Combined Treatment of 6-Gingerol Analog and Tobramycin for Inhibiting Pseudomonas aeruginosa Infections.

Pseudomonas aeruginosa is a ubiquitous human pathogen that causes severe infections. Although antibiotics, such as tobramycin, are currently used for infection therapy, their antibacterial activity has resulted in the emergence of multiple antibiotic-resistant bacteria. The 6-gingerol analog, a structural derivative of the main component of ginger, is a quorum sensing (QS) inhibitor. However, it has a lower biofilm inhibitory activity than antibiotics and the possibility to cause toxicity in humans. Therefore, novel and more effective approaches for decreasing dosing concentration and increasing biofilm inhibitory activity are required to alleviate P. aeruginosa infections. In this study, a 6-gingerol analog was combined with tobramycin to treat P. aeruginosa infections. The combined treatment of 6-gingerol analog and tobramycin showed strong inhibitory activities on biofilm formation and the production of QS-related virulence factors of P. aeruginosa compared to single treatments. Furthermore, the combined treatment alleviated the infectivity of P. aeruginosa in an insect model using Tenebrio molitor larvae without inducing any cytotoxic effects in human lung epithelial cells. The 6-gingerol analog showed these inhibitory activities at much lower concentrations when used in combination with tobramycin. Adjuvant effects were observed through increased QS-disrupting processes rather than through antibacterial action. In particular, improved RhlR inactivation by this combination is a possible target for therapeutic development in LasR-independent chronic infections. Therefore, the combined treatment of 6-gingerol analog and tobramycin may be considered an effective method for treating P. aeruginosa infections. IMPORTANCE Pseudomonas aeruginosa is a pathogen that causes various infectious diseases through quorum-sensing regulation. Although antibiotics are mainly used to treat P. aeruginosa infections, they cause the emergence of resistant bacteria in humans. To compensate for the disadvantages of antibiotics and increase their effectiveness, natural products were used in combination with antibiotics in this study. We discovered that combined treatment with 6-gingerol analog from naturally-derived ginger substances and tobramycin resulted in more effective reductions of biofilm formation and virulence factor production in P. aeruginosa than single treatments. Our findings support the notion that when 6-gingerol analog is combined with tobramycin, the effects of the analog can be exerted at much lower concentrations. Furthermore, its improved LasR-independent RhlR inactivation may serve as a key target for therapeutic development in chronic infections. Therefore, the combined treatment of 6-gingerol analog and tobramycin is suggested as a novel alternative for treating P. aeruginosa infections.

Drug repurposing for COVID-19 based on an integrative meta-analysis of SARS-CoV-2 induced gene signature in human airway epithelium.

Drug repurposing has the potential to bring existing de-risked drugs for effective intervention in an ongoing pandemic-COVID-19 that has infected over 131 million, with 2.8 million people succumbing to the illness globally (as of April 04, 2021). We have used a novel `gene signature'-based drug repositioning strategy by applying widely accepted gene ranking algorithms to prioritize the FDA approved or under trial drugs. We mined publically available RNA sequencing (RNA-Seq) data using CLC Genomics Workbench 20 (QIAGEN) and identified 283 differentially expressed genes (FDR<0.05, log2FC>1) after a meta-analysis of three independent studies which were based on severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) infection in primary human airway epithelial cells. Ingenuity Pathway Analysis (IPA) revealed that SARS-CoV-2 activated key canonical pathways and gene networks that intricately regulate general anti-viral as well as specific inflammatory pathways. Drug database, extracted from the Metacore and IPA, identified 15 drug targets (with information on COVID-19 pathogenesis) with 46 existing drugs as potential-novel candidates for repurposing for COVID-19 treatment. We found 35 novel drugs that inhibit targets (ALPL, CXCL8, and IL6) already in clinical trials for COVID-19. Also, we found 6 existing drugs against 4 potential anti-COVID-19 targets (CCL20, CSF3, CXCL1, CXCL10) that might have novel anti-COVID-19 indications. Finally, these drug targets were computationally prioritized based on gene ranking algorithms, which revealed CXCL10 as the common and strongest candidate with 2 existing drugs. Furthermore, the list of 283 SARS-CoV-2-associated proteins could be valuable not only as anti-COVID-19 targets but also useful for COVID-19 biomarker development.

The therapeutic potential of inorganic polyphosphate: A versatile physiological polymer to control coronavirus disease (COVID-19).

Rationale: The pandemic caused by the novel coronavirus SARS-CoV-2 is advancing rapidly. In particular, the number of severe courses of the disease is still dramatically high. An efficient drug therapy that helps to improve significantly the fatal combination of damages in the airway epithelia, in the extensive pulmonary microvascularization and finally multiorgan failure, is missing. The physiological, inorganic polymer, polyphosphate (polyP) is a molecule which could prevent the initial phase of the virus life cycle, the attachment of the virus to the target cells, and improve the epithelial integrity as well as the mucus barrier. Results: Surprisingly, polyP matches perfectly with the cationic groove on the RBD. Subsequent binding studies disclosed that polyP, with a physiological chain length of 40 phosphate residues, abolishes the binding propensity of the RBD to the ACE2 receptor. In addition to this first mode of action of polyP, this polymer causes in epithelial cells an increased gene expression of the major mucins in the airways, of MUC5AC and MUC1, as well as a subsequent glycoprotein production. MUC5AC forms a gel-like mucus layer trapping inhaled particles which are then transported out of the airways, while MUC1 constitutes the periciliary liquid layer and supports ciliary beating. As a third mode of action, polyP undergoes enzymatic hydrolysis of the anhydride bonds in the airway system by alkaline phosphatase, releasing metabolic energy. Conclusions: This review summarizes the state of the art of the biotherapeutic potential of the polymer polyP and the findings from basic research and outlines future biomedical applications.

Effect of Korean Red Ginseng and Rg3 on Asian Sand Dust-Induced MUC5AC, MUC5B, and MUC8 Expression in Bronchial Epithelial Cells.

Korean Red ginseng (KRG), commonly used in traditional medicine, has anti-inflammatory, anti- oxidative, and anti-tumorigenic properties. Asian sand dust (ASD) is known to aggravate upper and lower airway inflammatory responses. BEAS-2B cells were exposed to ASD with or without KRG or ginsenoside Rg3. Mucin 5AC (MUC5AC), MUC5B, and MUC8 mRNA and protein expression levels were determined using quantitative RT-PCR and enzyme-linked immunosorbent assay. Nuclear factor kappa B (NF-κB), activator protein 1, and mitogen-activated protein kinase expression and activity were determined using western blot analysis. ASD induced MUC5AC, MUC5B, and MUC8 mRNA and protein expression in BEAS-2B cells, which was significantly inhibited by KRG and Rg3. Although ASD-induced mucin expression was associated with NF-κB and p38 mitogen-activated protein kinase (MAPK) activity, KRG and Rg3 significantly suppressed only ASD-induced NF-κB expression and activity. KRG and Rg3 inhibited ASD-induced mucin gene expression and protein production from bronchial epithelial cells. These results suggest that KRG and Rg3 have potential for treating mucus-producing airway inflammatory diseases.

Pirfenidone mediates cigarette smoke extract induced inflammation and oxidative stress in vitro and in vivo.

BACKGROUND: Antioxidant and anti-inflammatory effects are two main pharmacological mechanisms of pirfenidone (PFD) besides the anti-fibrotic effect. This study aims to investigate whether PFD could mediate cigarette smoke extract (CSE) induced inflammation and oxidative stress in vitro and in vivo. METHODS: BALB/C mice and alveolar epithelial (A549) cells treated with CSE were established as disease models in vivo and in vitro. Effects of PFD treatment on disease models were further measured. Hematoxylin and eosin (HE) staining was used to evaluate the pathological changes in lung tissues of mice. CCK-8 assay kit was applied to measure the viability of A549 cells treated by different concentrations of PFD. Inflammation cytokine expression in cell supernatants was measured with ELISA kits. The mRNA and protein levels of inflammation and oxidative stress-related factors were determined by real-time quantitative polymerase chain reaction analysis (RT-qPCR) and Western blotting. Furthermore, myeloperoxidase (MPO), malondialdehyde (MDA), and total antioxidant capacity (T-AOC) were measured to detect the antioxidative activity of lung tissues. Moreover, an assay kit with fluorescent probe 2',7'-dichlorofluorescin diacetate (DCFH-DA) was used to evaluate the intracellular reactive oxygen species (ROS) generation. RESULTS: In vitro and in vivo, PFD significantly reversed TNF-α, IL-6, CCL2, SOD1, and CAT mRNA level changes led by CSE; in addition, PFD significantly decreased the ratios of p-p65 to p65, p-ikBα to ikBα and increased Nrf-2 protein level compared with CSE group. In mice, high-dose (100 mg/kg/d) PFD significantly reversed MPO and MDA increases induced by CSE. However, PFD didn't significantly reverse T-AOC decrease induced by CSE. In A549 cell supernatant, PFD dramatically reversed the elevated levels of TNF-α and IL-1ß induced by CSE. Furthermore, PFD could significantly reverse the increased level of ROS induced by CSE in A549 cells. CONCLUSION: Our study reveals the potential role of PFD in regulating inflammatory response and oxidative stress induced by CSE.

The MAPKinase Signaling and the Stimulatory Protein-1 (Sp1) Transcription Factor Are Involved in the Phototherapy Effect on Cytokines Secretion from Human Bronchial Epithelial Cells Stimulated with Cigarette Smoke Extract.

The present study was aimed to investigate the phototherapy effect with low-level laser on human bronchial epithelial cells activated by cigarette smoke extract (CSE). Phototherapy has been reported to actuate positively for controlling the generation/release of anti-inflammatory and pro-inflammatory mediators from different cellular type activated by distinct stimuli. It is not known whether the IL-8 and IL-10 release from CSE-stimulated human bronchial epithelium (BEAS) cells can be influenced by phototherapy. Human bronchial epithelial cell (BEAS) line was cultured in a medium with CSE and irradiated (660 nm) at 9 J. Apoptosis index was standardized with Annexin V and the cellular viability was evaluated by MTT. IL-8, IL-10, cAMP, and NF-κB were measured by ELISA as well as the Sp1, JNK, ERK1/2, and p38MAPK. Phototherapy effect was studied in the presence of mithramycin or the inhibitors of JNK or ERK. The IL-8, cAMP, NF-κB, JNK, p38, and ERK1/2 were downregulated by phototherapy. Both the JNK and the ERK inhibitors potentiated the phototherapy effect on IL-8 as well as on cAMP secretion from BEAS. On the contrary, IL-10 and Sp1 were upregulated by phototherapy. The mithramycin blocked the phototherapy effect on IL-10. The results suggest that phototherapy has a dual effect on BEAS cells because it downregulates the IL-8 secretion by interfering with CSE-mediated signaling pathways, and oppositely upregulates the IL-10 secretion through of Sp1 transcription factor. The manuscript provides evidence that the phototherapy can interfere with MAPK signaling via cAMP in order to attenuate the IL-8 secretion from CSE-stimulated BEAS. In addition, the present study showed that phototherapy effect is driven to downregulation of the both the IL-8 and the ROS secretion and at the same time the upregulation of IL-10 secretion. Besides it, the increase of Sp-1 transcription factor was crucial for laser effect in upregulating the IL-10 secretion. The dexamethasone corticoid produces a significant inhibitory effect on IL-8 as well as ROS secretion, but on the other hand, the corticoid blocked the IL-10 secretion. Taking it into consideration, it is reasonable to suggest that the beneficial effect of laser therapy on lung diseases involves its action on unbalance between pro-inflammatory and anti-inflammatory mediators secreted by human bronchial epithelial cells through different signaling pathway.

Retinoic acid improves baseline barrier function and attenuates TNF-α-induced barrier leak in human bronchial epithelial cell culture model, 16HBE 14o.

Retinoic acid (RA) has been shown to improve epithelial and endothelial barrier function and development and even suppress damage inflicted by inflammation on these barriers through regulating immune cell activity. This paper thus sought to determine whether RA could improve baseline barrier function and attenuate TNF-α-induced barrier leak in the human bronchial epithelial cell culture model, 16HBE14o- (16HBE). We show for the first time that RA increases baseline barrier function of these cell layers indicated by an 89% increase in transepithelial electrical resistance (TER) and 22% decrease in 14C-mannitol flux. A simultaneous, RA-induced 70% increase in claudin-4 attests to RA affecting the tight junctional (TJ) complex itself. RA was also effective in alleviating TNF-α-induced 16HBE barrier leak, attenuating 60% of the TNF-α-induced leak to 14C-mannitol and 80% of the leak to 14C-inulin. Interleukin-6-induced barrier leak was also reduced by RA. Treatment of 16HBE cell layers with TNF-α resulted in dramatic decrease in immunostaining for occludin and claudin-4, as well as a downward "band-shift" in occludin Western immunoblots. The presence of RA partially reversed TNF-α's effects on these select TJ proteins. Lastly, RA completely abrogated the TNF-α-induced increase in ERK-1,2 phosphorylation without significantly decreasing the TNF-driven increase in total ERK-1,2. This study suggests RA could be effective as a prophylactic agent in minimizing airway barrier leak and as a therapeutic in preventing leak triggered by inflammatory cascades. Given the growing literature suggesting a "cytokine storm" may be related to COVID-19 morbidity, RA may be a useful adjuvant for use with anti-viral therapies.

In search of preventive strategies: novel high-CBD Cannabis sativa extracts modulate ACE2 expression in COVID-19 gateway tissues.

With the current COVID-19 pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), there is an urgent need for new therapies and prevention strategies that can help curtail disease spread and reduce mortality. The inhibition of viral entry and thus spread is a plausible therapeutic avenue. SARS-CoV-2 uses receptor-mediated entry into a human host via the angiotensin-converting enzyme 2 (ACE2), which is expressed in lung tissue as well as the oral and nasal mucosa, kidney, testes and gastrointestinal tract. The modulation of ACE2 levels in these gateway tissues may be an effective strategy for decreasing disease susceptibility. Cannabis sativa, especially those high in the anti-inflammatory cannabinoid cannabidiol (CBD), has been found to alter gene expression and inflammation and harbour anti-cancer and anti-inflammatory properties. However, its effects on ACE2 expression remain unknown. Working under a Health Canada research license, we developed over 800 new C. sativa cultivars and hypothesized that high-CBD C. sativa extracts may be used to down-regulate ACE2 expression in target COVID-19 tissues. Using artificial 3D human models of oral, airway and intestinal tissues, we identified 13 high-CBD C. sativa extracts that decrease ACE2 protein levels. Some C. sativa extracts down-regulate serine protease TMPRSS2, another critical protein required for SARS-CoV-2 entry into host cells. While our most effective extracts require further large-scale validation, our study is important for future analyses of the effects of medical cannabis on COVID-19. The extracts of our most successful novel high-CBD C. sativa lines, pending further investigation, may become a useful and safe addition to the prevention/treatment of COVID-19 as an adjunct therapy.

Sanguinarine Rapidly Relaxes Rat Airway Smooth Muscle Cells Dependent on TAS2R Signaling.

Excessive contraction of airway smooth muscle cells (ASMCs) is a hallmark feature of asthma. Intriguing, the activation of bitter taste receptor (TAS2R) in ASMCs can relax ASMCs. However, there is a lack of potent TAS2R agonists that can be used in asthma therapies since those tested agonists cannot relax ASMCs at the dose below a few hundred micromolar. Considering that sanguinarine (SA) is a bitter substance often used in small doses for the treatment of asthma in folk medicine, the present study was to determine the rapid relaxation effect of SA on ASMCs and to reveal the underlying mechanisms associated with TAS2R signaling. Here, cell stiffness, traction force, calcium signaling, cAMP levels, and the mRNA expression were evaluated by using optical magnetic twisting cytometry, traction force microscopy, Fluo-4/AM labeling, enzyme-linked immunosorbent assay (ELISA), and quantitative (q)RT-PCR, respectively. We found that 0.5 µM SA immediately decreased cell stiffness and traction force, which is comparable with the effect of 5 µM isoproterenol. In addition, 0.5 µM SA immediately increased intracellular free calcium concentration ([Ca2+]i) and decreased the mRNA expression of contractile proteins such as calponin and α-smooth muscle actin after the treatment for 24 h. Furthermore, SA-mediated decrease in cell stiffness/traction force and increase in [Ca2+]i were significantly blunted by inhibiting the TAS2Rs signaling. These findings establish the rapid relaxation effect of SA at low concentration (<1 µM) on cultured ASMCs depending on TAS2R signaling, indicating that SA might be developed as a useful bronchodilator in asthma therapy.

Arsenic represses airway epithelial mucin expression by affecting retinoic acid signaling pathway.

Arsenic is a ubiquitous environmental toxicant, found in high concentrations worldwide. Although abundant research has dealt with arsenic-induced cancers, studies on mechanisms of non-malignant lung diseases have not been complete. In addition, decades of research have mostly concentrated on high-dose arsenic exposure, which has very limited use in modeling the biological effects of today's low-dose exposures. Indeed, accumulated evidence has shown that low-dose arsenic exposure (i.e. ≤100 ppb) may also alter lung homeostasis by causing host susceptibility to viral infection. However, the underlying mechanism of this alteration is unknown. In this study, we found that low-dose sodium arsenite (As (III)) repressed major airway mucins-MUC5AC and MUC5B at both mRNA and protein levels. We further demonstrated that this repression was not caused by cellular toxicity or mediated by the reduction of a common mucin-inducing pathway-EGFR. Other established mucin activators- dsRNA, IL1ß or IL17 were not able to override As (III)-induced mucin repression. Interestingly, the suppressing effect of As (III) appeared to be partially reversible, and supplementation of all trans retinoic acid (t-RA) doses dependently restored mucin gene expression. Further analyses indicated that As (III) treatment significantly reduced the protein level of retinoic acid receptors (RARα, γ and RXRα) as well as RARE promoter reporter activity. Therefore, our study fills in an important knowledge gap in the field of low-dose arsenic exposure. The interference of RA signaling, and mucin gene expression may be important pathogenic factors in low-dose arsenic induced lung toxicity.

Systems pharmacology-based study of Tanreqing injection in airway mucus hypersecretion.

ETHNOPHARMACOLOGICAL RELEVANCE: Mucus hypersecretion (MH) is recognized as a key pathophysiological and clinical feature of many airway inflammatory diseases. MUC5AC is a major component of airway mucus. Tanreqing injection (TRQ) is a widely used herbal formula for the treatment of respiratory inflammations for years in China. However, a holistic network pharmacology approach to understanding its therapeutic mechanisms against MH has not been pursued. AIM OF THE STUDY: This study aimed to explore the systems-level potential active compounds and therapeutic mechanisms of TRQ in the treatment of MH. MATERIALS AND METHODS: We established systems pharmacology-based strategies comprising compound screenings, target predictions, and pathway identifications to speculate the potential active compounds and therapeutic targets of TRQ. We also applied compound-target and target-disease network analyses to evaluate the possible action mechanisms of TRQ. Then, lipopolysaccharide (LPS)-induced Sprague-Dawley (SD) rat model was constructed to assess the effect of TRQ in the treatment of MH and to validate the possible molecular mechanisms as predicted in systems pharmacology approach. RESULTS: The comprehensive compound collection successfully generated 55 compound candidates from TRQ. Among them, 11 compounds with high relevance to the potential targets were defined as representative and potential active ingredients in TRQ formula. Target identification revealed 172 potential targets, including pro-inflammatory cytokines of tumor necrosis factor α (TNF-α), interleukin (IL)-6, and IL-8. Pathway analyses uncovered the possible action of TRQ in the regulation of IL-17 signaling pathway and its downstream protein MUC5AC. Then in vivo experiment indicated that TRQ could significantly inhibit LPS stimulated MUC5AC over-production as well as the expression of TNF-α, IL-6, IL-8, and IL-17A, in both protein and mRNA levels. CONCLUSIONS: Based on the systems pharmacology method and in vivo experiment, our work provided a general knowledge on the potential active compounds and possible therapeutic targets of TRQ formula in its anti-MH process. This work might suggest directions for further research on TRQ and provide more insight into better understanding the chemical and pharmacological mechanisms of complex herbal prescriptions in a network perspective.

Inula helenium and Grindelia squarrosa as a source of compounds with anti-inflammatory activity in human neutrophils and cultured human respiratory epithelium.

ETHNOPHARMACOLOGICAL RELEVANCE: During the epidemic season, over 90% of acute wheezing disease is associated with bronchial inflammation. Both neutrophil- and eosinophil-mediated inflammation have been involved in the pathophysiology of acute bronchitis, but neutrophil cell recruitment has been shown to be dominant. The ongoing inflammation increases the chemotaxis of neutrophils to inflamed site providing to their overaccumulation. The pharmacological reduction of neutrophil migration can be limited by suppression of major chemo-attractants and cytokines (IL-8, IL-1ß and TNF-α) release and downregulation of adhesive molecules. AIM OF THE STUDY: During a screening of plants traditionally used in respiratory tracts diseases (e.g. cough, rhinitis, bronchitis, throat infection, fever, influenza) in Europe, we have selected roots of Inula helenium and aerial parts of Grindelia squarrosa as a potential source of compounds limiting neutrophil migration. MATERIALS AND METHODS: The effect on IL-8, IL-1ß and TNF-α release by neutrophils and respiratory epithelium cell line (A549) was measured by enzyme-linked immunosorbent assay (ELISA). The surface expression of adhesive molecules was analyzed with flow cytometry, and the neutrophil attachment to the epithelial cells was assessed fluorimetrically. RESULTS: We confirmed the ability of selected extracts and compounds to suppress neutrophil binding to the epithelium surface via downregulation of ß2 integrin. Alantolactone and grindelic acid have shown significant suppression of IL-8, TNF-α and IL-1ß release comparable with budesonide, used as a positive control. CONCLUSIONS: The present study demonstrated that Inula helenium and Grindelia squarrosa, which have been traditionally used in Europe as medicinal plants, are a valuable source of active compounds with anti-inflammatory activity. Our observations justify the traditional use of I. helenium and G. squarrosa for a treatment of inflammation-based diseases in respiratory tract.

Esculetin regulates the phenotype switching of airway smooth muscle cells.

Airway remodeling is one important feature of childhood asthma, which is one of the most common chronic childhood diseases. Phenotype switching of airway smooth muscle cells (ASMCs), defined as a reversible switching between contractile and proliferative phenotypes, plays an important role in the process of airway remodeling. Esculetin has shown antiinflammatory action in animal models of asthma; however, the effects of esculetin on ASMC phenotype switching have not been investigated. In the present study, platelet-derived growth factor (PDGF) was used to induce the phenotype modulation of ASMCs. The results demonstrated that esculetin pretreatment mitigated the PDGF-caused inhibitory effects on expressions of contractile phenotype protein markers, including calponin and SM22α. Esculetin also inhibited PDGF-induced migration and proliferation of ASMCs. Besides, the PDGF-induced expressions of extracellular matrix components, collagen I and fibronectin, were attenuated by esculetin pretreatment. Furthermore, PDGF-caused activation of PI3K/Akt pathway in ASMCs was inhibited by esculetin. These findings suggest that esculetin might exert its inhibitory effect on PDGF-induced ASMC phenotype switching through inhibition of PI3K/Akt pathway.

Ergosterol attenuates cigarette smoke extract-induced COPD by modulating inflammation, oxidative stress and apoptosis in vitro and in vivo.

Cigarette smoke (CS) is the major cause of chronic obstructive pulmonary disease (COPD). CS heightens inflammation, oxidative stress and apoptosis. Ergosterol is the main bioactive ingredient in Cordyceps sinensis (C. sinensis), a traditional medicinal herb for various diseases. The objective of this work was to investigate the effects of ergosterol on anti-inflammatory and antioxidative stress as well as anti-apoptosis in a cigarette smoke extract (CSE)-induced COPD model both in vitro and in vivo Our results demonstrate that CSE induced inflammatory and oxidative stress and apoptosis with the involvement of the Bcl-2 family proteins via the nuclear factor kappa B (NF-κB)/p65 pathway in both 16HBE cells and Balb/c mice. CSE induced epithelial cell death and increased the expression of nitric oxide (NO), interleukin-6 (IL-6), tumor necrosis factor α (TNF-α), malondialdehyde (MAD) and the apoptosis-related proteins cleaved caspase 3/7/9 and cleaved-poly-(ADP)-ribose polymerase (PARP) both in vitro and in vivo, whereas decreased the levels of superoxide dismutase (SOD) and catalase (CAT). Treatment of 16HBE cells and Balb/c mice with ergosterol inhibited CSE-induced inflammatory and oxidative stress and apoptosis by inhibiting the activation of NF-κB/p65. Ergosterol suppressed apoptosis by inhibiting the expression of the apoptosis-related proteins both in vitro and in vivo Moreover, the usage of QNZ (an inhibitor of NF-κB) also partly demonstrated that NF-κB/p65 pathway was involved in the ergosterol protective progress. These results show that ergosterol suppressed COPD inflammatory and oxidative stress and apoptosis through the NF-κB/p65 pathway, suggesting that ergosterol may be partially responsible for the therapeutic effects of cultured C. sinensis on COPD patients.

Molecular Signature of Asthma-Enhanced Sensitivity to CuO Nanoparticle Aerosols from 3D Cell Model.

More than 5% of any population suffers from asthma, and there are indications that these individuals are more sensitive to nanoparticle aerosols than the healthy population. We used an air-liquid interface model of inhalation exposure to investigate global transcriptomic responses in reconstituted three-dimensional airway epithelia of healthy and asthmatic subjects exposed to pristine (nCuO) and carboxylated (nCuOCOOH) copper oxide nanoparticle aerosols. A dose-dependent increase in cytotoxicity (highest in asthmatic donor cells) and pro-inflammatory signaling within 24 h confirmed the reliability and sensitivity of the system to detect acute inhalation toxicity. Gene expression changes between nanoparticle-exposed versus air-exposed cells were investigated. Hierarchical clustering based on the expression profiles of all differentially expressed genes (DEGs), cell-death-associated DEGs (567 genes), or a subset of 48 highly overlapping DEGs categorized all samples according to "exposure severity", wherein nanoparticle surface chemistry and asthma are incorporated into the dose-response axis. For example, asthmatics exposed to low and medium dose nCuO clustered with healthy donor cells exposed to medium and high dose nCuO, respectively. Of note, a set of genes with high relevance to mucociliary clearance were observed to distinctly differentiate asthmatic and healthy donor cells. These genes also responded differently to nCuO and nCuOCOOH nanoparticles. Additionally, because response to transition-metal nanoparticles was a highly enriched Gene Ontology term (FDR 8 × 10-13) from the subset of 48 highly overlapping DEGs, these genes may represent biomarkers to a potentially large variety of metal/metal oxide nanoparticles.

Antitussive, expectorant, and anti-inflammatory effects of Adenophorae Radix powder in ICR mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Adenophora triphylla var. japonica is frequently used as an oriental medicinal plant in Korea, China, and Japan for its anti-inflammatory, antitussive, and hepatoprotective effects. AIM OF THE STUDY: In the present study, the antitussive, expectorant, and anti-inflammatory effects of AR powder were investigated using animal models to evaluate their potential to treat respiratory disorders. MATERIALS AND METHODS: AR powder was administered orally to mice once daily for 11 days, at dose levels of 400, 200, and 100 mg/kg. Theobromine (TB), ambroxol (AM) and dexamethasone (DEXA) were used as standard drugs for antitussive effects, expectorant effects and anti-inflammatory effects, respectively. Evaluations of antitussive effects were based on changes in body weight, the number of cough responses and the histopathology of the lung and trachea. Expectorant effects were based on changes in the body weight, macroscopic observations of body surface redness, the mucous secretion of the trachea and histopathology of lung (secondary bronchus). Anti-inflammatory effects were based on changes in the body weight, macroscopic observations involving redness and edema of the treated ear, absolute and relative ear weights and histopathology of the treated ears. RESULTS: Allergic acute inflammation and coughing induced by exposure to NH4OH and symptoms of xylene-induced contact dermatitis were significantly inhibited by treatment with AR powder in a dose-dependent manner. Histological analyses revealed that AR powder decreased the OD values in trachea lavage fluid, reduced body surface redness, thicknesses of intrapulmonary secondary bronchus mucosa, and the number of PAS-positive mucous producing cells. Overall, AR powder administered at 200 mg/kg displayed superior antitussive and expectorant effects as compared to TB (50 mg/kg), and AM (250 mg/kg). At the highest concentration (400 mg/kg) AR powder displayed only moderately improved anti-inflammatory activities as compared to DEXA (1 mg/kg). CONCLUSION: The results obtained in this study suggest that AR powder exerts dose-dependent, favorable antitussive, expectorant, and anti-inflammatory activities achieved through modulation of the activity of mast cells and respiratory mucous production. Therefore, AR powder may serve as a therapeutic agent in various respiratory disorders, especially those that occur as a result of environmental toxicants.

Pretreatment with a grape seed proanthocyanidin extract downregulates proinflammatory cytokine expression in airway epithelial cells infected with respiratory syncytial virus.

Respiratory syncytial virus (RSV) infections are associated with significant morbidity and mortality. Inflammation is mediated by cytokine secretion from RSV­infected airway epithelial cells. Grape seed proanthocyanidin extract (GSPE) exhibits potent antioxidant capacity, as well as anti­bacterial, anti­viral, anti­carcinogenic, anti­inflammatory and anti­allergic actions. However, few studies have explored the anti­inflammatory effects of GSPE on airway epithelial cells infected with RSV. Airway epithelial A549 cells were pretreated with GSPE and its effects on cytokine production during RSV infection were investigated. A549 cells were infected with RSV, with or without GSPE pretreatment, and cultured for 24, 48 and 72 h. The expression of interleukin (IL)­1ß, IL­6 and IL­8, were measured by reverse transcription­quantitative polymerase chain reaction, ELISA and western blotting. RSV infection induced significant increases in proinflammatory cytokine expression. However, GSPE pretreatment decreased the mRNA and protein expression levels of IL­1ß, IL­6 and IL­8. GSPE regulated the immune response by reducing the RSV­induced transcription of proinflammatory cytokines in airway epithelial cells, suggesting that GSPE helps to prevent RSV­induced airway disease.

Anti-inflammatory and anti-remodeling effects of myrtenol in the lungs of asthmatic rats: Histopathological and biochemical findings.

INTRODUCTION: Asthma is a chronic inflammatory disease of the airways. In this study, we evaluated the anti-inflammatory effects of myrtenol on the inflammatory indices in the pulmonary parenchyma and airways and on the inflammatory and oxidative indices of the bronchoalveolar lavage fluid (BALF) of asthmatic rats. METHODS: The allergic asthma was induced by sensitization (two weeks) followed by the inhalation of ovalbumin (four weeks). Animals were divided into two main groups: (1) Histopathology, and (2) measurement of inflammatory and oxidative biomarkers in the BALF. Each main group was subdivided into four subgroups: Control, Asthma, Asthma+Dexamethasone and Asthma+Myrtenol. (-)-Myrtenol (50mg/kg) or Dexamethasone (2.5mg/kg) was administered intraperitoneally once a day for one week, at the end of the inhalation period. On day 50, lung histopathologic parameters and inflammatory indices in BALF including INF-γ, IL-10, IL-1ß, and TNF-α and oxidative stress biomarkers (MDA, SOD, and GPX) were measured. RESULT: In the Asthma group, leukocyte infiltration, the thickness of smooth muscle and epithelium of airways wall and the number of goblet cells increased. Myrtenol reduced all of the above-mentioned indices except the epithelium thickness. It also inhibited the increase in BALF IL-1ß, TNF-α and MDA and increased the levels of INF-γ, IL-10 and SOD. CONCLUSION: Our results suggest that myrtenol reduced damage caused by experimental asthma by reducing the inflammatory indices, normalizing the level of interleukins and balancing oxidative stress in the lungs. It also prevented airway remodeling. Myrtenol may be suggested as a potent herbal medicine for the treatment of allergic asthma.

Pingchuanning decoction attenuates airway inflammation by suppressing autophagy via phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin signaling pathway in rat models of asthma.

BACKGROUND: Pingchuanning decoction is a well-known traditional Chinese medicine for the treatment of airway inflammatory diseases, including asthma. However, the potential mechanism by which Pingchuanning decoction contributes to the amelioration of airway inflammation remains unknown. METHODS: A rat model of asthma was well established by inducing ovalbumin. Lipopolysaccharide-stimulated rat tracheal epithelial (RTE) cells were used as cellular model. Lung histopathology and goblet cell hyperplasia were assessed by hematoxylin-eosin (HE) and periodic acid Schiff staining, respectively. Total inflammatory cells count and RTE cell apoptosis were analyzed by flow cytometry. The autophagic activities were evaluated by immunohistochemical and immunofluorescence analysis and Western blot analysis of autophagy-related proteins. We also detected the effects of Pingchuanning decoction on phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) and high-mobility group box 1 (HMGB1)-mediated toll-like receptor 4 (TLR4)/NF-κB pathways-related proteins and inflammatory cytokines using the Western blot analysis and enzyme-linked immunosorbent assay. RESULTS: Pingchuanning decoction effectively attenuated pulmonary pathology and autophagy. Treatment with Pingchuanning decoction activated PI3K/Akt/mTOR pathway and inhibited HMGB1/TLR4/NF-κB pathway, which could be overturned by LY294002, a PI3K antagonist, or rapamycin (Rapa), an autophagy inducer. CONCLUSION: Pingchuanning decoction exerted a therapeutic effect on asthma by inhibiting autophagy via PI3K/Akt /mTOR signaling pathway.