Anti-fibrosis effect for Hirsutella sinensis mycelium based on inhibition of mTOR p70S6K phosphorylation

Hirsutella sinensis, cultured in vitro, is an attractive substitute for Cordyceps sinensis as health supplement. The aim of this study was to demonstrate whether H. sinensis mycelium (HSM) attenuates murine pulmonary fibrosis induced by bleomycin and to explore the underlying molecular mechanisms. Using lung fibrosis modle induced by intratracheal instillation of bleomycin (BLM; 4 mg/kg), we observed that the administration of HSM reduced HYP, TGF-β1 and the production of several pro-fibrosis cytokines (α-smooth muscle actin, fibronectin and vimentin) in fibrotic mice lung sections. Histopathological examination of lung tissues also demonstrated that HSM improved BLM-induced pathological damage. Concurrently, HSM supplementation markedly reduced the chemotaxis of alveolar macrophages and potently suppressed the expression of inflammatory cytokines. Also, HSM influenced Th1/Th2 and Th17/Treg imbalance and blocked the phosphorylation of mTOR pathway in vivo. Alveolar epithelial A549 cells acquired a mesenchymal phenotype and an increased expression of myofibroblast markers of differentiation (vimentin and fibronectin) after treatment with TGF-β1. HSM suppressed these markers and blocked the phosphorylation of mTOR pathway in vitro. The results provide evidence supporting the use of HSM in the intervention of pulmonary fibrosis and suggest that HSM is a potential therapeutic agent for lung fibrosis.

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Knockdown of Long Noncoding RNA H19 Represses the Progress of Pulmonary Fibrosis through the Transforming Growth Factor β/Smad3 Pathway by Regulating MicroRNA 140

Molecular and Cellular Biology ◽

10.1128/mcb.00143-19 ◽

2019 ◽

Vol 39 (12) ◽

Cited By ~ 14

Author(s):

Xi Wang ◽

Zhe Cheng ◽

Lingling Dai ◽

Tianci Jiang ◽

Liuqun Jia ◽

...

Keyword(s):

Growth Factor ◽

Pulmonary Fibrosis ◽

Noncoding Rna ◽

Molecular Mechanisms ◽

Transforming Growth Factor ◽

Animal Experiments ◽

A549 Cells ◽

Transforming Growth Factor Β ◽

Tgf Β1

ABSTRACT Long noncoding RNAs (lncRNAs) are involved in various human diseases. Recently, H19 was reported to be upregulated in fibrotic rat lung and play a stimulative role in bleomycin (BLM)-induced pulmonary fibrosis in mice. However, its expression in human fibrotic lung tissues and mechanism of action remain unclear. Here, our observations showed that H19 expression was significantly upregulated and that of microRNA 140 (miR-140) was markedly reduced in pulmonary fibrotic tissues from idiopathic pulmonary fibrosis (IPF) patients and transforming growth factor β1 (TGF-β1)-induced HBE and A549 cells. Moreover, the expression of H19 was negatively correlated with the expression of miR-140 in IPF tissues. H19 knockdown attenuated TGF-β1-induced pulmonary fibrosis in vitro. Furthermore, animal experiments showed that H19 knockdown attenuated BLM-induced pulmonary fibrosis in mice. The study of molecular mechanisms showed that H19 functioned via reduction of miR-140 expression by binding to miR-140. The increase of miR-140 inhibited TGF-β1-induced pulmonary fibrosis, and H19 upregulation diminished the inhibitory effects of miR-140 on TGF-β1-induced pulmonary fibrosis, which was involved in the TGF-β/Smad3 pathway. Taken together, our findings showed that H19 knockdown attenuated pulmonary fibrosis via the regulatory network of lncRNA H19–miR-140–TGF-β/Smad3 signaling, and H19 and miR-140 might represent therapeutic targets and early diagnostic and prognostic biomarkers for patients with pulmonary fibrosis.

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Inhibition of Raf1 ameliorates bleomycin-induced pulmonary fibrosis through attenuation of TGF-β1 signaling

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00093.2018 ◽

2018 ◽

Vol 315 (2) ◽

pp. L241-L247 ◽

Cited By ~ 3

Author(s):

Shuang Li ◽

Jia Liu ◽

Jiangning Tan ◽

Lian Li ◽

Mary J. Kaltreider ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Lung Fibrosis ◽

Molecular Mechanisms ◽

Human Lung ◽

Smooth Muscle Actin ◽

Lung Fibroblasts ◽

Human Lung Fibroblasts ◽

And Migration ◽

Fibrotic Lung Disease ◽

Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease associated with aberrant activation and differentiation of fibroblasts, leading to abnormal extracellular matrix production. Currently, it is still an untreatable disease (except for lung transplantation). Here, we demonstrate that the Raf1 inhibitor GW5074 ameliorates lung fibrosis in bleomycin-induced pulmonary fibrosis. Posttreatment with GW5074 reduced fibronectin (FN) expression, collagen deposition, and inflammatory cell infiltration in bleomycin-challenged mice, suggesting an antifibrotic property of GW5074. To determine the molecular mechanisms by which inhibition of Raf1 ameliorates lung fibrosis, we investigated the role of Raf1 in TGF-β1 signaling in human lung fibroblasts. GW5074 or downregulation of Raf1 by siRNAs significantly attenuated TGF-β1-induced smooth muscle actin, FN, and collagen I expression, whereas overexpression of Raf1 promoted the effects of TGF-β1 in lung fibroblasts. Furthermore, we found that Raf1-promoted TGF-β1 signaling was through the Raf1/ERK/Smad pathway and contributed to the cell proliferation and migration in human lung fibroblasts. This study provides preclinical and mechanistic evidence for development of Raf1 inhibitors as potential antifibrotic drugs for the treatment of IPF.

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Anti-Fibrosis Effects of Magnesium Lithospermate B in Experimental Pulmonary Fibrosis: By Inhibiting TGF-βRI/Smad Signaling

Molecules ◽

10.3390/molecules26061715 ◽

2021 ◽

Vol 26 (6) ◽

pp. 1715

Author(s):

Xin Luo ◽

Qiangqiang Deng ◽

Yaru Xue ◽

Tianwei Zhang ◽

Zhitao Wu ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Cell Line ◽

Molecular Mechanisms ◽

Transforming Growth Factor ◽

Salvia Miltiorrhiza ◽

A549 Cells ◽

Epithelial Cell Line ◽

Human Lung Fibroblast ◽

Magnesium Lithospermate B

Pulmonary fibrosis is a severe and irreversible interstitial pulmonary disease with high mortality and few treatments. Magnesium lithospermate B (MLB) is a hydrosoluble component of Salvia miltiorrhiza and has been reported to have antifibrotic effects in other forms of tissue fibrosis. In this research, we studied the effects of MLB on pulmonary fibrosis and the underlying mechanisms. Our results indicated that MLB treatment (50 mg/kg) for seven days could attenuate bleomycin (BLM)-induced pulmonary fibrosis by reducing the alveolar structure disruption and collagen deposition in the C57 mouse model. MLB was also found to inhibit transforming growth factor-beta (TGF-β)-stimulated myofibroblastic transdifferentiation of human lung fibroblast cell line (MRC-5) cells and collagen production by human type II alveolar epithelial cell line (A549) cells, mainly by decreasing the expression of TGF-β receptor I (TGF-βRI) and regulating the TGF-β/Smad pathway. Further studies confirmed that the molecular mechanisms of MLB in BLM-induced pulmonary fibrosis mice were similar to those observed in vitro. In summary, our results demonstrated that MLB could alleviate experimental pulmonary fibrosis both in vivo and in vitro, suggesting that MLB has great potential for pulmonary fibrosis treatment.

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A Novel CD206 Targeting Peptide Inhibits Bleomycin Induced Pulmonary Fibrosis in Mice

10.1101/2020.07.27.218115 ◽

2020 ◽

Author(s):

Anghesom Ghebremedhin ◽

Ahmad Bin Salam ◽

Benjamin Adu-Addai ◽

Steve Noonan ◽

Richard Stratton ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Lung Fibrosis ◽

Early Stage ◽

Blood Chemistry ◽

Dependent Manner ◽

Similar Reduction ◽

Clinical Scenarios ◽

Approved Drugs ◽

Tgf Β1

AbstractActivated M2 polarized macrophages are drivers of pulmonary fibrosis in several clinical scenarios such as Acute Respiratory Disease Syndrome (ARDS) and Idiopathic Pulmonary Fibrosis (IPF), through the production of inflammatory and fibrosis-inducing cytokines. In this study, we investigated the effect of targeting the CD206 receptor with a novel fragment of a Host Defense Peptide (HDP), RP-832c to decrease cytokines that cause fibrosis. RP-832c selectively binds to CD206 on M2 polarized bone marrow derived macrophages (BMDM) in vitro, resulting in a time-dependent decrease in CD206 expression, and a transient increase in M1 marker TNFα, which resolves over a 24hr period. To elucidate the antifibrotic effect of RP-832c, we used a murine model of bleomycin (BLM) -induced early-stage pulmonary fibrosis. RP-832c significantly reduced bleomycin-induced fibrosis in a dosage dependent manner, as well as decreased CD206, TGF-β1 and α-SMA expression in mouse lungs. Interestingly we did not observe any changes in the resident alveolar macrophage marker CD170 expression. Similarly, in an established model of lung fibrosis, RP-832c significantly decreased fibrosis in the lung, as well as significantly decreased inflammatory cytokines TNFα, IL-6, IL-10, INF-γ, CXCL1/2, and fibrosis markers TGF-β1 and MMP-13. In comparison with FDA approved drugs, Nintedanib and Pirfenidone, RP-832c exhibited a similar reduction in fibrosis compared to Pirfenidone, and to a greater extent than Nintedanib, with no apparent toxicities observed on body weight or blood chemistry. In summary, RP-832c is a potential agent to mitigate the overactivity of M2 macrophages in pathogenesis several pulmonary fibrotic diseases, including SARS-CoV-2 induced lung fibrosis.

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Bleomycin induces epithelial-to-mesenchymal transition via bFGF/PI3K/ESRP1 signaling in pulmonary fibrosis

Bioscience Reports ◽

10.1042/bsr20190756 ◽

2020 ◽

Vol 40 (1) ◽

Cited By ~ 1

Author(s):

Chang-Mei Weng ◽

Qing Li ◽

Kui-Jun Chen ◽

Cheng-Xiong Xu ◽

Meng-Sheng Deng ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Epithelial Mesenchymal Transition ◽

Regulatory Protein ◽

A549 Cells ◽

Akt Signaling ◽

High Rate ◽

Akt Inhibitor ◽

Mesenchymal Transition ◽

Tgf Β1

Abstract Idiopathic pulmonary fibrosis (IPF) is a fatal and chronic disease with a high rate of infection and mortality; however, its etiology and pathogenesis remain unclear. Studies have revealed that epithelial–mesenchymal transition (EMT) is a crucial cellular event in IPF. Here, we identified that the pulmonary fibrosis inducer bleomycin simultaneously increased the expression of bFGF and TGF-β1 and inhibited epithelial-specific regulatory protein (ESRP1) expression in vivo and in vitro. In addition, in vitro experiments showed that bFGF and TGF-β1 down-regulated the expression of ESRP1 and that silencing ESRP1 promoted EMT in A549 cells. Notably, we determined that bFGF activates PI3K/Akt signaling, and treatment with the PI3K/Akt inhibitor LY294002 inhibited bleomycin-induced cell morphology changes and EMT. In addition, the effects of LY294002 on bleomycin-induced EMT were inhibited by ESRP1 silencing in A549 cells. Taken together, these findings suggest that bleomycin induced EMT through down-regulating ESRP1 by simultaneously increasing bFGF and TGF-β1 in pulmonary fibrosis. Additionally, our findings indicated that bFGF inhibits ESRP1 by activating PI3K/Akt signaling.

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Arctigenin Suppressed Epithelial-Mesenchymal Transition Through Wnt3a/β-Catenin Pathway in PQ-Induced Pulmonary Fibrosis

Frontiers in Pharmacology ◽

10.3389/fphar.2020.584098 ◽

2020 ◽

Vol 11 ◽

Author(s):

Fei Gao ◽

Yun Zhang ◽

Zhizhou Yang ◽

Mengmeng Wang ◽

Zhiyi Zhou ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Signaling Pathway ◽

Lung Fibrosis ◽

Epithelial Mesenchymal Transition ◽

A549 Cells ◽

Underlying Mechanism ◽

Alveolar Type ◽

Mesenchymal Transition

Arctigenin (ATG), a major bioactive substance of Fructus Arctii, counters renal fibrosis; however, whether it protects against paraquat (PQ)-induced lung fibrosis remains unknown. The present study was to determine the effect of ATG on PQ-induced lung fibrosis in a mouse model and the underlying mechanism. Firstly, we found that ATG suppressed PQ-induced pulmonary fibrosis by blocking the epithelial-mesenchymal transition (EMT). ATG reduced the expressions of Vimentin and α-SMA (lung fibrosis markers) induced by PQ and restored the expressions of E-cadherin and Occludin (two epithelial markers) in vivo and in vitro. Besides, the Wnt3a/β-catenin signaling pathway was significantly activated in PQ induced pulmonary fibrosis. Further analysis showed that pretreatment of ATG profoundly abrogated PQ-induced EMT-like phenotypes and behaviors in A549 cells. The Wnt3a/β-catenin signaling pathway was repressed by ATG treatment. The overexpression of Wnt3a could weaken the therapeutic effect of ATG in A549 cells. These findings suggested that ATG could serve as a new therapeutic candidate to inhibit or even reverse EMT-like changes in alveolar type II cells during PQ-induced lung fibrosis, and unraveled that the Wnt3a/β-catenin pathway might be a mechanistic tool for ATG to control pulmonary fibrosis.

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Downregulation of S100A2 alleviates pulmonary fibrosis through inhibiting wnt/β-catenin signaling pathway

10.21203/rs.3.rs-23375/v1 ◽

2020 ◽

Author(s):

Guichuan Huang ◽

Jing Zhang ◽

Gang Qing ◽

Daishun Liu ◽

Xin Wang ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Western Blot ◽

Molecular Mechanisms ◽

Transforming Growth Factor ◽

Epithelial Mesenchymal Transition ◽

A549 Cells ◽

Mesenchymal Transition ◽

Qrt Pcr ◽

Tgf Β1

Abstract Background：Pulmonary fibrosis (PF) is a progressive and lethal disease with poor prognosis. S100A2 plays an important role in the progression of cancer. However, the role of S100A2 in PF has not been reported yet. In this study, we explored the potential role of S100A2 in PF and its potential molecular mechanisms. Methods: First, we analyzed S100A2 expression of patients with PF by retrieving RNA-sequencing datasets from Gene Expression Omnibus (GEO) database. Next, we detected the expression of S100A2 in patients with PF using quantitative real time PCR (qRT-PCR). Then, S100A2 expression was determined with or without the treatment of transforming growth factor-β1 (TGF-β1) in A549 cells. Epithelial-mesenchymal transition (EMT) biomarkers, including E-cadherin，vimentin, and α smooth muscle actin (α-SMA), were identified using qRT-PCR and western blot. Finally, the relevant signalling pathway indicators were detected by western blot. Results: Increased expression of S100A2 was first observed in lung tissues of PF patients. Meanwhile, we found that downregulation of S100A2 inhibited the TGF-β1-induced EMT in A549 cells. Mechanically, TGF-β1 up-regulated β-catenin and phosphorylation of GSK-3β, which was blocked by silencing S100A2 in vitro. Conclusion: These findings demonstrate that downregulation of S100A2 alleviate pulmonary fibrosis via inhibiting EMT. S100A2 is a promising potential target for further understanding the mechanism and developing strategy for the treatment of PF and other EMT-associated disease.

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Pterostilbene Alleviates Pulmonary Fibrosis by Regulating ASIC2

10.21203/rs.3.rs-127867/v1 ◽

2020 ◽

Author(s):

Yanfang Peng ◽

Yingwen Zhang ◽

Yabing Zhang ◽

Xiuping Wang ◽

Yukun Xia ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Western Blot ◽

High Throughput ◽

Protein Level ◽

High Throughput Sequencing ◽

Epithelial Mesenchymal Transition ◽

A549 Cells ◽

Mesenchymal Transition ◽

Tgf Β1

Abstract Background: Idiopathic pulmonary fibrosis (IPF) is a serious chronic disease of the respiratory system, and its current treatment have certain shortcomings and adverse effects. In this study, we evaluate the anti-fibrotic activity of pterostilbene (PTE) using an IPF model induced by TGF-β1 in vitro.Methods: A549 and AEC cells were incubated with 10 ng/ml TGF-β1 to induce lung fibroblast activation. 30 μmol/L PTE was used to treat the cells. The epithelial-mesenchymal transition (EMT), accumulation of extracellular matrix (ECM) and autophagy of cells were suggested by western blot. The apoptosis was proved by flow cytometry analysis and western blot. Transcriptome high-throughput sequencing on A549 cells incubated with TGF-β1 alone or TGF-β1 and PTE (TGF-β1+PTE) was performed, and differentially expressed genes caused by PTE were identified. The ASIC2 overexpression plasmid was used to rescue the protein level of ASIC2 in A549 and AEC cells.Results: TGF-β1 caused the EMT and accumulation of ECM, and blocked the autophagy and apoptosis of A549 and AEC cells. Most importantly, 30 μmol/L PTE inhibited the pulmonary fibrosis induced by TGF-β1. Compared with cells treated with TGF-β1, PTE treatment inhibited the EMT and accumulation of ECM, and rescued cell apoptosis and autophagy. The results of transcriptome high-throughput sequencing performed that PTE greatly reduced the protein level of ASIC2. In addition, compared with the TGF-β1+PTE group, the transfection of ASIC2 overexpression plasmid stimulated the EMT and accumulation of ECM, and inhibited apoptosis and autophagy, suggesting that PTE inhibited pulmonary fibrosis by down-regulating ASIC2. Conclusions: In conclusion, our study suggests that PTE and ASIC2 inhibitors may benefit future IPF treatments.

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Tetrandrine modulates SQSTM1-mediated selective autophagy and protects pulmonary fibrosis

10.22541/au.161185451.12306983/v1 ◽

2021 ◽

Author(s):

Yuanyuan Liu ◽

Wenshan Zhong ◽

Jinming Zhang ◽

Weimou Chen ◽

Ye Lu ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Lung Fibrosis ◽

Smooth Muscle Actin ◽

Lung Fibroblasts ◽

Therapeutic Effects ◽

The Novel ◽

Induced Expression ◽

Tgf Β1

Background and Purpose Idiopathic pulmonary fibrosis is a progressive fatal disease characterized by interstitial remodeling, with high lethality and a lack of effective medical therapies. Tetrandrine has been proposed to present anti-fibrotic effects, but the efficacy and mechanisms of tetrandrine against lung fibrosis has not been systematically evaluated. We sought to study the potential therapeutic effects and mechanisms of tetrandrine in lung fibrosis. Experimental Approach The anti-fibrotic effects of tetrandrine were evaluated in bleomycin-induced mouse models and TGF-β1-stimulated murine lung fibroblasts. We performed Chromatin Immunoprecipitation (ChIP), Immunoprecipitation (IP) and mRFP-GFP-MAP1LC3B adenovirus construct to investigate the novel mechanisms of tetrandrine-induced autophagy. Key Results Tetrandrine decreased TGF-β1-induced expression of α-smooth muscle actin, fibronectin, vimentin and type 1 collagen and proliferation in fibroblasts. Tetrandrine restored TGF-β1-induced impaired autophagy, accompanied by the up-regulation and enhanced interaction of SQSTM1 and MAP1LC3-Ⅱ. ChIP studies revealed that NRF2 bound to SQSTM1 promoter in tetrandrine-induced autophagy. Furthermore, TGF-β1-induced phosphorylated mTOR was inhibited by tetrandrine, with reduced activation levels of Rheb. In vivo tetrandrine suppressed the bleomycin-induced expression of fibrotic markers and improved pulmonary function. Conclusion and Implications Our data suggest that tetrandrine might be recognized as a novel autophagy inducer, thus attenuating lung fibrosis. Tetrandrine should be investigated as a novel therapeutic strategy for IPF.

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Dual Effects of Hypoxia-Inducible Factors-1 Alpha in Bleomycin-Induced Pulmonary Fibrosis Treated by Human Umbilical Cord Mesenchymal Stem Cells

Stem Cells International ◽

10.1155/2021/6658855 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Hongpeng Zhang ◽

Hao Wang ◽

Yong Xia ◽

Nianmin Qi

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Pulmonary Fibrosis ◽

Umbilical Cord ◽

Smooth Muscle Actin ◽

A549 Cells ◽

Human Umbilical Cord ◽

Therapeutic Administration

Pulmonary fibrosis (PF) is a kind of lung disease characterized by scar formation and inflammation damage. Mesenchymal stem cells (MSCs) are considered a promising therapy because of multidirectional differentiation and immune regulation. Our research was designed for identifying the preventative defensive ability and therapeutic effect of human umbilical cord mesenchymal stem cells (HUCMSCs). HUCMSCs were administered before or after bleomycin injection in different groups of C57BL/6 mice. We calculated the survival time of mice, the lung coefficients, contents of hydroxyproline, and pathological scores. The expression levels of HIF-1α (hypoxia-inducible factor-1α), α-SMA (α-smooth muscle actin), γH2AFX (γH2A histone family, member X), ZO-1 (zonula occludens-1), ROS (reactive oxygen species) content, and proliferation ability of A549 cells were detected after treatment with bleomycin and HUCMSCs conditioned medium (HUCMSCs-CM), respectively, or together in vitro. In addition, we examined the secretome of HUCMSCs in regular and inflammatory stimulation conditions. Our results demonstrated that prophylactic HUCMSC administration before bleomycin-induced modeling process could significantly meliorate damage to pulmonary fibrosis. After the deletion of HIF-1α, damage markers in A549 cells were significantly reduced in therapeutic administration condition. However, it was the opposite in prophylactic administration condition. The results confirmed that HUCMSCs had available preventive effect on bleomycin-induced pulmonary fibrosis in vivo and in vitro. However, it may have a negative effect in therapeutic administration condition because of the dual effect of HIF-1α.

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