Anti-pulmonary Fibrosis Effect Research Articles

Evaluating the Therapeutic Mechanisms of Selected Active Compounds in Houttuynia cordata Thunb. in Pulmonary Fibrosis via Network Pharmacology Analysis.

Pulmonary fibrosis, a common outcome of pulmonary interstitial disease of various different etiologies, is one of the most important causes of respiratory failure. Houttuynia cordata Thunb. (family: Saururaceae) (H. cordata), as has been reported, is a Chinese herbal medicine commonly used to treat upper respiratory tract infection and bronchitis. Our previous study has proven that sodium houttuyfonate (an additional compound from sodium bisulfite and houttuynin) had beneficial effects in the prevention of pulmonary fibrosis (PF) induced by bleomycin (BLM) in mice. In the present study, network pharmacology was used to investigate the efficiency and potential mechanisms of H. cordata in PF treatment. Upon manual collection from the literature and databases such as TCMSP and TCM-ID, 10 known representative ingredients of H. cordata species were screened. Then, the prediction of the potential active ingredients, action targets, and signaling pathways were conducted through the Gene Ontology (GO), protein–protein interaction (PPI),and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. The results of network pharmacology prediction suggested that H. cordata may act through multiple signaling pathways to alleviate PF, including the phosphatidylinositol 3-kinase-protein kinase B (PI3K/AKT) pathways, mitogen-activated protein kinase (MAPK) pathways, the tumor necrosis factor (TNF) pathways, and interleukin-17 (IL-17) signaling pathways. Molecular docking experiments showed that the chemical constituents of H. cordata had good affinity with TNF, MAPK1, and AKT1, and using lipopolysaccharide (LPS)-induced A549 cells, a model was established to verify the anti-pulmonary fibrosis effects and related mechanisms of H. cordata–relevant constituents. Finally, these evidences collectively suggest H. cordata may alleviate PF progression via PI3K/Akt, MAPK, and TNF signaling pathways and provide novel insights to verify the mechanism of H. cordata in the treatment of PF.

Mangostanin Derivatives as Novel and Orally Active Phosphodiesterase 4 Inhibitors for the Treatment of Idiopathic Pulmonary Fibrosis with Improved Safety.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease, and its incidence rate is rapidly rising. However, effective therapies for the treatment of IPF are still lacking. Phosphodiesterase 4 (PDE4) inhibitors were reported to be potential anti-fibrotic agents, but their clinical use was hampered by side effects like emesis and nausea. Herein, structure-based hit-to-lead optimizations of natural mangostanin resulted in the novel and orally active PDE4 inhibitor 18a with potent inhibitory affinity (IC50 = 4.2 nM), favorable physico-chemical properties, and a different binding pattern from roflumilast. Emetic activity tests on dogs demonstrated that 18a cannot cause emesis even at an oral dose of 10 mg/kg, whereas rolipram had severe emetic effects at an oral dose of 1 mg/kg. Finally, the oral administration of 18a (10 mg/kg) exhibited comparable anti-pulmonary fibrosis effects with pirfenidone (150 mg/kg) in a bleomycin-induced IPF rat model, indicating its potential as a novel anti-IPF agent with improved safety.

Mechanistic Investigation of Xuebijing for Treatment of Paraquat-Induced Pulmonary Fibrosis by Metabolomics and Network Pharmacology.

After paraquat (PQ) poisoning, it is difficult to accurately diagnose patients’ condition by only measuring their blood PQ concentration. Therefore, it is important to establish an accurate method to assist in the diagnosis of PQ poisoning, especially in the early stages. In this study, a gas chromatography–mass spectrometry (GC–MS) metabonomics strategy was established to obtain metabolite information. A random forest algorithm was used to search for potential biomarkers of PQ poisoning, and data mining and network pharmacological analysis were used to evaluate the active components, drug–disease targets, and key pathways of Xuebijing (XBJ) injection in the treatment of PQ-induced pulmonary fibrosis. Targets from the network pharmacology analysis and metabolites from plasma metabolomics were jointly analyzed to select crucial metabolic pathways. Finally, molecular docking technology and in vitro experiments were used to verify the pathway targets to further reveal the potential mechanisms underlying the antipulmonary fibrosis effect of XBJ. Metabonomics studies showed that l-valine, glycine, citric acid, d-mannose, d-galactose, maltose, l-tryptophan, and arachidonic acid contributed more to the differentiation of different groups than other metabolites. Compared with the control group, the PQ poisoning group had higher levels of l-valine, glycine, citric acid, l-tryptophan, and arachidonic acid, and lower levels of d-mannose, d-galactose, and maltose. After treatment with XBJ injection, the relative levels of these metabolites were reversed. The network pharmacological analysis screened a total of 180 targets, mainly involving multiple signaling pathways and metabolic pathways, which jointly played an antipulmonary fibrosis effect. Based on the combined analysis of 180 targets and 8 different metabolites, arachidonic acid metabolism was selected as the key metabolic pathway. Molecular docking analysis showed that the XBJ compound had strong binding activity with the target protein. Western blot results showed that XBJ injection could reduce the inflammatory response by downregulating the expressions of p-p65, p-IKBα, and p-IKKβ, thus inhibiting the development of PQ-induced pulmonary fibrosis. In summary, the combined results from metabolomics and network pharmacology studies showed that Xuebijing has the characteristics of multitarget, multichannel, and multicomponent action in the treatment of pulmonary fibrosis caused by PQ.

PPARγ mediates the anti-pulmonary fibrosis effect of icaritin

BackgroundPulmonary fibrosis is a fatal lung disease with limited treatment options. Icaritin is the active ingredient derived from the traditional Chinese medical plant Epimedium and possesses many biomedical activities. This study aimed to investigate the effects and molecular mechanisms of icaritin on bleomycin-induced pulmonary fibrosis in mice. MethodsTo assess its preventative effects, bleomycin treated mice received 0, 0.04, 0.2, and 1 mg/kg of icaritin from day 1 onwards. To assess its therapeutic effects, bleomycin treated mice received 0 and 1 mg/kg of icaritin from day 15 onwards. Mice were sacrificed on day 21 and lung tissues were collected, stained with HE, Masson and immunohistochemistry. Q-PCR was used to measure Collagen I and Collagen III expression, western blotting was used to quantify α-SMA, Collagen I expression. Hydroxyproline content was measured using a biochemical method. NIH3T3 and HLF-1 cells were treated with TGF-β1with or without icaritin, and α-SMA, Collagen I were tested. PPARγ antagonist GW9662 and PPARγ-targeted siRNA were used to investigate the mechanism of icaritin in inhibiting myofibroblast differentiation. ResultsBoth preventative and therapeutic administration of icaritin improved the histopathological changes, decreased Collagen and α-SMA, lowered hydroxyproline content in bleomycin-treated lung tissues. Icaritin decreased α-SMA and Collagen I expression in TGF-β1-stimulated NIH3T3 and HLF-1 cells. However, its effect in reducing α-SMA and Collagen I expression was suppressed when expression or activity of PPARγ was inhibited. ConclusionsIcaritin has therapeutic potential against pulmonary fibrosis via the inhibition of myofibroblast differentiation, which may be mediated by PPARγ.

Mechanism of Dandelion Sterol in Treating Pulmonary Fibrosis Through Transforming Growth Factor-β Signaling Pathway

Background: The paper aimed to elucidate the molecular mechanism of Dandelion sterol in the treatment of pulmonary fibrosis, to study its effect on EMT of lung epithelial cells, and to find its target and downstream signaling pathways. Material and methods: The effects of Dandelion sterol on parathyroid (PQ)-induced EMT in lung epithelial cells were studied by immunofluorescence method. Immunohistochemistry and western-blot methods were used to verify that Dandelion sterol inhibited TGF-β1-induced EMT at the cellular level in animals, demonstrating that Dandelion sterol targets TGF-β1 to exert an anti-pulmonary fibrosis effect. Results: Dandelion sterol significantly inhibited PQ-induced migration and invasion of lung epithelial cells, and also inhibited the induced EMT. Dandelion sterol had a proper binding activity with the lung fibrosis-inducing factor TGF-β1. Dandelion sterol inhibited the TGF-β1-induced EMT process, and acted to treat pulmonary fibrosis by inhibiting the TGF-β1/Smad3 signaling pathway. Conclusion: Dandelion sterol can inhibit the pulmonary fibrosis by inhibiting the EMT process of lung epithelial cells through targeting the TGF- β1/Smad signaling pathway.

Effect of Fluorofenidone Against Paraquat-Induced Pulmonary Fibrosis Based on Metabolomics and Network Pharmacology.

BackgroundFluorofenidone (AKF-PD) is an anti-fibrotic small-molecule compound. Its mechanism of action on paraquat (PQ)-induced pulmonary fibrosis is still unclear.Material/MethodsForty-eight SD rats were divided into 4 groups: control group, PQ group, PQ+AKF-PD group, and AKF-PD group. The pathological changes of lung tissues were observed by Masson and HE staining. The UPLC-QTOF-MS analysis was performed to detect the differences in metabolites among groups, then the possible mechanisms of the anti-pulmonary fibrosis effects of fluorofenidone were further revealed by network pharmacology analysis. Biological methods were used to verify the results of the network pharmacology analysis.ResultsThe results showed that fluorofenidone treatment significantly alleviated paraquat-induced pulmonary fibrosis. Metabolomics analysis showed that 18 metabolites were disordered in the serum of paraquat-poisoned rats, of which 13 were restored following fluorofenidone treatment. Network pharmacology analysis showed that the drug screened a total of 12 targets and mainly involved multiple signaling pathways and metabolic pathways to jointly exert anti-pulmonary fibrosis effects. Autophagy is the main pathway of fluorofenidone in treatment pulmonary fibrosis. The western blot results showed that fluorofenidone upregulated the expression of LC3-II/I and E-cadherin, and downregulated the expression of p62, α-SMA, and TGF-β1, which validated that fluorofenidone could inhibit the development of paraquat-induced pulmonary fibrosis by increasing autophagy.ConclusionsIn conclusion, metabolomics combined with network pharmacology research strategy revealed that fluorofenidone has a multi-target and multi-path mechanism of action in the treatment of pulmonary fibrosis.

Chrysin inhibited epithelial-mesenchymal transition of type Ⅱ alveolar epithelial cell by regulating NF-κB/Twist 1 signaling pathway

This study aimed to assess whether chrysin(ChR) can inhibit epithelial-mesenchymal transition(EMT) of type Ⅱ alveolar epithelial cell and produce anti-pulmonary fibrosis effect by regulating the NF-κB/Twist 1 signaling pathway. Sixty rats were randomly divided into the control group, the bleomycin(BLC) group, BLC+ChR(50 mg·kg~(-1)) group and BLC+ChR(100 mg·kg~(-1)) group, with 15 rats in each group. The pulmonary fibrosis model was induced by intratracheal injection of BLC(7 500 U·kg~(-1)). Rats were orally administered with different doses of ChR after BLC injection for 28 days. The cells were divided into control group, TGF-β1 group(5 ng·mL~(-1)), and TGF-β1+ChR(1, 10, 100 μmol·L~(-1)) groups. The type Ⅱ alveolar epithelial cells were treated with TGF-β1 for 24 h, and then treated with TGF-β1 for 48 h in the presence or absence of different doses of ChR(1, 10 and 100 μmol·L~(-1)). The morphological changes and collagen deposition in lung tissues were analyzed by HE staining, Masson staining and immunohistochemistry. The mRNA and protein expression levels of collagen Ⅰ, E-cadherin, zonula occludens-1(ZO-1), vimentin, alpha smooth muscle actin(α-SMA), inhibitor of nuclear factor kappa B alpha(IκBα), nuclear factor-kappa B p65(NF-κB p65), phospho-NF-κB p65(p-p65) and Twist 1 in lung tissues and cells were detected by qPCR and Western blot, respectively. The animal experiment results showed that as compared with the BLC group, after administration of ChR for 28 days, bleomycin-induced pulmonary fibrosis in rats was significantly relieved, collagen Ⅰ expression in lung tissues was significantly reduced(P<0.05 or P<0.01), and EMT of alveolar epithelial cells was obviously inhibited [the expression levels of E-cadherin and ZO-1 were increased and the expression levels of vimentin and α-SMA were decreased(P<0.05 or P<0.01)], concomitantly with significantly reduced IκBα and p65 phosphorylation level in cytoplasm and decreased NF-κB p65 and Twist 1 expression in nucleus(P<0.05 or P<0.01). The cell experiment results showed that different doses of ChR(1, 10 and 100 μmol·L~(-1)) significantly reduced TGF-β1-induced collagen Ⅰ expression(P<0.05 or P<0.01), significantly inhibited EMT of type Ⅱ alveolar epithelial cells[the expression levels of E-cadherin and ZO-1 were increased and the expression levels of vimentin and α-SMA were decreased(P<0.05 or P<0.01)], and inhibited IκBα and p65 phosphorylation in cytoplasm and down-regulated NF-κB p65 and Twist 1 expression in nucleus induced by TGF-β1(P<0.05 or P<0.01). The results suggest that ChR can reverse EMT of type Ⅱ alveolar epithelial cell and alleviate pulmonary fibrosis in rats, and its mechanism may be associated with reducing IκBα phosphorylation and inhibiting NF-κB p65 phosphorylation and nuclear transfer, thus down-regulating Twist 1 expression.

N-BuOH extract of Bletilla striata exerts chemopreventive effects on lung against SiO2 nanoparticles through activation of Nrf2 pathway

BackgroundSiO2 nanoparticles (nm SiO2) are ubiquitous in daily life and are acknowledged to be detrimental to human health. Bletilla striata is a traditional medicine used for generations in China and its polysaccharide has the anti-pulmonary fibrosis effect. PurposeTo investigate the lung protective effect of the small molecules (n-BuOH extract) of B. striata and clarify the underlying mechanism. Study design and methodsC57BL/6 mice were subjected to intratracheal instillation with nm SiO2 nanoparticle suspension (7 mg/kg) to construct the in vivo model of nm SiO2-induced lung injury. The chemical profile of the n-BuOH extract of B. striata was investigated by HPLC analysis using authentic samples isolated from B. striata. Gymnoside II with the most potent chemoprotective capacity in the n-BuOH extract was used to clarify the potential bio-active molecular basis of the n-BuOH extract using in vitro experiments. The cytotoxicity, apoptosis, oxidative stress, and the Nrf2 signaling pathway were examined in SiO2-induced A549 cells. ML385 was adopted to down-regulate the Nrf2 expression. ResultsThe n-BuOH extract of B. striata (40 mg/kg) could alleviate the SiO2-induced lung injury by increasing Nrf2 expression and thereby suppressing Bax/Bcl-2 pathway in the nm SiO2-induced mice model. The chemical profile study showed that militarine, gymnoside II, and 4-allyl-2, 6-dimethoxyphenol glucoside were the main constituents of n-BuOH extract. Studies on gymnoside II revealed that it could partially restore the SiO2-induced decline in cell viability while did not affect the growth of normal A549 cells within the concentration range of 1-50 μM, suggesting a protective effect against nm SiO2 in lung A549 cells. The hoechst 33258 staining, flow cytometry, and western blot experiments demonstrated that gymnoside II (25 μM) could partially reverse the SiO2-induced cell apoptosis and ROS production by enhancing Nrf2, HO-1, and γ-GCSc expressions and Nrf2 silencing by ML385 abrogated the effects of gymnoside II (25 μM) on apoptosis and ROS production in A549 cells. ConclusionThe present study suggests that in addition to the polysaccharide, small molecules (n-BuOH extract) of B. striata can also elicit a protective effect on lung injuries through the Nrf2-dependent mechanism and gymnoside II is one of the main bio-active constituents contributing to the n-BuOH extract-elicited lung protective effect against nm SiO2.

Betulinic acid attenuated bleomycin-induced pulmonary fibrosis by effectively intervening Wnt/β-catenin signaling

BackgroundIdiopathic pulmonary fibrosis (IPF) is a fatal and progressive fibrotic lung disease lacking a validated and effective therapy. Aberrant activation of the Wnt/β-catenin signaling cascade plays the key role in the pathogenesis of IPF. Betulinic acid is a natural pentacyclic triterpenoid molecule that has excellent antitumor and antiviral activities. HypothesisWe hypothesized that BA has an anti-pulmonary fibrosis effect mediated by the suppression of the Wnt/β-catenin pathway.Study design Pulmonary fibrosis markers were detected in vitro and in vivo to confirm the antifibrotic effect of BA. The Wnt/β-catenin pathway-related proteins were overexpressed to determine the effect of BA on Wnt signaling. Methods and resultsBA dose-dependently inhibited Wnt3a-induced fibroblast activation in vitro. Moreover, BA decreased Wnt3a- and LiCl-induced transcriptional activity, as assessed by the TOPFlash assay in fibroblasts, and repressed the expression of the Wnt target genes cyclin D1, axin 2, and S100A4. Further investigation indicated that BA restrained the nuclear accumulation of β-catenin, mainly by increasing the phospho-β-catenin ratio (S33/S37/T41 and S45), inhibited the phosphorylation of DVL2 and LRP, and decreased the levels of Wnt3a and LRP6. In agreement with the results of the in vitro assays, the in vivo experiments indicated that BA significantly decreased bleomycin-induced pulmonary fibrosis in mice and suppressed myofibroblast activation by inhibiting Wnt/β-catenin signaling. ConclusionBA may directly interfere with the Wnt/β-catenin pathway to subsequently repress myofibroblast activation and pulmonary fibrosis.

Pharmacological effects of indole alkaloids from Alstonia scholaris (L.) R. Br. on pulmonary fibrosis in vivo

Ethnopharmacological relevanceAlstonia scholaris (L.) R. Br. (Apocynaceae) is a Dai folk medicine for the treatment of lung diseases in China.Aim of the study: The present study investigated the anti-pulmonary fibrosis effects of total alkaloids (TA) and the potential active ingredients and its possible mechanism. Materials and methodsAfter intratracheal instillation of bleomycin (BLM, 5 mg/kg), mice were divided into ten groups, and orally treated with the corresponding samples once daily for 28 days. The effect of indole alkaloids was determined through analysis of cytokines, as well as histopathological examinations and gene expressions. ResultsSevere lung fibrosis was observed in the BLM-treated mice on day 28. However, the administration of TA significantly ameliorated the pathological changes in the lungs, decreased the content of Krebs von den Lungen-6, lactate dehydrogenase, transforming growth factor-β (TGF-β), hydroxyproline, type I collagen, and malonaldehyde, and enhanced the activity of superoxide dismutase in the serum and lung tissues. In addition, the enhanced TGF-β and matrix metalloproteinase-1 (MMP-1) expressions in BLM-induced mice were obviously weakened by indole alkaloids, as well as the ratio of matrix metalloproteinase-1 to tissue inhibitor of metalloproteinase-1 was decreased. Moreover, picrinine and scholaricine yielded markedly better values in the aforementioned indices than those in other samples, indicating that they may be the active ingredients of alkaloids. ConclusionsTA exerted protective effects against BLM-induced pulmonary fibrosis by reducing collagen deposition through TGF-β/MMP-1 pathway.

A dry powder inhalable formulation of salvianolic acids for the treatment of pulmonary fibrosis: safety, lung deposition, and pharmacokinetic study.

Salvianolic acids (SAL), the main bioactive component extracted from Salvia miltiorrhiza, is a natural product with a reported anti-pulmonary fibrosis (PF) effect. SAL is commonly administrated orally; however, it has a low oral bioavailability (less than 5%). The objective of this work was to develop a new dry powder inhalable formulation intended to facilitate the access of SAL to the target place. We prepared the new SAL powder formulation containing L-arginine and 2% of lecithin using the ball milling technique. L-arginine was used to regulate the strong acidity of the SAL solution, and lecithin was added to disperse the powder and improve the flowability. The resulting powder had a content in salvianolic acid B (SALB, the main active principle of SAL) of 66.67%, a particle size of less than 5μm and a good flowability. In vivo fluorescence imaging showed that the powder could be successfully aerosolized and delivered to the lung. The acute lung irritation study proved that the presence of L-arginine improved the biocompatibility of the powder. Finally, according to the pharmacokinetic study, the new SAL powder formulation was found to significantly increase drug concentration in the lung and the bioavailability. In conclusion, the new dry powder inhalable formulation of SAL developed in this study could be a strategy to enhance the performance of SAL at the lung level. Graphical abstract.

Discovery of Novel Selective and Orally Bioavailable Phosphodiesterase-1 Inhibitors for the Efficient Treatment of Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and devastating lung disease lacking effective therapy. To identify whether phosphodiesterase-1 (PDE1) inhibition could act as a novel target for the treatment of IPF, hit-to-lead structural optimizations were performed on the PDE9/PDE1 dual inhibitor (R)-C33, leading to compound 3m with an IC50 of 2.9 nM against PDE1C, excellent selectivity across PDE subfamilies, reasonable drug-like properties, and remarkable pharmacodynamic effects as an anti-IPF agent. Oral administration of compound 3m (10 mg/kg) exerted more significant anti-pulmonary fibrosis effects than pirfenidone (150 mg/kg) in a bleomycin-induced IPF rat model and prevented transforming growth factor-β-induced fibroblast-to-myofibroblast conversion in vitro, indicating that PDE1 inhibition could serve as a novel target for the efficient treatment of IPF.

Use of Chou's 5-Steps Rule to Reveal Active Compound and Mechanism of Shuangshen Pingfei San on Idiopathic Pulmonary Fibrosis.

Shuangshen Pingfei San (SPS) is the derivative from the classic formula Renshen Pingfei San in treating idiopathic pulmonary fibrosis (IPF). In this study, Chou's 5-steps rule was performed to explore the potential active compound and mechanism of SPS on IPF. Compound-target network, target- pathway network, herb-target network and the core gene target interaction network were established and analyzed. A total of 296 compounds and 69 candidate therapeutic targets of SPS in treating IPF were obtained. Network analysis revealed that the main active compounds were flavonoids (such as apigenin, quercetin, naringenin, luteolin), other clusters (such as ginsenoside Rh2, diosgenin, tanshinone IIa), which might also play significant roles. SPS regulated multiple IPF relative genes, which affect fibrosis (PTGS2, KDR, FGFR1, TGFB, VEGFA, MMP2/9) and inflammation (PPARG, TNF, IL13, IL4, IL1B, etc.). In conclusion, anti-pulmonary fibrosis effect of SPS might be related to the regulation of inflammation and pro-fibrotic signaling pathways. These findings revealed that the potential active compounds and mechanisms of SPS on IPF were a benefit to further study.

β-carboline alkaloids attenuate bleomycin induced pulmonary fibrosis in mice through inhibiting NF-kb/p65 phosphorylation and epithelial-mesenchymal transition

Ethnopharmacological relevanceThe plant Arenaria kansuensis is used in traditional medicine to treat lung inflammation for a long time. However, the anti-pulmonary fibrosis effect and its corresponding bioactive constituents of this plant have not been studied extensively. Aim of the studyThe purpose of this study was to investigate the anti-pulmonary fibrosis effect and its corresponding bioactive constituents of A. kansuensis and its possible mechanism. Materials and methodsIn vivo experiment, the anti-pulmonary fibrosis effects of the fraction (Part1) enriched from ethyl acetate extracts of the whole plant A. kansuensis were evaluated through bleomycin (BLM)-induced pulmonary fibrosis mice (five groups, n = 10) daily at doses of 50, 100 and 150 mg/kg for 15 days. In vitro experiment, the anti-inflammation and reversed epithelial-mesenchymal transition (EMT) effect of 12 β-carboline alkaloids isolated from Part1 were evaluated through lipopolysaccharide (LPS)-induced RAW264.7 inflammatory cell model and TGF-β1 induced A549 cell model. ResultsIn this study, a fraction named Part1 extracted from Arenaria kansuensis presented strong anti-pulmonary fibrosis effect at the dose of 150 mg/kg. Vivo experiments showed that the survival rate and body weight of mice significantly increased after Part1 treatment. Part1 could significantly inhibit the initial of inflammation, deposition of collagen and expression of TGF-β1 and α-SMA, moreover, the expression of E-cadherin was significantly elevated after administration of Part1. All the cure effects of Part1 were in dose dependent manner. A total of 12 β-carboline alkaloids were identified in Part1 and they all showed suppressive effect on inflammatory cytokines including MCP-1, TNF-α, IL-6 and IL-1β through inhibition of NF-kb/p65 phosphorylation, and that epithelial-mesenchymal transition (EMT) process was reversed by different compounds in different levels. The expression of indicators of EMT including α-SMA, vimentin and E-cadherin was significantly improved after given different β-carboline alkaloids. ConclusionsThis study showed that antifibrogenic effect of β-carboline alkaloids was due to inhibiting the initial of inflammation through NF-kb/p65 pathway and reversing the process of EMT.

Mangiferin attenuates bleomycin-induced pulmonary fibrosis in mice through inhibiting TLR4/p65 and TGF-β1/Smad2/3 pathway.

Investigating the antipulmonary fibrosis effect of mangiferin from Mangifera indica and the possible molecular mechanism. In vivo, bleomycin (BLM)-induced pulmonary fibrosis experimental model was used for evaluating antipulmonary fibrosis effect of mangiferin. Histopathologic examination and collagen deposition were investigated by HE and Masson staining as well as detecting the content of hydroxyproline. The expression of transforming growth factor-β1 (TGF-β1), α-smooth muscle actin (α-SMA), TLR4 and p-P65 in lung tissue was analysed through immunofluorescence. Leucocytes and inflammatory cytokines including IL-1β, IL-6, TNF-α and MCP-1 in bronchoalveolar lavage fluid were detected by cell counting and enzyme-linked immunosorbent assay. In vitro, TGF-β1-induced A549 epithelial-mesenchymal transition (EMT) cell model was used for investigating the possible molecular mechanism. Reactive oxygen species (ROS) generation was detected by DCFH-DA assay. Expression of all proteins was examined by Western blot. Oral administration of mangiferin could attenuate the severity of BLM-induced pulmonary fibrosis through increasing the survival rate, improving histopathological lesion and body weight loss as well as decreasing pulmonary index visibly. Pulmonary hydroxyproline content, TGF-β1, and α-SMA levels were reduced significantly. The molecular mechanism of mangiferin for inhibiting pulmonary fibrosis is that it could obviously inhibit the occurrence of inflammation and the secretion of inflammatory cytokine through inhibiting activation of TLR4 and phosphorylation of p65. Meanwhile, EMT process was suppressed obviously by mangiferin through blocking the phosphorylation of Smad2/3 and reducing MMP-9 expression. Besides, mangiferin could significantly inhibit the process of oxidant stress through downregulating the intracellular ROS generation. Mangiferin attenuates BLM-induced pulmonary fibrosis in mice through inhibiting TLR4/p65 and TGF-β1/Smad2/3 pathway.

Paeoniflorin inhibits IL-1β-induced MMP secretion via the NF-κB pathway in chondrocytes.

Paeoniflorin serves important cellular roles, exerting anti-cancer, anti-inflammatory and anti-pulmonary fibrosis effects and possesses immune-modulatory properties. However, the exact role of paeoniflorin in the pathogenesis of osteoarthritis (OA) remains unclear. The aim of the present study was to investigate the effects of paeoniflorin on articular surfaces in vitro. Rat chondrocytes were cultured in vitro and an MTT assay was performed to assess chondrocyte survival. Following treatment with interleukin (IL)-1β and paeoniflorin, the production of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases-1 (TIMP-1) was examined using reverse transcription-quantitative polymerase chain reaction and western blotting. The interleukin (IL)-1β-induced nuclear factor (NF)-κB pathway activation was also investigated. The results demonstrated that paeoniflorin was able to downregulate the expression of MMP and increase the expression of TIMP-1ntmRNA and protein in IL-1β-induced rat chondrocytes. Furthermore, treating chondrocytes with paeoniflorin blocked the activation of NF-κB. These results suggest that paeoniflorin may serve am anti-catabolic role in the progression of OA and may be an effective preventative treatment for OA.

Transplantation of adipose-derived mesenchymal stem cells attenuates pulmonary fibrosis of silicosis via anti-inflammatory and anti-apoptosis effects in rats

BackgroundSilicosis has been topping the list of high-incidence occupational diseases in developing countries and cannot be completely cured. Recent advances in stem cell research have made possible the treatment of various diseases including lung fibrosis. The application of stem cell therapy in occupational diseases, in particular the use of adipose-derived mesenchymal stem cells (AD-MSCs) in treatment of silicosis, has not yet been reported. The aim of the study is to explore the intervening effect of silica-induced lung fibrosis in rats.MethodsIn this study, we investigated the anti-pulmonary fibrosis effects of the transplantation of AD-MSCs in rats in which lung fibrosis was induced by oral tracheal intubation with silica suspension. Twenty rats were divided into four groups: control group (n = 5), exposure group (n = 5), vehicle group (n = 5) and treatment group (n = 5). AD-MSCs were given to rats after exposure to silica for 24 h. Twenty-eight days after AD-MSC transplantation, we examined the organ coefficient, inflammatory cytokines, apoptosis, pathological and fibrotic changes in lung tissue.ResultsResults showed that exposure to silica for 28 days induced an increase of the lung coefficient with significant pulmonary fibrosis. Treatment with AD-MSC transplantation led to a remissive effect on pulmonary fibrosis. We found that after AD-MSC transplantation the inflammatory response decreased and Caspase-3 protein expression significantly decreased with a significant increase of the Bcl-2/Bax ratio.ConclusionsAnti-inflammatory and anti-apoptosis of AD-MSCs may play important roles in their anti-pulmonary fibrosis effect. Our data suggest that transplantation of AD-MSCs holds promise for potential interference in the formation of silicosis through regulating inflammatory and apoptotic processes.

Madecassoside ameliorates bleomycin-induced pulmonary fibrosis in mice through promoting the generation of hepatocyte growth factor via PPAR-γ in colon.

Madecassoside has potent anti-pulmonary fibrosis (PF) effects when administered p.o., despite having extremely low oral bioavailability. Herein, we explored the mechanism of this anti-PF effect with regard to gut hormones. A PF model was established in mice by intratracheal instillation of bleomycin. Haematoxylin and eosin stain and Masson's trichrome stain were used to assess histological changes in the lung. Quantitative-PCR and Western blot detected mRNA and protein levels, respectively, and cytokines were measured by ELISA. Small interfering RNA was used for gene-silencing. EMSA was applied to detect DNA-binding activity. Administration of madecassoside, p.o., but not its main metabolite madecassic acid, exhibited a direct anti-PF effect in mice. However, i.p. madecassoside had no anti-PF effect. Madecassoside increased the expression of hepatocyte growth factor (HGF) in colon tissues, and HGF receptor antagonists attenuated its anti-PF effect. Madecassoside facilitated the secretion of HGF from colonic epithelial cells by activating the PPAR-γ pathway, as shown by an up-regulation of PPAR-γ mRNA expression, nuclear translocation and DNA-binding activity both in vitro and in vivo. Also GW9662, a selective PPAR-γ antagonist, almost completely prevented the madecassoside-induced increased expression of HGF and amelioration of PF. The potent anti-PF effects induced by p.o. madecassoside in mice are not mediated by its metabolites or itself after absorption into blood. Instead, madecassoside increases the activity of PPAR-γ, which subsequently increases HGF expression in colonic epithelial cells. HGF then enters into the circulation and lung tissue to exert an anti-PF effect.

Fluorofenidone attenuates TGF-β1-induced lung fibroblast activation via restoring the expression of caveolin-1.

Caveolin-1 plays an important role in the pathogenesis of idiopathic pulmonary fibrosis. We previously showed that fluorofenidone (FD), a novel pyridine agent, can attenuate bleomycin-induced experimental pulmonary fibrosis and restore the production of caveolin-1. In this study, we explore mainly whether caveolin-1 plays a critical role in the anti-pulmonary fibrosis effects of FD in vitro. The normal human lung fibroblasts (NHLFs) were cultured with transforming growth factor-β1 (TGF-β1) and then were treated with FD. Subsequently, NHLFs transfected with cav-1-siRNA were treated with TGF-β1 and/or FD. The expressions of α-smooth muscle actin (α-SMA), fibronectin, collagen I, caveolin-1, phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated c-Jun N-terminal kinase (p-JNK), and phosphorylated P38 were measured by Western blot and/or real-time polymerase chain reaction. Fluorofenidone attenuated TGF-β1-induced expressions of α-SMA, fibronectin, and collagen I; inhibited phosphorylation of ERK, JNK, and P38; and restored caveolin-1 protein expression but cannot increase caveolin-1 mRNA level in vitro. After caveolin-1 was silenced, FD could not downregulate TGF-β1-induced expressions of α-SMA, fibronectin, and collagen I or phosphorylation of ERK, JNK, and P38. These studies demonstrate that FD, a potential antifibrotic agent, may attenuate TGF-β1-induced activation of NHLFs by restoring the expression of caveolin-1.

Prevention of pulmonary fibrosis with salvianolic acid A by inducing fibroblast cell cycle arrest and promoting apoptosis

Ethnopharmacological relevanceDanshen (Salvia miltiorrhiza Bunge) is widely used in traditional Chinese medicine (TCM), often in combination with other herbs, to treat a diversity of ailments. More recent studies have focused on its possible roles in the treatment of respiratory diseases (pneumonia and pulmonary fibrosis) and found that it has pharmacological activity that protects pulmonary morphology and function. However, the mechanism underlying this activity has not yet been clarified. Materials and methodsThe purpose of this study was to investigate the anti-pulmonary fibrosis effects exerted by salvianolic acid A (SAA), the ingredient responsible for the pharmacological activity of Danshen, and the underlying mechanisms. Bleomycin (BLM)-induced rat pulmonary fibrosis was used to evaluate the antifibrotic role of SAA, and fibroblast cells were used to study the mechanism involved. ResultsBLM-treated rats exhibited increased alveolar wall thickness and collagen deposition in lung tissues, but these pathologies were greatly attenuated by daily administration of SAA. We also found that SAA significantly inhibited the proliferation, adhesion and migration of fibroblasts in vitro. This was partly due to a strong induction of cell cycle arrest and apoptosis upon SAA treatment. Consistent with these phenotypes, we observed decreased expression of the cell cycle-related proteins cyclin D1, cyclin E1, and cyclin B1, and increased expression of p53 and p21 in SAA-treated cells. In addition, the anti-apoptotic Bcl-2 protein decreased in a dose-dependent manner, while cleaved caspase-3 protein increased upon SAA treatment. ConclusionsThese results suggest that the alleviation of rat pulmonary fibrosis by SAA is due to the inhibition of fibroblast proliferation and induction of apoptosis, which occurs mainly through p53-dependent growth arrest and apoptosis. We suggest that SAA should be considered as a potential novel therapeutic agent for the treatment of fibrotic lung diseases.