ordering, reconfigurable multi-skyrmion clustering and large-scale cohesion mediated by out-of-equilibrium elastic interactions. Remarkably, this behavior arises under conditions similar to those in liquid crystal displays and may enable dynamic materials with strong emergent electro-optic responses.

Berberine as a chemical and pharmacokinetic marker of the butanol-extracted Food Allergy Herbal Formula-2.

RATIONALE: Food Allergy Herbal Formula-2 (FAHF-2) provided protection against peanut anaphylaxis in a murine model and induced beneficial immune-modulation in humans. Butanol-refined FAHF-2, B-FAHF-2, retained safety and efficacy in the peanut allergic murine model at only 1/5 of FAHF-2 dosage. One compound, berberine, was isolated and identified in vitro as a bioactive component present in FAHF-2 and B-FAHF-2. The aim of this study was to investigate berberine as a chemical and pharmacokinetic marker of B-FAHF-2. METHODS: The consistency of constituents between B-FAHF-2 and FAHF-2 was tested. Peanut allergic C3H/HeJ mice were orally administered with 1mg of berberine or B-FAHF-2 containing an equivalent amount of berberine, and the ability to protect against peanut anaphylaxis and pharmacokinetic parameters were determined. Human intestinal epithelial cells (Caco-2) were cultured with berberine with or without the nine individual herbal constituents in B-FAHF-2, and the absorbed berberine levels were determined. RESULTS: Berberine is one of the major components in B-FAHF-2 and FAHF-2 formula. In a peanut allergic mouse model, B-FAHF-2, but not berberine, protected mice from anaphylaxis reactions. Pharmacokinetic profiles showed that the Cmax of B-FAHF-2 fed mice was 289.30±185.40ng/mL; whereas berberine alone showed very low bioavailability with Cmax value of 35.13±47.90ng/mL. Caco-2 cells influx assay showed that 7 of 9 herbal constituents in B-FAHF-2 increased berberine absorption at rates ranging from 18 to 205%. CONCLUSIONS: B-FAHF-2 remarkably increased the bioavailability of berberine. Berberine can be used as chemical and pharmacokinetic marker of B-FAHF-2. Other herbal components in B-FAHF-2 may facilitate the absorption of berberine.

The Flavonoid 7,4'-Dihydroxyflavone Prevents Dexamethasone Paradoxical Adverse Effect on Eotaxin Production by Human Fibroblasts.

Eotaxin/CCL-11 is a major chemoattractant that contributes to eosinophilic inflammation in asthma. Glucocorticoids inhibit inflammation, but long-time exposure may cause paradoxical adverse effects by augmenting eotaxin/CCL-11production. The aim of this study was to determine if 7,4'-dihydroxyflavone (7,4'-DHF), the eotaxin/CCL11 inhibitor isolated from Glycyrrhiza uralensis, reduces in vitro eotaxin production induced by long-time dexamethasone (Dex) exposure, and if so, to elucidate the mechanisms of this inhibition. Human lung fibroblast-1 cells were used to identify the potency of 7,4'-DHF compared with other compounds from G. uralensis, to compare 7,4'-DHF with Dex on eotaxin production following 24-h short-time culture and 72-h longer-time (LT) culture, and to determine the effects of the 7,4'-DHF on Dex LT culture augmented eotaxin production and molecule mechanisms. 7,4'-DHF was the most potent eotaxin/CCL-11 inhibitor among the ten compounds and provided continued suppression. In contrast to short-time culture, Dex LT culture increased constitutively, and IL-4/TNF-α stimulated eotaxin/CCL11 production by human lung fibroblast-1 cells. This adverse effect was abrogated by 7,4'-DHF co-culture. 7,4'-DHF significantly inhibited Dex LT culture augmentation of p-STAT6 and impaired HDAC2 expression. This study demonstrated that 7,4'-DHF has the ability to consistently suppress eotaxin production and prevent Dex-paradoxical adverse effects on eotaxin production. Copyright © 2017 John Wiley & Sons, Ltd.

Ganoderic acid C1 isolated from the anti-asthma formula, ASHMI™ suppresses TNF-α production by mouse macrophages and peripheral blood mononuclear cells from asthma patients.

Asthma is a heterogeneous airway inflammatory disease, which is associated with Th2 cytokine-driven inflammation and non-Th2, TNF-α mediated inflammation. Unlike Th2 mediated inflammation, TNF-α mediated asthma inflammation is generally insensitive to inhaled corticosteroids (ICS). ASHMITM, aqueous extract of three medicinal herbs-Ganoderma lucidum (G. lucidum), Sophora flavescens Ait (S. flavescens) and Glycyrrhiza uralensis Fischer (G. uralensis), showed a high safety profile and was clinically beneficial in asthma patients. It also suppresses both Th2 and TNF-α associated inflammation in murine asthma models. We previously determined that G. uralensis flavonoids are the key active compounds responsible for ASHMITM suppression of Th2 mediated inflammation. Until now, there are limited studies on anti-TNF-α compounds presented in ASHMITM. The objective of this study was to isolate and identify TNF-α inhibitory compounds in ASHMITM. Here we report that G. lucidum, but not the other two herbal extracts, S. flavescens or G. uralensis inhibited TNF-α production by murine macrophages; and that the methylene chloride (MC)-triterpenoid-enriched fraction, but not the polysaccharide-enriched fraction, contained the inhibitory compounds. Of the 15 triterpenoids isolated from the MC fraction, only ganoderic acid C1 (GAC1) significantly reduced TNF-α production by murine macrophages (RAW 264.7 cells) and peripheral blood mononuclear cells (PBMCs) from asthma patients. Inhibition was associated with down-regulation of NF-κB expression, and partial suppression of MAPK and AP-1 signaling pathways. Ganoderic acid C1 may have potential for treating TNF-α mediated inflammation in asthma and other inflammatory diseases.

Anti-inflammatory Effects of Ganoderma lucidum Triterpenoid in Human Crohn's Disease Associated with Downregulation of NF-κB Signaling.

BACKGROUND: Crohn's disease (CD) is a chronic inflammatory disease of the gastrointestinal tract. Current medications have potentially serious side effects. Hence, there is increasing interest in alternative therapies. We previously demonstrated the anti-inflammatory effects of Food Allergy Herbal Formula-2 in vitro on peripheral blood mononuclear cells (PBMCs) and mucosa from CD subjects. Here, we investigated the anti-inflammatory effects of a bioactive compound isolated from Ganoderma lucidum (G. lucidum), a key herbal constituent of Food Allergy Herbal Formula-2, in CD in vitro. METHODS: Triterpene ganoderic acid C1 (GAC1) was isolated from G. lucidum. Stimulated RAW 264.7 macrophages were treated with GAC1. Human PBMCs and colonic biopsies were obtained from children with CD and cultured with or without GAC1. TNF-α and other proinflammatory cytokine levels were measured in the culture supernatant. NF-κB signaling was investigated in PBMCs and colonic mucosa treated with GAC1 by In-Cell Western and Western blot analysis. RESULTS: GAC1 decreased TNF-α production by macrophages and PBMCs from CD subjects. GAC1 significantly decreased TNF-α, IFN-γ, and IL-17A production by inflamed colonic biopsies from CD subjects. These effects were due to downregulation of the NF-κB signaling pathway. CONCLUSIONS: GAC1 inhibited production of TNF-α and other proinflammatory cytokines by PBMCs and inflamed CD colonic mucosa due to blockage of NF-κB activation. GAC1 is a key beneficial constituent in G. lucidum and the Food Allergy Herbal Formula-2 in suppressing the inflammatory cytokines found in CD and warrants clinical investigation for the treatment of CD.

The Flavonoid 7,4'-Dihydroxyflavone Inhibits MUC5AC Gene Expression, Production, and Secretion via Regulation of NF-κB, STAT6, and HDAC2.

Mucus overproduction is a significant component of the pathophysiology of obstructive lung diseases. Currently, there are only a few medications available that inhibit mucus production. Previous studies showed that glycyrrhizin, a triterpenoid in Glycyrrhiza uralensis inhibits mucin 5AC (MUC5AC) mRNA and protein expression. Other potential mucus production inhibitory compounds contained within in G. uralensis have not been fully investigated. The aim of the present study was to determine if the G. uralensis flavonoid 7,4'-dihydroxyflavone (7,4'-DHF) inhibits MUC5AC gene expression, mucus production, and secretion, and if so, to elucidate the mechanism of this inhibition. 7,4'-Dihydroxyflavone significantly decreased phorbol 12-myristate 13-acetate-stimulated NCI-H292 human airway epithelial cell MUC5AC gene expression and mucus production, at a 28-fold lower concentration than glycyrrhizin (The half maximal inhibitory concentration IC50 value of 1.4 µM vs 38 µM, respectively); 7,4'-DHF also inhibited MUC5AC mucus secretion. Inhibition was associated with the suppression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), signal transducer and activator of transcription 6 (STAT6) activation, and enhanced histone deacetylase 2 (HDAC2) expression. In a murine model of asthma, 7,4'-DHF-treated mice exhibited a marked reduction in MUC5AC secretion in the bronchoalveolar lavage fluid compared with control mice. These findings, together with previous findings linking NF-κB, STAT6, and HDAC2 modulation to the control of MUC5AC expression, demonstrate that 7,4'-DHF is a newly identified component of G. uralensis that regulates MUC5AC expression and secretion via regulation of NF-κB, STAT6, and HDAC2.

Maternal allergy increases susceptibility to offspring allergy in association with TH2-biased epigenetic alterations in a mouse model of peanut allergy.

BACKGROUND: Although maternal atopy is a risk factor for the development of peanut allergy, this phenomenon has not been well characterized experimentally, and the mechanisms underlying offspring risk are unclear. OBJECTIVE: We sought to determine whether offspring of mothers with peanut allergy (O-PAM mice) are more susceptible to peanut allergy than offspring of naive mothers (O-NM mice) in a murine model and, if so, whether the susceptibility is linked to TH2-biased epigenetic alterations. METHODS: Five-week-old O-PAM and O-NM mice were intragastrically sensitized to and challenged with peanut. Serum peanut-specific IgE levels, plasma histamine levels, anaphylactic reactions, and splenocyte and MLN cell cytokine production were measured. DNA methylation levels of the Il4 gene promoter from splenocytes and MLN cells from sensitized offspring and splenocytes from unsensitized neonatal offspring were determined by means of pyrosequencing. RESULTS: O-PAM mice exhibited 3-fold higher peanut-specific IgE levels after peanut sensitization, as well as 5-fold higher histamine levels and significantly higher anaphylactic symptom scores after challenge than O-NM mice (P < .05-.01). Cultured splenocytes and MLNs from O-PAM mice produced significantly more TH2 cytokines than cells from O-NM mice (P < .05-.01). Cells from O-PAM mice exhibited significantly reduced DNA methylation at CpG sites of the Il4 gene promoter than cells from O-NM mice. DNA methylation levels were inversely correlated with IL-4 and IgE production. O-PAM neonatal splenocyte hypomethylation of the Il4 gene promoter was also present. CONCLUSION: This study is the first to demonstrate that increased susceptibility to peanut allergy in O-PAM mice is associated with epigenetic alteration of the Il4 gene promoter. This finding might provide insight into preventing the development of early-life allergy.

Berberine and limonin suppress IgE production by human B cells and peripheral blood mononuclear cells from food-allergic patients.

BACKGROUND: Currently, there is no satisfactory treatment for IgE-mediated food allergy. Food Allergy Herbal Formula 2 (FAHF-2) and butanol-purified FAHF-2 (B-FAHF-2) have been shown to protect against peanut-induced anaphylaxis and inhibit IgE synthesis in a murine model. OBJECTIVE: To determine which herbs and compounds in FAHF-2 and B-FAHF-2 suppress IgE production. METHODS: The effect of FAHF-2 and B-FAHF-2 on IgE production was determined using a human B-cell line (U266). Individual compounds were isolated and identified using column chromatography, liquid chromatographic mass spectrometry, and nuclear magnetic resonance techniques. The potency of compounds on IgE suppression were investigated using U266 cells and verified using human peripheral blood mononuclear cells (n = 25) from peanut-allergic patients. Epsilon germline transcript expression was determined. Phosphorylated IκBα level was analyzed using the In-Cell Western assay. The mRNA expression of signal transducer and activator of transcription-3, T-box transcription factor TBX21, interferon-γ, forkhead box P3, GATA-binding protein 3, interleukin-10, and interleukin-5 also were analyzed using real-time polymerase chain reaction. RESULTS: FAHF-2 and B-FAHF-2 inhibited IgE production by U266 cells. B-FAHF-2 was 9 times more effective than FAHF-2. Two compounds that inhibited IgE production were isolated from Philodendron chinensis and identified as berberine and limonin. Berberine was more potent and inhibited IgE production by peripheral blood mononuclear cells by 80% at 0.62 µg/mL. Berberine significantly inhibited ε-germline transcript expression by peripheral blood mononuclear cells. Phosphorylated IκBα level was significantly suppressed and mRNA expressions of T-box transcription factor TBX21 and signal transducer and activator of transcription-3 were significantly increased by berberine. CONCLUSION: Berberine and limonin mediated IgE suppression. The mechanism by which berberine modulates ε-germline transcript expression might be through regulating the phosphorylated IκBα level and the expressions of signal transducer and activator of transcription-3 and T-box transcription factor TBX21. TRIAL REGISTRATION: Clinicaltrials.gov identifier NCT00602160.

Weight loss herbal intervention therapy (W-LHIT) a non-appetite suppressing natural product controls weight and lowers cholesterol and glucose levels in a murine model.

BACKGROUND: The prevalence of obesity is increasing in industrialized countries. Obesity increases the risk of coronary artery disease, stroke, cancer, hypertension, and type-2 diabetes. Unfortunately, conventional obesity drug treatment is often associated with adverse effects. The objective of this study was to evaluate a novel natural formula, Weight loss herbal intervention therapy (W-LHIT), developed from traditional Chinese medicine, for weight control in a high-fat-diet (HFD) induced obesity murine model. METHODS: Two sets of experiments were performed. In experiment 1, 14-week-old C57BL/6 J male mice were fed with HFD for 21 days and then separated into 3 weight-matched groups. One group continued on the HFD as obese-controls. Two groups were switched from HFD to normal fat level diet (NFD) and sham or W-LHIT treated. In experiment 2, 25-week-old obese mice, following 2 weeks acclimatization, received either W-LHIT or sham treatment while maintained on HFD. In both sets of experiments, NFD fed, age matched normal weight mice served as normal controls. Body weight and food intake were recorded. Epididymal fat pad weight, serum glucose and cholesterol levels, as well as PPARγ and FABP4 gene expression in epididymal fat tissue were analyzed at the end of the experiment. RESULTS: In experiment 1, W-LHIT treated obese mice lost body weight 12.2 ± 3.8% whereas sham treated mice lost 5.5 ± 2.8% by day 10 after switching from the HFD to the NFD, without reduction of chow consumption. In experiment 2, W-LHIT treated obese mice maintained on the HFD had significantly lower body weight (8 fold less) than the sham treated mice. W-LHIT treatment also reduced epididymal fat pad weight, blood cholesterol and glucose levels versus sham treated mice without reduced chow consumption. In addition, significantly increased PPARγ (peroxisome proliferator activated receptor γ) and FABP4 (fatty acid binding protein 4) gene expression were found in epdidymal fat tissues. Liver and kidney function and hematology testing results of W-LHIT treated mice were within the normal range. CONCLUSIONS: W-LHIT significantly and safely reduced body weight, normalized glucose and cholesterol levels in obese mice, without suppression of appetite, and increased adipocyte PPARγ and FABP4 gene expression.

Effect of Antiasthma Simplified Herbal Medicine Intervention on neutrophil predominant airway inflammation in a ragweed sensitized murine asthma model.

BACKGROUND: Neutrophil-predominant asthma is less responsive to steroids and associated with poorer disease control. The effects of Antiasthma Simplified Herbal Medicine Intervention (ASHMI), a traditional Chinese medicine formula reported to be efficacious in asthmatic patients and murine asthma models, on neutrophil predominant asthma are unknown. OBJECTIVE: To determine the effects of standard ASHMI and refined formula ASHMI (ASHMI(II)) in a neutrophil-predominant murine model of ragweed (RW) asthma and explore underlying mechanisms. METHODS: BALB/c mice were systemically sensitized, intranasally challenged with RW extract, and orally treated with ASHMI, ASHMI(II), or vehicle (water). In a separate experiment, some RW sensitized mice were treated with dexamethasone before challenge. After RW challenge, airway hyperreactivity (AHR), total and differential bronchoalveolar lavage fluid leukocyte counts, lung histologic features, and bronchoalveolar lavage fluid cytokine and chemokine levels were assessed. RW stimulation of the murine macrophage cell line RAW264.7 was used to determine effects of ASHMI active compound ganoderic acid C1 (GAC1) on tumor necrosis factor α (TNF-α) production and regulation of phosphorylated IκB and histone deacetylase 2 (HDAC2) levels. RESULTS: ASHMI and ASHMI(II) markedly reduced AHR, mucous production, neutrophilic inflammation, and TNF-α, interleukin 8, and interleukin 17 levels and decreased eosinophilic inflammation and TH2 responses in vivo (P < .01-.001 for all). GAC1 inhibited TNF-α production in RW-stimulated RAW264.7 cells in association with suppression of phosphorylated IκB and increased HDAC2 expression. Dexamethasone failed to reduce AHR and neutrophilic inflammation. CONCLUSION: ASHMI treatment was efficacious in a murine model of neutrophil-predominant asthma via modulation of innate chemokines, TH2 responses, nuclear factor-κB, and HDAC2. ASHMI, and/or its constituent GAC1, may be a valuable option for treating neutrophil-predominant asthma.

Ethacrynic acid and a derivative enhance apoptosis in arsenic trioxide-treated myeloid leukemia and lymphoma cells: the role of glutathione S-transferase p1-1.

PURPOSE: Arsenic trioxide (ATO) as a single agent is used for treatment of acute promyelocytic leukemia (APL) with minimal toxicity, but therapeutic effect of ATO in other types of malignancies has not been achieved. We tested whether a combination with ethacrynic acid (EA), a glutathione S-transferase P1-1 (GSTP1-1) inhibitor, and a reactive oxygen species (ROS) inducer will extend the therapeutic effect of ATO beyond APL. EXPERIMENTAL DESIGN: The combined apoptotic effects of ATO plus ethacrynic acid were tested in non-APL leukemia and lymphoma cell lines. The role of ROS, GSTP1-1, glutathione (GSH), and Mcl-1 in apoptosis was determined. The selective response to this combination of cells with and without GSTP1-1 expression was compared. RESULTS: ATO/EA combination synergistically induced apoptosis in myeloid leukemia and lymphoma cells. This treatment produced high ROS levels, activated c-jun-NH(2)-kinase (JNK), and reduced Mcl-1 protein. This led to the decrease of mitochondrial transmembrane potential, release of cytochrome c, and subsequently, to activation of caspase-3 and -9. Induction of apoptosis in leukemia and lymphoma cells expressing GSTP1-1 required high ethacrynic acid concentrations to be combined with ATO. Silencing of GSTP1 in leukemia cells sensitized them to ATO/EA-induced apoptosis. In a subgroup of B-cell lymphoma, which does not express GSTP1-1, lower concentrations of ethacrynic acid and its more potent derivative, ethacrynic acid butyl-ester (EABE), decreased intracellular GSH levels and synergistically induced apoptosis when combined with ATO. CONCLUSION: B-cell lymphoma cells lacking GSTP1-1 are more sensitive than myeloid leukemia cells to ATO/EA-induced apoptosis.

A study on the chemical constituents of Veratrum nigrum L. processed by rice vinegar.

One new steroid alkaloid, 12beta-hydroxylveratroylzygadenine (1) and four known compounds, verdine (2), jervine (3), veramarine (4), and veratroylzygadenine (5), have been isolated from the roots and rhizomes of Veratrum nigrum L. processed by rice vinegar. Their structures were established through a combined analysis of physicochemical properties and spectroscopic evidence. The assay results revealed that compounds 1, 4, and 5 exhibited cell toxicity against human HL-60 cells with IC50 values 52.67, 52.90, and 56.51 micromol/l, respectively.