Jiawei Yanghe Decoction attenuate allergic airway inflammation by suppressing group 2 innate lymphoid cells responses.

ETHNOPHARMACOLOGICAL RELEVANCE: Jiawei Yanghe Decoction (JWYHD) is modified Yanghe Decoction (YHD). YHD historically utilized as a potent medicinal solution for addressing chronic inflammatory conditions, holds promising therapeutic potential in the treatment of asthma. However, the mechanisms underlying JWYHD's effects on allergic asthma remain unclear. AIM OF THE STUDY: To investigate the therapeutic effect as well as the underlying mechanisms of JWYHD on asthmatic mice. MATERIALS AND METHODS: The ovalbumin (OVA)-induced mouse model was utilized, followed by the administration of JWYHD to allergic asthmatic mice. Subsequently, inflammatory cells in the bronchoalveolar lavage fluid (BALF) and lung tissues were conducted. The levels of various cytokines including interleukin (IL)-4, IL-5, IL-13, IL-33, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ in BALF, as well as the total immunoglobulin E (IgE) content in serum, were assessed. Lung function and tissue pathology examinations were performed to assess the protective impacts of JWYHD. The chemical components of JWYHD and its lung prototype compounds (referred to the chemical components present in JWYHD that were observed in the lung) were explored by ultra-high performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). RNA-seq analysis revealed the regulation mechanisms of JWYHD treating asthma. Furthermore, the effect of JWYHD on type 2 innate lymphoid cells (ILC2s) in asthmatic mice was detected by flow cytometry and Smart-RNA-seq analysis. Then molecular docking analysis was used to show the interaction between identified compounds and key targets. RESULTS: JWYHD significantly attenuated the airway inflammation of asthmatic mice, reduced the levels of inflammatory cells in BALF, as well the levels of the cytokines IL-4, IL-5, IL-13, IL-33, and TNF-α in BALF and IgE in serum. Airway hyperresponsiveness (AHR) and lung inflammation infiltration were also alleviated by JWYHD. Moreover, RNA-seq analysis revealed that JWYHD attenuated airway inflammation in asthmatic mice via regulating immunity. Flow cytometry confirmed that JWYHD could inhibit ILC2 responses. ILC2 Smart-RNA-seq analysis showed that JWYHD impaired the inflammation reaction-related signaling pathways in ILC2s, and neuropilin-1 (Nrp1), endothelial transcription factor 3 (GATA3) and interleukin 1 receptor like protein 1 (ST2) might be the key targets. The molecular docking analysis investigating the connection between the primary targets and JWYHD's prototype compounds in the lung demonstrated that liquiritin apioside, icariin, glycyrrhizic acid, and uralsaponin B, identified through UPLC-Q-TOF/MS, exhibited significant affinity in binding to the mentioned key targets. CONCLUSION: Our results suggested that the mechanism of JWYHD in treating asthma might be related to limiting ILC2 responses. Our findings provided some pharmacological evidence for the clinical application of JWYHD in the treatment of asthma.

Multi-omics analysis of lung tissue metabolome and proteome reveals the therapeutic effect of Shegan Mahuang Decoction against asthma in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Shegan Mahuang Decoction (SMD) is a classic traditional Chinese medicine (TCM) formula for asthma treatment, but the anti-asthma mechanism of SMD is still not fully studied. AIMS OF THE STUDY: In this study, we established an ovalbumin (OVA)-induced asthma rat model and treated it with SMD to observe its anti-asthma effect and explore the related mechanism. MATERIALS AND METHODS: We evaluated the anti-inflammatory effect of SMD via testing the levels of immunoglobulin E (IgE), C-reactive protein (CRP), interleukin-4 (IL-4), interleukin-6 (IL-6) in serum and performing the hematoxylin-eosin (H&E) staining of lung tissue slices. We analyzed the variations of metabolites and proteins in the lung tissue of different groups using liquid chromatography-mass spectrometry (LC-MS)-based untargeted metabolomics and TMT-based proteomics approaches. The metabolic biomarkers and differentially expressed proteins (DEPs) were picked, and the related signal transduction pathways were also investigated. In addition, the key proteins on the signaling pathway were validated through western blotting (WB) experiment to reveal the anti-asthma mechanism of SMD. RESULTS: The results showed that the SMD could significantly reduce the serum levels of IgE, CRP, IL-4, and IL-6 and attenuate the OVA-induced pathological changes in lung tissue. A total of 34 metabolic biomarkers and 84 DEPs were screened from rat lung tissue, which were mainly associated with lipid metabolism, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation, the excessive production of reactive oxygen species (ROS), and lysosome pathway. Besides, SMD could inhibit the myeloid differentiation factor 88 (MyD88)/inhibitor of kappa B kinase (IKK)/nuclear factor-kappa B (NF-κB) signaling pathway to exhibit anti-inflammatory activities. CONCLUSIONS: SMD exhibited a therapeutic effect on asthma, which possibly be exerted by inhibiting the MyD88/IKK/NF-κB signaling pathway.

Curcumin and PCI-34051 combined treatment ameliorates inflammation and fibrosis by affecting MAP kinase pathway.

OBJECTIVE: Bronchoconstriction, along with inflammation and hyperresponsiveness is the characteristic feature associated with asthma, contributing to variable airflow obstruction, which manifests shortness of breath, cough and wheeze, etc. Histone deacetylases 8 (HDAC8) is the member of class I HDAC family and known to regulate microtubule integrity and muscle contraction. Therefore, we aimed to investigate the effects of HDAC8 inhibition in murine model of asthma using Pan-HDAC inhibitor curcumin (CUR) and HDAC8-specific inhibitor PCI-34051 (PCI), alone and in combination. MATERIALS AND METHODS: To develop asthmatic mouse model, Balb/c mice were sensitized and challenged with ovalbumin (OVA). CUR (10 mg/kg, pre, post, alone and combined treatment) and PCI (0.5 mg/kg), were administered through intranasal (i.n) route, an hour before OVA aerosol challenge. Effects of HDAC8 inhibition by CUR and PCI pretreatments were evaluated in terms of inflammation, oxidative stress and fibrosis markers. Efficacy of curcumin post-treatment (CUR(p)) was also evaluated simultaneously. RESULTS: Inflammatory cell recruitment, oxidative stress (reactive oxygen species, nitric oxide), histamine and Immunoglobulin E (IgE) levels and expression of fibrosis markers including hydroxyproline, matrix metalloproteinases-9 and alpha smooth muscle actin (MMP-9 and α-SMA) were significantly reduced by CUR, CUR(p), PCI-alone and combined treatments. Protein expressions of HDAC8, Nuclear factor-κB (NF-κB) accompanied by MAPKs (mitogen-activated protein kinases) were significantly reduced by the treatments. Structural alterations were examined by histopathological analysis and linked with the fibrotic changes. CONCLUSIONS: Present study indicates protective effects of HDAC8 inhibition in asthma using HDAC8 using CUR and PCI alone or in combination, attenuates airway inflammation, fibrosis and remodeling; hence, bronchoconstriction was accompanied through modulation of MAP kinase pathway.

In vitro anti-inflammatory activity and ameliorative effects on gastric ulcers of Licania rigida benth seed extract.

Licania rigida Benth., a Brazilian endemic plant, has been traditionally used for treating inflammation and stomach pain. This work investigates the anti-inflammatory and gastroprotective activities of the ethanolic extract from L. rigida seeds (EELr) by in vitro and in vivo methods. The phytochemical profile was determined and the in vitro antioxidant activity was investigated by radical scavenging and thiobarbituric acid reactive substances methods. The ovalbumin denaturation method was used with sodium diclofenac as standard for the in vitro anti-inflammatory activity assessment. Acetylsalicylic acid was used to induce gastric ulcers in male mice and then to evaluate the preventive and therapeutic gastroprotective effect of EELr, using omeprazole as the reference drug. The extract exhibited relevant amount of phenolic compounds and flavonoids, in particular, demonstrating in vitro antioxidant capacity. EELr was able to inhibit almost 60% of ovalbumin denaturation at a concentration considered low. It also prevented the decrease of biochemical markers for oxidative stress such as superoxide dismutase (SOD) and reduced glutathione (GSH) in the stomach and SOD and catalase (CAT) in the liver. EELr also significantly decreased the number of lesions as well as reduced the ulcerated area when used as therapy. The observed effect may be due to its phenolic compounds, such as chlorogenic acid, caffeic acid and tannins, as previously reported. EELr is a potential source of compounds with anti-inflammatory activity, protects the liver from oxidative damage and improves healing of aspirin-induced ulcers. This work contributes to the knowledge of L. rigida species.

The Programming Effect of Plant-Based DHA, Along with Equivalent AA, on Immune System and Oral Tolerance Development in Six-Week Allergy Prone BALB/c Pups.

BACKGROUND: Docosahexaenoic acid (DHA) and arachidonic acid (AA) on oral tolerance (OT) development in allergy-prone infants is less known. OBJECTIVES: We aim to determine the effects of early life DHA supplementation (1% of total fat, from novel canola oil), along with AA, on OT toward ovalbumin (ova, egg protein) in allergy-prone BALB/c pups at 6-wk. METHODS: Breastfeeding dams (n ≥ 10/diet) were fed DHA+AA (1% DHA, 1% AA wt/wt of total fat) or control (0% DHA, 0% AA) suckling period diet (SPD) during which pups consumed dam's milk. At 3-wk, pups from each SPD group were assigned to either the control or DHA+AA weaning diet. For OT, pups from each diet group were either orally fed ova or placebo daily from 21-25 d. Systemic immunization to ova was induced through intraperitoneal injections before euthanizing 6-wk pups. Ova-specific immunoglobulin (ova-Ig) and splenocytes ex-vivo cytokine response to different stimuli were analyzed using a 3-factor analysis of variance. RESULTS: OT-induced suppression was seen in ova-stimulated splenocyte ex-vivo response, where ova-tolerized pups showed significantly lower total immunoglobulin (Ig)G, IgG1, interleukin (IL)-2 and IL-6 production than sucrose (placebo) pups. DHA+AA SPD was associated with 3 times lower plasma concentrations of ova-IgE (P = 0.03) than controls. DHA+AA weaning diet resulted in lower T helper type-2 cytokines (IL-4 and IL-6) with ova stimulation than controls, which may benefit OT. DHA+AA SPD resulted in significantly higher T cell cytokine response [IL-2, interferon-gamma, (IFNγ) and IL-1ß] to anti-CD3/CD28 stimulation than controls. The splenocytes stimulated with lipopolysaccharide produced lower inflammatory cytokines (IFNγ, tumor necrosis factor-alpha, IL-6, and C-X-C motif ligand 1), which may be because of lower CD11b+CD68+ splenocytes proportion in pups from DHA+AA SPD than control (all P < 0.05). CONCLUSIONS: DHA and AA in early life may influence OT in allergy-prone BALB/c mouse offspring, as they effectively promote T helper type-1 immune responses.

Pinellia ternata (Thunb.) Breit. attenuates the allergic airway inflammation of cold asthma via inhibiting the activation of TLR4-medicated NF-kB and NLRP3 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Pinellia ternata (Thunb.) Breit. (PT) has been demonstrated to be effective against the allergic airway inflammation (AAI) in clinical practices, especially in cold asthma (CA). Until now, the active ingredients, protective effect, and possible mechanism of PT against CA remain unknown. AIM OF THE STUDY: The aim of this investigation was to examine the therapeutic impact and elucidate the underlying mechanism of PT on the AAI of CA. METHODS: The compositions of PT water extract were determined via the UPLC-Q-TOF-MS/MS. The ovalbumin (OVA) and cold-water baths were used to induce CA in female mice. Morphological characteristic observations, expectorant effect, bronchial hyperreactivity (BHR), excessive mucus secretion, and inflammatory factors were used to uncover the treatment effect of PT water extract. In addition, the mucin 5AC (MUC5AC) mRNA and protein levels and the aquaporin 5 (AQP5) mRNA and protein levels were detected via qRT-PCR, immunohistochemistry (IHC), and western blotting. Moreover, the protein expressions associated with the TLR4, NF-κB, and NLRP3 signaling pathway were monitored by western blot analysis. RESULTS: Thirty-eight compounds were identified from PT water extract. PT showed significant therapeutic effects on mice with cold asthma in terms of expectorant activity, histopathological changes, airway inflammation, mucus secretion, and hyperreactivity. PT exhibited good anti-inflammatory effects in vitro and in vivo. The expression levels of MUC5AC mRNA and protein decreased significantly, while AQP5 expression levels increased significantly in the lung tissues of mice after administration with PT as compared to mice induced by CA. Furthermore, the protein expressions of TLR4, p-iκB, p-p65, IL-1ß, IL-18, NLRP3, cleaved caspase-1, and ASC were markedly reduced following PT treatment. CONCLUSIONS: PT attenuated the AAI of CA by modulating Th1- and Th2-type cytokines. PT could inhibit the TLR4-medicated NF-kB signaling pathway and activate the NLRP3 inflammasome to reduce CA. This study provides an alternative therapeutic agent of the AAI of CA after administration with PT.

Tingli Dazao Xiefei Decoction ameliorates asthma in vivo and in vitro from lung to intestine by modifying NO-CO metabolic disorder mediated inflammation, immune imbalance, cellular barrier damage, oxidative stress and intestinal bacterial disorders.

ETHNOPHARMACOLOGICAL RELEVANCE: Asthma is a chronic airway inflammatory disease. Current treatment of mainstream medications has significant side effects. There is growing evidence that the refractoriness of asthma is closely related to common changes in the lung and intestine. The lungs and intestines, as sites of frequent gas exchange in the body, are widely populated with gas signaling molecules NO and CO, which constitute NO-CO metabolism and may be relevant to the pathogenesis of asthma in the lung and intestine. The Chinese herbal formula Tingli Dazao Xiefei Decoction (TD) is commonly used in clinical practice to treat asthma with good efficacy, but there are few systematic evaluations of the efficacy of asthma on NO-CO metabolism, and the mode of action of its improving effect on the lung and intestine is unclear. AIM OF THE STUDY: To investigate the effect of TD on the lung and intestine of asthmatic rats based on NO-CO metabolism. MATERIALS AND METHODS: In vivo, we established a rat asthma model by intraperitoneal injection of sensitizing solution with OVA atomization, followed by intervention by gavage administration of TD. We simultaneously examined alterations in basal function, pathology, NO-CO metabolism, inflammation and immune cell homeostasis in the lungs and intestines of asthmatic rats, and detected changes in intestinal flora by macrogenome sequencing technology, with a view to multi-angle evaluation of the treatment effects of TD on asthmatic rats. In vitro, lung cells BEAS-2B and intestinal cells NCM-460 were used to establish a model of lung injury causing intestinal injury using LPS and co-culture chambers, and lung cells or intestinal cells TD-containing serum was administered to intervene. Changes in inflammatory, NO-CO metabolism-related, cell barrier-related and oxidative stress indicators were measured in lung cells and intestinal cells to evaluate TD on intestinal injury by way of amelioration and in-depth mechanism. RESULTS: In vivo, our results showed significant basal functional impairment in the lung and intestine of asthmatic rats, and an inflammatory response, immune cell imbalance and intestinal flora disturbance elicited by NO-CO metabolic disorders were observed (P < 0.05 or 0.01). The administration of TD was shown to deliver a multidimensional amelioration of the impairment induced by NO-CO metabolic disorders (P < 0.05 or 0.01). In vitro, the results showed that LPS-induced lung cells BEAS-2B injury could cause NO-CO metabolic disorder-induced inflammatory response, cell permeability damage and oxidative stress damage in intestinal cells NCM-460 (P < 0.01). The ameliorative effect on intestinal cells NCM-460 could only be exerted when TD-containing serum interfered with lung cells BEAS-2B (P < 0.01), suggesting that the intestinal ameliorative effect of TD may be exerted indirectly through the lung. CONCLUSION: TD can ameliorate NO-CO metabolism in the lung and thus achieve the indirectly amelioration of NO-CO metabolism in the intestine, ultimately achieving co-regulation of lung and intestinal inflammation, immune imbalance, cellular barrier damage, oxidative stress and intestinal bacterial disorders in asthma in vivo and in vitro. Targeting lung and intestinal NO-CO metabolic disorders in asthma may be a new therapeutic idea and strategy for asthma.

Shaoyao-Gancao-Tang regulates the T-helper-type 1/T-helper-type 2 ratio in the lung and gut and alters gut microbiota in rats with ovalbumin-induced asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Shaoyao-Gancao Tang (SGT) is a traditional Chinese medicine formulation. It has been used to treat kinds of pain and to alleviate asthma in clinic. However, the mechanism of action is not known. AIM OF THE STUDY: To investigate the anti-asthma effect of SGT involving modulation of the T-helper type 1 (Th1) Th1/Th2 ratio in the gut-lung axis and alteration of the gut microbiota (GM) in rats with ovalbumin (OVA)-induced asthma. MATERIALS AND METHODS: The main constituents of SGT were analyzed by high-performance liquid chromatography (HPLC). A model of asthma was established in rats by OVA-induced allergen challenge. Rats suffering from asthma (RSAs) were treated with SGT (2.5, 5.0 and 10.0 g/kg), dexamethasone (1 mg/kg) or physiologic saline for 4 weeks. The level of immunoglobulin (Ig)E in bronchoalveolar lavage fluid (BALF) and serum was determined by enzyme-linked immunosorbent assay. Histology of lung and colon tissues was investigated using staining (hematoxylin and eosin and periodic acid-Schiff). The Th1/Th2 ratio and levels of cytokines (interferon (IFN)-γ and interleukin (IL)-4) in the lung and colon were detected by immunohistochemistry. The GM in fresh feces was analyzed by 16 S rRNA gene sequencing. RESULTS: Twelve main constituents (gallic acid, albiflorin, paeoniflorin, liquiritin apioside, liquiritin, benzoic acid, isoliquiritin apioside, isoliquiritin, liquiritigenin, glycyrrhizic acid, isoliquiritigenin and glycyrrhetinic acid) of SGT were simultaneously determined by HPLC. SGT treatment (5.0 and 10.0 g/kg) was found to reduce the IgE level (a vital marker of hyper-responsiveness) in BALF and serum, improve typical morphological changes (inflammatory-cell infiltration and goblet cell metaplasia) in the lung and colon, alleviate airway remodeling (including bronchiostenosis and basement membrane-thickening) in the lung, significantly decrease the IL-4 level and increase the IFN-γ level in the lung and colon, which led to restoration of the IFN-γ/IL-4 ratio. The dysbiosis and dysfunction of GM in RSAs were modulated by SGT. The abundance of bacteria of the genera Ethanoligenens and Harryflintia was increased in RSAs and was decreased upon SGT treatment. The abundance of Family_XIII_AD3011_group was decreased in RSAs and increased upon SGT treatment. Moreover, SGT therapy increased the abundance of bacteria of the genera Ruminococcaceae_UCG-005 and Candidatus_Sacchrimonas, and decreased that of Ruminococcus_2 and Alistipes. CONCLUSIONS: SGT ameliorated rats with OVA-induced asthma via regulation of the Th1/Th2 ratio in the lung and gut, and modulated the GM.

Modified Dingchuan Decoction treats cough-variant asthma by suppressing lung inflammation and regulating the lung microbiota.

ETHNOPHARMACOLOGICAL RELEVANCE: Modified Dingchuan Decoction (MDD) is a Chinese medicine formula containing 11 materials with cough suppression, asthma relief, and anti-inflammatory effects. AIM OF THE STUDY: This study aimed to evaluate the therapeutic effect of MDD on cough-variant asthma (CVA) and to investigate its mechanism of action. MATERIALS AND METHODS: The chemical constituents of MDD were analyzed by ultra-performance liquid chromatography-quadrupole/electrostatic field orbitrap high-resolution mass spectrometry (UPLC-Q-Orbitrap HRMS). A guinea pig CVA model was established using an intramuscular injection of ovalbumin (OVA), combined with an intraperitoneal injection of aluminum hydroxide [Al(OH)3] and nebulized OVA. At the beginning of day 18, the low, medium, and high MDD groups were gavaged with 7.23 g/kg, 14.46 g/kg, and 28.92 g/kg of MDD, respectively, and the positive group was gavaged with 5 mg/kg of prednisone acetate combined with 1 mg/kg of montelukast sodium; the normal and model groups were given an equal volume of distilled water, once a day for 21 days. The cough was induced by 10-3 mol/L capsaicin solution 1 h after the last administration, and the number of coughs and the latency of coughs were evaluated. Hematoxylin and eosin staining (H&E) was used to observe pathological changes in the lungs and airways. The concentration of inflammatory factors in bronchoalveolar lavage fluid (BALF) was measured by enzyme-linked immunosorbent assay (ELISA). We analyzed the lung microbiota using 16 S ribosomal DNA (16 S rDNA) high-throughput sequencing. RESULTS: The 38 chemical components were found in MDD, and MDD reduced the number of coughs in guinea pigs with CVA, prolonged cough latency, improved pathological damage to the lungs and airways, regulated inflammatory factor levels in BALF, and modulated the lung microbiota. CONCLUSIONS: This study demonstrated that treating CVA with MDD may be related to inhibiting lung inflammation and regulating lung microbiota.

The extract from Hyssopus cuspidatus Boriss. Prevents bronchial airway remodeling by inhibiting mouse bronchial wall thickening and hASMC proliferation and migration.

ETHNOPHARMACOLOGICAL RELEVANCE: Bronchial asthma, a non-communicable chronic respiratory disease, affects people of all ages. An important pathological feature of bronchial asthma is airway remodeling. Hyssopus cuspidatus Boriss. has been used to treat bronchial asthma for over 100 years in Uygur medicine. The ethanol extract of Hyssopus cuspidatus Boriss.(JAX2) can improve airway inflammation in asthma. However, the anti-asthmatic airway-remodeling effect of JAX2 is unclear. AIM OF THE STUDY: The current study investigated the anti-airway remodeling effect of JAX2 and elucidated its mechanism of action. MATERIALS AND METHODS: The present study established an ovalbumin-induced mouse model of asthma and platelet-derived growth factor-BB-induced human airway smooth muscle cells (hASMCs) proliferation model, with dexamethasone (DEX) and feining tablets (FNP) designated as positive control drugs. Pathological changes in lung tissues were observed using hematoxylin and eosin staining. Interleukin (IL)-5, IL-10, IL-13, and IL-33 levels in the bronchoalveolar lavage fluid (BALF) and serum of mice were determined using enzyme-linked immunosorbent assay (ELISA). Changes in the expression and distribution of TGF-ß1, p-ERK1/2, Smad2/3, and p-Smad3 in lung tissues were determined using immunohistochemistry. Western blotting (WB) was used to determine the protein levels of p-ERK1/2 in lung tissues and cells. MTS assay was used to determine the effects of JAX2 on cell proliferation. IL-5, IL-10, IL-13, MMP-2, and MMP-9 levels in the cell supernatant were determined using ELISA. HASMCs migration was observed using the scratch and transwell methods. The effect of JAX2 on the hASMCs cycle was determined using flow cytometry. RESULTS: JAX2 significantly improved the pathological status of lung tissues in asthmatic mice. It could also significantly reduce IL-5, IL-13, and IL-33 levels in the BALF and serum of asthmatic mice in a dose-dependent manner and significantly increase IL-10 levels. TGF-ß1, p-ERK1/2, Smad2/3, and p-Smad3 expression in lung tissues were decreased in a dose-dependent manner. The protein level of p-ERK1/2 in lung tissues was also reduced. JAX2 could significantly inhibit the proliferation and migration of PDGF-BB-induced hASMCs. IL-5, IL-13, MMP-9, and MMP-2 levels decreased significantly, and IL-10 levels increased significantly in a dose-dependent manner in the cell supernatant. JAX2 could block hASMCs in the G0/G1 phase, thereby inhibiting cell proliferation. p-ERK1/2 protein levels were found to decrease in a dose-dependent manner. CONCLUSIONS: JAX2 significantly inhibits airway remodeling in asthma. Its mechanism of action may be inhibiting the proliferation and migration of hASMCs, releasing inflammatory factors and metalloproteinases, activating the ERK1/2 signal pathway, and promoting the secretion of anti-inflammatory factors.

Anti-asthmatic activity of standardized hydro-ethanolic and aqueous extracts of Stachytarpheta cayennensis (Rich.) Vahl in a murine model.

ETHNOPHARMACOLOGICAL RELEVANCE: Stachytarpheta cayennensis (Verbenaceae) has been used in Brazilian traditional medicine to treat asthma and other respiratory diseases. AIMS OF THE STUDY: To investigate the effects of different doses of standardized hydro-ethanolic (SCH) and aqueous (SCA) extracts of aerial parts of S. cayennensis using a murine ovalbumin (OVA)-induced asthma model. MATERIALS AND METHODS: The major constituents of the plant extracts were identified and standardized by ultra-performance liquid chromatography coupled with mass spectrometry. Balb/c mice were challenged with OVA solution and treated concomitantly by intraperitoneal injection of standardized SCH or SCA extracts at 50, 100, and 200 mg/kg concentrations. OVA-challenged control animals were treated with either dexamethasone (OVA-DEX) or saline solution (OVA-SAL). After challenge, we assessed in vivo bronchial hyperresponsiveness, airway inflammation (number of cells), peribronchial inflammation (histological analysis) and production of OVA-specific IgE and interleukin (IL)-4, IL-5, and IL-13 (ELISA). RESULTS: Acteoside, isoacteoside, and ipolamiide were the major constituents of SCH and SCA. The respective concentrations of acteoside in SCH and SCA were 78 and 98 µg/mL, while those of ipolamiide were 30 and 19 µg/mL. Treatment with 200 mg/kg of SCH or SCA decreased IL-4, IL-5, and IL-13 in lung homogenates. These reductions were accompanied by a lower influx of inflammatory cells (eosinophils, lymphocytes, and macrophages) to the airways and lungs. In addition to the anti-inflammatory effects, administration of SCA, but not SCH, ameliorated the parameters of bronchial hyperresponsiveness and decreased levels of circulating OVA-specific IgE. CONCLUSION: The results presented herein demonstrate for the first time the anti-asthmatic activity of S. cayennensis extracts in a murine model, thereby supporting the ethnopharmacological uses of the plant.

Effects of Houpo Mahuang Decoction on serum metabolism and TRPV1/Ca2+/TJs in asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Houpo Mahuang Decoction (HPMHD is one of the classic traditional Chinese prescriptions that has been used in the treatment of asthma. The therapeutic effects and mechanism of HPMHD in aggravated asthma remain to be explored, especially from the perspective of metabolomics and Transient Receptor Potential Vanilloid-1 (TRPV1)/Ca2+/Tight junction (TJ) regulation. AIM OF THE STUDY: To investigate the therapeutic and metabolic regulatory effects and the underlying mechanism of HPMHD in asthmatic rats. MATERIALS AND METHODS: The asthmatic rats were administered with the corresponding HPMHD (at dosages of 5.54, 11.07, 22.14 mg/kg). Then inflammatory cells in peripheral blood and bronchoalveolar lavage fluid (BALF) were counted, the levels of interleukin (IL)-4 and IL-13 in BALF were measured, and the changes in enhanced pause (Penh) and pathological damage of lung tissues were also detected to evaluate the protective effects of HPMHD. The serum metabolic profile of HPMHD in asthmatic rats was explored using Ultra-High-Performance Liquid Chromatography-mass spectrometer (UHPLC-MS), and the regulatory effects on TRPV1 and TJs of HPMHD in asthmatic rats were detected by Western blotting analysis. In vitro, 16HBE cells were stimulated with IL-4 plus SO2 derivatives and then administered HPMHD. The intracellular Ca2+ regulated by TRPV1, and the expression levels of TRPV1 and TJ proteins (TJs) were then detected by calcium imaging and Western blotting. The effects were verified by inhibition of TRPV1 and in short hairpin RNA (shRNA)-mediated TRPV1 silencing cells. RESULTS: HPMHD significantly attenuated the airway inflammation of asthmatic rats, and reduced the levels of inflammatory cells in peripheral blood and BALF as well as the levels of IL-4 plus IL-13 in BALF. In addition, the airway hyperresponsiveness and lung pathological damage were alleviated. Serum metabolomic analysis showed that 31 metabolites were differentially expressed among the normal saline-, model-, and HPMHD-treated rats. Pathway enrichment analysis showed that the metabolites were involved in 45 pathways, among which, TJs regulation-relevant pathway was associated with the Ca2+ concentration change mediated by the TRP Vanilloid channel. In vivo and in vitro experiments indicated that HPMHD reduced the concentration of intracellular Ca2+ via suppressing the expression and activation of TRPV1, increased the expression of ZO-1, Occludin, and Claudin-3, and protected the integrity of TJs. CONCLUSION: The current study indicates that HPMHD alleviates rat asthma and participates in the regulation of serum metabolism. The anti-asthma effects of HPMHD might be related to the protection of TJs by inhibiting the intracellular Ca2+ concentration via TRPV1.

Artemisia gmelinii Extract Alleviates Allergic Airway Inflammation via Balancing TH1/TH2 Homeostasis and Inhibiting Mast Cell Degranulation.

A new terminology "combined allergic rhinitis and asthma syndrome (CARAS)" was introduced to describe patients suffering from both allergic rhinitis (AR) and asthma. The pathogenesis of allergic airway inflammation has been well known, with the main contribution of TH1/TH2 imbalance and mast cell degranulation. Artemisia gmelinii has been used as an herbal medicine with its hepaprotective, anti-inflammatory, and antioxidant properties. In this study, the effect of A. gmelinii extracts (AGE) on the ovalbumin (OVA)-induced CARAS mouse model was investigated. AGE administration significantly alleviated the nasal rubbing and sneezing, markedly down-regulated both OVA-specific IgE, IgG1, and histamine levels, and up-regulated OVA-specific IgG2a in serum. The altered histology of nasal and lung tissues of CARAS mice was effectively ameliorated by AGE. The AGE treatment group showed markedly increased levels of the TH1 cytokine interleukin (IL)-12 and TH1 transcription factor T-bet. In contrast, the levels of the TH2 cytokines, including IL-4, IL-5, IL-13, and the TH2 transcription factor GATA-3, were notably suppressed by AGE. Moreover, AGE effectively prevented mast cell degranulation in vitro and mast cell infiltration in lung tissues in vivo. Based on these results, we suggest that AGE could be a potential therapeutic agent in OVA-induced CARAS by virtue of its role in balancing the TH1/TH2 homeostasis and inhibiting the mast cell degranulation.

Antiallergic Effects of N,N-dicoumaroylspermidine Isolated from Lithospermum erythrorhizon on Mast Cells and Ovalbumin-Induced Allergic Rhinitis.

In East Asia, the dried root of Lithospermum erythrorhizon has been utilized as an anti-inflammatory, antipyretic, detoxifying, and anti-inflammatory agent. Recently, we reported that L. erythrorhizon protects against allergic rhinitis; however, the component within L. erythrorhizon that exerts antiallergic activity remains unknown. The purpose of the current study was to isolate and characterize the antiallergic active components in an ethanolic extract of L. erythrorhizon roots. We examined the antiallergic effects of L. erythrorhizon reflux ethanol extracts in an ovalbumin (OVA)-induced allergic rhinitis mouse model, and compared the chemical compounds extracted using the hot reflux and cold extraction methods. Chromatographic separation identified two novel anthraquinones, erythrin A and B, one newly discovered compound from the Lithospermum genus, N1″,N3″-dicoumaroylspermidine, and nineteen other recognized compounds. Their chemical structures were elucidated by single (1D) and 2D analysis of nuclear magnetic resonance (NMR) spectroscopic data, as well as high resolution mass spectrometry. Among the identified compounds, N,N'-dicoumaroylspermidine strongly inhibited the release of ß-hexosaminidase, as well as the production of IL-3, IL-4, and IL-13 by IgE-sensitized and BSA-stimulated RBL-2H3 cells. Using the OVA-induced allergic rhinitis mouse model, we showed that N,N'-dicoumaroylspermidine reduced the production of serum OVA-specific IgE and the number of inflammatory cells in nasal lavage fluid. N,N'-dicoumaroylspermidine isolated from L. erythrorhizon exhibits antiallergic properties, making it potentially effective for allergic rhinitis.

[Effect of heat-reinforcing needling on expression of serum inflammatory factors and autophagy of knee joint synovial tissue in rheumatoid arthritis rabbits with cold syndrome].

OBJECTIVE: To observe the effect of heat-reinforcing needling on the expression of serum inflammatory factors and autophagy of knee synovial tissue in rheumatoid arthritis (RA) rabbits with cold syndrome, so as to explore its mechanism of anti-inflammatory in the treatment of RA. METHODS: Fifty rabbits were randomly divided into normal, model, heat-reinforcing needling, inhibitor and agonist groups (n=10 rabbits in each group). The model of RA with cold syndrome was established by Freund's adjuvant and ovalbumin mixed solution injection combined with freezing and wind-cold dampness method. Heat-reinforcing needling was applied at "Zusanli" (ST36) for 30 min, once a day for 14 days. Rabbits of the inhibitor and agonist groups were given intraperitoneally injected with autophagy inhibitor 3-methyladenine (3-MA) or autophagy agonist rapamycin, once every 2 days for 7 days. The knee circumference and skin temperature of the rabbits in each group were measured. Color doppler ultrasonography was applied to examine the synovial membrane, joint effusion and blood flow signals in the knee joints of the rabbits in each group. Serum tumor necrosis factor (TNF) -α, interleukin (IL)-1ß, IL-6 and C-creactive protein (CRP) were detected by ELISA. Transmission electron microscopy was applied to observe the ultrastructure and autophagosomes of synovial cells. The protein expressions of autophagy-related protein Atg5, serine/threonine protein kinase-dysregulated 51-like kinase 1 (ULK1), microtubule-associated protein light chain 3B (LC3B), and Beclin-1 were detected by Western blot. Fluorescence quantitative PCR was used to detect the mRNA expressions of NOD-like receptor 3 (NLRP3) and nuclear factor-κB (NF-κB). RESULTS: Compared with the normal group, the circumference of the knee joint was increased (P<0.01), the skin temperature was decreased (P<0.01), the knee joint synovium was thickened and the blood flow signal was abundant, the contents of serum TNF-α, IL-1ß, IL-6, and CRP were increased (P<0.01), the protein expressions of Atg5, ULK1, Beclin-1 and LC3BⅡ/LC3BⅠof synovial tissue were significantly decreased (P<0.01), the mRNA expressions of NLRP3 and NF-κB were increased (P<0.01) in the model group. In comparison with the model and inhibitor groups, the circumference of the knee joint was decreased (P<0.01), whlie the skin temperature was increased (P<0.01), the synovial membrane became thinner and the blood flow signal was wea-kened, the contents of TNF-α, IL-1ß, IL-6 and CRP were decreased (P<0.01), the protein expressions of Atg5, ULK1, Beclin-1 and LC3B Ⅱ/LC3B Ⅰ were increased (P<0.01), and the mRNA expressions of NLRP3 and NF-κB were decreased (P<0.01) in the heat-reinforcing needling and agonist groups. CONCLUSION: Heat-reinforcing needling can alleviate the inflammatory response of the knee joint synovium in RA rabbits with cold syndrome, which may be related to its function in enhancing the autophagy activity of synovial cells and inhibiting the synthesis and release of inflammatory factors TNF-α, IL-1ß, IL-6 and CRP.

HPLC-DAD analysis of Quercus leucotrichophora extract and appraisal of its antiasthmatic potential via modulation of aquaporins, inflammatory, and oxidative stress biomarkers in Albino mice.

Herbal drugs offer an alternative approach for the treatment of diseases like asthma due to low cost and comparatively less adverse effects in contrast to synthetic drugs. Leaves of Quercus leucotrichophora are traditionally used for the treatment of asthma. The study was aimed to assess the anti-asthmatic activity of Quercus leucotrichophora (QL) methanolic (QLME) and aqueous extracts (QLAE) in ovalbumin-(OVA) induced asthma and chemical characterization of QL extract by High Performance Liquid Chromatography-Diode array detector (HPLC-DAD). Animals were inoculated with OVA (i.p) on day 1 and 14 followed by intranasal challenge on 27th and 29th day. Both extracts of QL at 600, 300 and 150 mg/kg and dexamethasone (2 mg/kg) l were administered consecutively from days 15-26 via oral gavage. The QL extracts notably reduced (p < 0.0001-p < 0.05) total and differential leukocyte counts in blood and BALF and serum IgE levels in contrast to disease control. Both extracts and Dex substantially improved activities of superoxide dismutase, catalase, and GSH, while reduced malondialdehyde level in treated mice. Treatment with extracts and Dex caused significant (p < 0.0001-p < 0.05) downregulation of tumor necrosis factor-α, interleukin-4, - 5, - 13, - 6, - 1ß, and NF-κB whereas, increased expression of Aquaporin (AQP) 1 and AQP5 in contrast to disease control. It was inferenced from findings that both extract of QL exhibited notable antiasthmatic potential might be due to presence of Daidzein-glucuronic acid, 3-Hydroxyphloretin 6'-hexoside, Catechin, Quercetin, and Kaemferol.

Processed product (Pinelliae Rhizoma Praeparatum) of Pinellia ternata (Thunb.) Breit. Alleviates the allergic airway inflammation of cold phlegm via regulation of PKC/EGFR/MAPK/PI3K-AKT signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Pinelliae Rhizoma Praeparatum (PRP) is a traditional processed product of Pinellia ternata (Thunb.) Berit., which mainly used for treating cold asthma (CA). However, the mechanism of action of PRP for treating CA have not been fully elucidated. AIM OF THE STUDY: To investigate the core active constituents and the pharmacological mechanism of PRP against CA. MATERIALS AND METHODS: Ovalbumin (OVA) and cold water-induced cold asthma model were established in male mice. The effects of water extract from PRP were evaluated by general morphological observation, expectorant activity, airway hyperresponsiveness, mucus hypersecretion, inflammatory cytokines, etc. Additionally, the mRNA and protein expression of mucin 5AC (MUC5AC) and aquaporin 5 (AQP5) in vivo and in vitro were detected by immunohistochemistry (IHC), qRT-PCR, and western blotting. The mechanisms of action were investigated through network pharmacology and transcriptomic, and validated through western blotting and molecular docking. RESULTS: PRP exhibited a favorable expectorant activity, and significantly reduced the airway inflammation, mucus secretion, and hyperresponsiveness in cold asthma model. It also reduced the levels of IL-4, IL-5, IL-8, and IL-13 in bronchoalveolar lavage fluid (BALF) and IL-4 and total IgE in serum, while obviously increased the levels of IL-10 and IFN-γ in serum for asthmatic mice. Meanwhile, PRP also attenuated the pathological changes and mucus production in cold asthmatic mice. Moreover, the downregulation of MUC5AC and upregulation of AQP 5 were detected by western blotting and qRT-PCR after administration with PRP both in vivo and in vitro. PRP expectedly inhibited the protein expression of PKC-α, SRC, p-EGFR, p-ERK1/2, p-JNK, p-p38, p-PI3K, and p-Akt levels in vivo. CONCLUSIONS: These combined data showed that PRP suppressed the allergic airway inflammation of CA by regulating the balance of Th1 and Th2 cytokines and the possible involvement of the PKC/EGFR/MAPK/PI3K-Akt signaling pathway. Pentadecanoic acid, licochalcone A, ß-sitosterol, etc. were considered as main active ingredients of PRP against CA. This study provides a novel perspective of the classical herbal processed product PRP in the treatment of CA.

Effect of Houpo-Mahuang Decoction on aggravated asthma induced by cigarette smoke and the expression of TRPA1 and tight junctions in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Cigarette smoke (CS) is a common environmental irritant and a risk factor for asthma, as it induces as well as aggravates asthmatic attacks. The injured airway epithelial tight junctions (TJs) aggravate asthma. CS can aggravate asthma by activating the transient receptor potential ankyrin A1 (TRPA1) channel and enhancing TJs destruction. Houpo Mahuang decoction (HPMHD) is a classic traditional Chinese prescription for the treatment of asthma. However, its underlying action mechanism is unclear. AIM OF THE STUDY: The present study aimed to evaluate the effect of HPMHD on the asthma phenotype and the regulation of TRPA1 and TJs in a CS-induced mouse model of aggravated asthma. MATERIALS AND METHODS: Under optimized chromatographic and mass spectrometry conditions, the ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) technique was used to detect and analyze the major chemical components of HPMHD. C57BL/6 female mice were randomly divided into seven groups, viz, normal saline (NS) group, ovalbumin (OVA) + CS group, dexamethasone group, HPMHD high-dose group and low-dose groups, n-butanol extract group, and ethyl acetate extract group, with 10 mice in each group. OVA sensitization and challenge, and CS exposure were used to establish the aggravated asthma model. As the main indices to evaluate the protective effect of HPMHD, the eosinophils count in peripheral blood, percentages of inflammatory cells classified and the levels of interleukin (IL)-4, IL-5, IL-13 in the bronchoalveolar lavage fluid (BALF), airway responsiveness enhanced pause (Penh), and changes in lung histopathology were determined and compared among the groups. The mRNA and protein expression of TRPA1 and TJs in lung tissue was also examined. RESULTS: Using UPLC-QTOF-MS, the chemical components of HPMHD, including ephedrine, pseudoephedrine, laetrile, and amygdalin amide, were identified by 51 signal peaks. Compared with those in the NS group, the eosinophil number in the peripheral blood and the eosinophils and neutrophils percentages in BALF of the OVA + CS group were remarkably increased. Following the inhalation of 50 µl of acetylcholine chloride (ACH) at doses of 25 and 50 mg/mL, the Penh increased significantly (p < 0.01). Moreover, in the OVA + CS group, hematoxylin and eosin (H&E) staining of lung tissue showed a significant number of infiltrated inflammatory cells, increased mucus secretion in the lumen, damaged bronchial mucosa, increased thickness of tracheal wall, and increased score of lung damage (p < 0.01). The IL-4/5/13 levels were also remarkably increased (p < 0.01). The protein as well as gene expression of both ZO-1 and occludin decreased markedly in the lung tissue, while the expression of TRPA1 and claudin-2 was increased (p < 0.05, p < 0.01). Next, the OVA + CS group and the treatment groups were compared. The inflammatory cells, Penh value, and levels of IL-4/5/13 were significantly reduced, and less lung injury was observed in the treatment groups. The gene and protein levels of TRPA1 and TJs were corrected (p < 0.05, p < 0.01); the effects on the HPMHD high-dose and ethyl acetate extract groups were particularly remarkable. CONCLUSIONS: HPMHD reduced airway hyperresponsiveness, inflammatory cell recruitment and Th2 cytokine secretion in CS-induced aggravated asthma mice, in a manner potentially dependent on regulation of the expression of TRPA1 and TJ proteins. Both the n-butanol and ethyl acetate extracts contained the active ingredients, especially the ethyl acetate extract.

The effect of Ma-Xin-Gan-Shi decoction on asthma exacerbated by respiratory syncytial virus through regulating TRPV1 channel.

ETHNOPHARMACOLOGICAL RELEVANCE: The incidence and mortality of bronchial asthma are increasing, and respiratory syncytial virus (RSV) is widely regarded as the common cause of clinical exacerbation of asthma. Ma-Xing-Gan-Shi decoction (MXGSD), a classic traditional Chinese medicine prescription, is well-known for treating respiratory diseases, while the mechanism of effecting on RSV-exacerbated asthma remains to be explored. AIM OF THE STUDY: In this study, we investigated the mechanism by which MXGSD exerts a protective effect on asthma exacerbated by RSV in vivo and in vitro. MATERIALS AND METHODS: MXGSD is composed of four Chinese medicine, including Ephedra intermedia Schrenk & C.A.Mey. (herbaceous stem, 27g), Prunus armeniaca L. (dry seed, 27g), Glycyrrhiza uralensis Fisch. (radix and rhizome, 18g), and Gypsum fibrosum (main component: CaSO4·2H2O, 54g). In the present study, the exacerbated asthmatic mice model with the treatment of OVA plus RSV was replicated, and accompanied by the TMT proteomic analysis and further experimental investigations. Then, the protective effect of MXGSD (13.2, 6.6, 3.3 g/kg/d, 7d) on the mice treated by OVA plus RSV, and the mechanism of regulating TRPV1 was explored. In addition, the intracellular Ca2+ concentration of 16HBE cells pretreated with MXGSD medicated serum was also tested after stimulation with the TRPV1 agonist capsaicin. RESULTS: The results suggested that MXGSD could reduce the levels of inflammation cells, airway hyperresponsiveness, and pathological damage of lung tissue. TMT quantitative proteomics analysis and further experimental exploration revealed that MXGSD could reduce the levels of IL-4, IL-13, PGE2, and SP in BAL and down-regulate the expression of TRPV1 mRNA and protein in lung tissue. Furthermore, 16HBE cells stimulated by capsaicin showed an increased intracellular Ca2+ concentration, while the pretreatment of MXGSD medicated serum could reduce it. CONCLUSION: MSGSD showed a protective effect on RSV-exacerbated asthma, which may be related to its regulation of TRPV1 expression and reduction of Th2 cytokines and neurogenic inflammatory mediators. It may provide an objective basis and reference for the clinical application of MXGSD.

Polystichum braunii ameliorates airway inflammation by attenuation of inflammatory and oxidative stress biomarkers, and pulmonary edema by elevation of aquaporins in ovalbumin-induced allergic asthmatic mice.

Asthma is a chronic inflammation of pulmonary airways associated with bronchial hyper-responsiveness. The study was aimed to validate the folkloric use of Polystichum braunii (PB) against ovalbumin (OVA)-induced asthmatic and chemical characterization OF both extracts. Allergic asthma was developed by intraperitoneal sensitization with an OVA on days 1 and 14 followed by intranasal challenge. Mice were treated with PB methanolic (PBME) and aqueous extract (PBAE) orally at 600, 300, and 150 mg/kg and using dexamethasone (2 mg/kg) as standard from day 15 to 26. High performance liquid chromatography-diode array detector analysis revealed the presence of various bioactive compounds such as catechin, vanillic acid, and quercetin. The PBME and PBAE profoundly (p < 0.0001-0.05) declined immunoglobulin E level, lungs wet/dry weight ratio, and total and differential leukocyte count in blood and bronchial alveolar lavage fluid of treated mice in contrast to disease control. Histopathological examination showed profoundly decreased inflammatory cell infiltration and goblet cell hyperplasia in treated groups. Both extracts caused significant (p < 0.0001-0.05) diminution of IL-4, IL-5, IL-13, IL-6, IL-1ß, TNF-α, and NF-κB and upregulation of aquaporins (1 and 5), which have led to the amelioration of pulmonary inflammation and attenuation of lung edema in treated mice. Both extracts profoundly (p < 0.0001-0.05) restored the activities of SOD, CAT, GSH and reduced the level of MDA dose dependently. Both extracts possessed significant anti-asthmatic action mainly PBME 600 mg/kg might be due to phenols and flavonoids and could be used as a potential therapeutic option in the management of allergic asthma.