The Extracellular Matrix Enriched With Exosomes for the Treatment on Pulmonary Fibrosis in Mice

Pulmonary fibrosis (PF) is a severe respiratory disease caused by lung microenvironment changes. TGF-β/Smad3 signaling pathway plays a critical role in the fibrotic process. MicroRNA-29 (miR-29) has proved to alleviate the occurrence of PF by downregulating TGF-β/Smad3 signaling pathway. The miRNA application encounters obstacles due to its low stability in body and no targeting to lesions. Exosomes can be used for therapeutic delivery of miRNA due to their favorable delivery properties. However, low efficiency of separation and production impedes the therapeutic application of exosomes. In this study, we developed a liquid natural extracellular matrix (ECM) enriched with miR-29-loaded exosomes for PF treatment. The collagen-binding domain (CBD)-fused Lamp2b (CBD-Lamp2b) and miR-29 were overexpressed in human foreskin fibroblast (HFF) host cells for the entrapment of miR-29-loaded exosomes in ECM of the cells. The repeated freeze-thaw method was performed to prepare the liquid ECM enriched with exosomes without destroying the exosomal membrane. In summary, this study developed a novel functional ECM biomaterial for therapy of PF, and also provided a promising gene therapy platform for different diseases by treatment with liquid ECM that is, enriched with exosomes loaded with different functional miRNAs.

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The promise of mTOR as a therapeutic target pathway in idiopathic pulmonary fibrosis

European Respiratory Review ◽

10.1183/16000617.0269-2020 ◽

2020 ◽

Vol 29 (157) ◽

pp. 200269

Author(s):

Manuela Platé ◽

Delphine Guillotin ◽

Rachel C Chambers

Keyword(s):

Extracellular Matrix ◽

Idiopathic Pulmonary Fibrosis ◽

Pulmonary Fibrosis ◽

Therapeutic Target ◽

Critical Role ◽

Mammalian Target Of Rapamycin ◽

Lung Parenchyma ◽

Extracellular Signals ◽

Novel Therapeutic Strategies ◽

Novel Therapeutic

Idiopathic pulmonary fibrosis (IPF) is characterised by the progressive deposition of excessive extracellular matrix proteins within the lung parenchyma and represents the most rapidly progressive and fatal of all fibrotic conditions. Current anti-fibrotic drugs approved for the treatment of IPF fail to halt disease progression and have significant side-effect profiles. Therefore, there remains a pressing need to develop novel therapeutic strategies for IPF. Mammalian target of rapamycin (mTOR) forms the catalytic subunit of two complexes, mTORC1 and mTORC2. mTORC1 acts as critical cellular sensor which integrates intracellular and extracellular signals to reciprocally regulate a variety of anabolic and catabolic processes. The emerging evidence for a critical role for mTORC1 in influencing extracellular matrix production, metabolism, autophagy and senescence in the setting of IPF highlights this axis as a novel therapeutic target with the potential to impact multiple IPF pathomechanisms.

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Extracellular CIRP Induces an Inflammatory Phenotype in Pulmonary Fibroblasts via TLR4

Frontiers in Immunology ◽

10.3389/fimmu.2021.721970 ◽

2021 ◽

Vol 12 ◽

Author(s):

Siavash Bolourani ◽

Ezgi Sari ◽

Max Brenner ◽

Ping Wang

Keyword(s):

Pulmonary Fibrosis ◽

Inflammatory Cytokines ◽

Rna Binding ◽

Critical Role ◽

Dependent Manner ◽

Gene Expressions ◽

Pulmonary Fibroblasts ◽

Molecular Pattern ◽

Fibrotic Process ◽

Inflammatory Phenotype

Extracellular cold-inducible RNA-binding protein (eCIRP), a new damage-associated molecular pattern (DAMP), has been recently shown to play a critical role in promoting the development of bleomycin-induced pulmonary fibrosis. Although fibroblast activation is a critical component of the fibrotic process, the direct effects of eCIRP on fibroblasts have never been examined. We studied eCIRP’s role in the induction of inflammatory phenotype in pulmonary fibroblasts and its connection to bleomycin-induced pulmonary fibrosis in mice. We found that eCIRP causes the induction of proinflammatory cytokines and differentially expression-related pathways in a TLR4-dependent manner in pulmonary fibroblasts. Our analysis further showed that the accessory pathways MD2 and Myd88 are involved in the induction of inflammatory phenotype. In order to study the connection of the enrichment of these pathways in priming the microenvironment for pulmonary fibrosis, we investigated the gene expression profile of lung tissues from mice subjected to bleomycin-induced pulmonary fibrosis collected at various time points. We found that at day 14, which corresponds to the inflammatory-to-fibrotic transition phase after bleomycin injection, TLR4, MD2, and Myd88 were induced, and the transcriptome was differentially enriched for genes in those pathways. Furthermore, we also found that inflammatory cytokines gene expressions were induced, and the cellular responses to these inflammatory cytokines were differentially enriched on day 14. Overall, our results show that eCIRP induces inflammatory phenotype in pulmonary fibroblasts in a TLR4 dependent manner. This study sheds light on the mechanism by which eCIRP induced inflammatory fibroblasts, contributing to pulmonary fibrosis.

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Dynamic Observation of Autophagy and Transcriptome Profiles in a Mouse Model of Bleomycin-Induced Pulmonary Fibrosis

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2021.664913 ◽

2021 ◽

Vol 8 ◽

Author(s):

Yani Wang ◽

Siqi Hu ◽

Lisha Shen ◽

Song Liu ◽

Linyan Wan ◽

...

Keyword(s):

Gene Expression ◽

Pulmonary Fibrosis ◽

Mouse Model ◽

Muscle Contraction ◽

Signaling Pathway ◽

Lung Tissue ◽

Critical Role ◽

Mouse Lung ◽

Mouse Lung Tissue

Pulmonary fibrosis is a group of progressive, fibrotic, and fatal lung diseases, and the role of autophagy in pulmonary fibrosis is controversial. In the current research, we dynamically observed a bleomycin-induced pulmonary fibrosis mouse model after 3, 7, 14, 21, and 28 days and investigated the expression of autophagy markers. We found that autophagy markers were not significantly changed on the indicated days in the mouse lung tissue. Then, RNA-Seq was used to analyze the gene expression and associated functions and pathways in fibrotic lung tissue on different days post-bleomycin. In addition, short time series expression miner (STEM) analysis was performed to explore the temporal post-bleomycin gene expression. Through STEM, continually up- or downregulated profiles did not demonstrate the critical role of autophagy in the development of fibrosis. Furthermore, gene ontology (GO) annotations showed that continually upregulated profiles were mainly related to fibrosis synthesis, extracellular space, and inflammation, while enriched pathways were mainly related to the PI3K-Akt signaling pathway, ECM–receptor interactions, and focal adhesion signaling pathway. For continually downregulated profiles, GO annotations mainly involved sarcomere organization, muscle contraction, and muscle fiber development. The enriched KEGG signaling pathways were the cAMP signaling pathway, cGMP-PKG signaling pathway, calcium signaling pathway, and cardiac muscle contraction. Moreover, we analyzed autophagy-related genes’ expression in specific cells from a publicly available database of three human and one animal study of pulmonary fibrosis using single-cell sequencing technology. All results consistently demonstrated no critical role of autophagy in the pathogenesis of pulmonary fibrosis. In summary, autophagy may not critically and consistently change during the development of pulmonary fibrosis at different stages post-bleomycin in a mouse model. These continually up- or downregulated profiles, including gene profiles, and the corresponding functions and pathways may provide mechanistic insights into IPF therapy.

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Ionizing radiation induces myofibroblast differentiation via lactate dehydrogenase

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00153.2015 ◽

2015 ◽

Vol 309 (8) ◽

pp. L879-L887 ◽

Cited By ~ 17

Author(s):

J. L. Judge ◽

K. M. Owens ◽

S. J. Pollock ◽

C. F. Woeller ◽

T. H. Thatcher ◽

...

Keyword(s):

Extracellular Matrix ◽

Lactate Dehydrogenase ◽

Pulmonary Fibrosis ◽

Ionizing Radiation ◽

Lactate Production ◽

Lung Fibroblasts ◽

Myofibroblast Differentiation ◽

Dependent Manner ◽

Fibrotic Process ◽

Radiation Induced

Pulmonary fibrosis is a common and dose-limiting side-effect of ionizing radiation used to treat cancers of the thoracic region. Few effective therapies are available for this disease. Pulmonary fibrosis is characterized by an accumulation of myofibroblasts and excess deposition of extracellular matrix proteins. Although prior studies have reported that ionizing radiation induces fibroblast to myofibroblast differentiation and collagen production, the mechanism remains unclear. Transforming growth factor-β (TGF-β) is a key profibrotic cytokine that drives myofibroblast differentiation and extracellular matrix production. However, its activation and precise role in radiation-induced fibrosis are poorly understood. Recently, we reported that lactate activates latent TGF-β through a pH-dependent mechanism. Here, we wanted to test the hypothesis that ionizing radiation leads to excessive lactate production via expression of the enzyme lactate dehydrogenase-A (LDHA) to promote myofibroblast differentiation. We found that LDHA expression is increased in human and animal lung tissue exposed to ionizing radiation. We demonstrate that ionizing radiation induces LDHA, lactate production, and extracellular acidification in primary human lung fibroblasts in a dose-dependent manner. We also demonstrate that genetic and pharmacologic inhibition of LDHA protects against radiation-induced myofibroblast differentiation. Furthermore, LDHA inhibition protects from radiation-induced activation of TGF-β. We propose a profibrotic feed forward loop, in which radiation induces LDHA expression and lactate production, which can lead to further activation of TGF-β to drive the fibrotic process. These studies support the concept of LDHA as an important therapeutic target in radiation-induced pulmonary fibrosis.

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mTORC1 Signaling Pathway Mediated SIRT6 Overexpression in TGF-β1-Induced Pulmonary Fibrosis

10.21203/rs.3.rs-1149371/v1 ◽

2021 ◽

Author(s):

Ji Zhang ◽

Yi Hu ◽

Huiping Huang ◽

Qun Liu ◽

Yang Du ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Signaling Pathway ◽

Critical Role ◽

Mtor Signaling ◽

Lung Fibroblasts ◽

Dependent Manner ◽

Mtor Signaling Pathway ◽

Mtorc1 Signaling ◽

Mtorc1 Pathway ◽

Tgf Β1

Abstract Fibroblast-to-myofibroblast transdifferentiation and myofibroblast hyperproliferation play a major role in Idiopathic pulmonary fibrosis (IPF). It was also reported that mTOR signaling pathway and SIRT6 have a critical role in pulmonary fibrosis. However, the mechanisms whether mTOR signaling pathway and SIRT6 affect the myofibroblasts differentiation in IPF remain unclear. The results show that SIRT6 is significantly upregulated by TGF-β1 with a time and concentration-dependent manner in MRC5 line and primary lung fibroblasts isolated from IPF patients. SIRT6 protein is also increased in IPF fibrotic lung tissues and bleomycin-challenged mice lung tissues. Also, the activity of mTOR signaling is activated in MRC5 and primary lung fibroblasts. Furthermore, the inhibitor of mTOR, rapamycin treatment significantly suppress mTORC1 pathway activity and SIRT6 protein expression. SIRT6 siRNA failed to mediate the activity of mTORC1 pathway and autophagy induction. Finally, deficiency of SIRT6 could promote TGF-β1 induced pro-fibrotic cytokines. In summary, the study have suggested that SIRT6 is a downstream of mTORC1 signaling pathway in the pulmonary fibrosis caused by TGF-β1-induced. Deficiency of SIRT6 mediated myofibroblasts differentiation through induced pro-fibrotic cytokines production but not induced-autophagy. It was indicated that manipulations of SIRT6 expression may provide a new therapeutic strategy to reverse the progression of pulmonary fibrosis.

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TRIM21 Promotes Innate Immune Response to RNA Viral Infection through Lys27-Linked Polyubiquitination of MAVS

Journal of Virology ◽

10.1128/jvi.00321-18 ◽

2018 ◽

Vol 92 (14) ◽

Cited By ~ 25

Author(s):

Binbin Xue ◽

Huiyi Li ◽

Mengmeng Guo ◽

Jingjing Wang ◽

Yan Xu ◽

...

Keyword(s):

Immune Response ◽

Innate Immunity ◽

Viral Infection ◽

Signaling Pathway ◽

Innate Immune Response ◽

Rna Virus ◽

Critical Role ◽

Innate Immune ◽

Host Cells ◽

Lysine Residues

ABSTRACT Human innate immunity responds to viral infection by activating the production of interferons (IFNs) and proinflammatory cytokines. The mitochondrial adaptor molecule MAVS plays a critical role in innate immune response to viral infection. In this study, we show that TRIM21 (tripartite motif-containing protein 21) interacts with MAVS to positively regulate innate immunity. Under viral infection, TRIM21 is upregulated through the IFN/JAK/STAT signaling pathway. Knockdown of TRIM21 dramatically impairs innate immune response to viral infection. Moreover, TRIM21 interacts with MAVS and catalyzes its K27-linked polyubiquitination, thereby promoting the recruitment of TBK1 to MAVS. Specifically, the PRY-SPRY domain of TRIM21 is the key domain for its interaction with MAVS, while the RING domain of TRIM21 facilitates the polyubiquitination chains of MAVS. In addition, the MAVS-mediated innate immune response is enhanced by both the PRY-SPRY and RING domains of TRIM21. Mutation analyses of all the lysine residues of MAVS further revealed that Lys325 of MAVS is catalyzed by TRIM21 for the K27-linked polyubiquitination. Overall, this study reveals a novel mechanism by which TRIM21 promotes the K27-linked polyubiquitination of MAVS to positively regulate innate immune response, thereby inhibiting viral infection. IMPORTANCE Activation of innate immunity is essential for host cells to restrict the spread of invading viruses and other pathogens. MAVS plays a critical role in innate immune response to RNA viral infection. In this study, we demonstrated that TRIM21 targets MAVS to positively regulate innate immunity. Notably, TRIM21 targets and catalyzes K27-linked polyubiquitination of MAVS and then promotes the recruitment of TBK1 to MAVS, leading to upregulation of innate immunity. Our study outlines a novel mechanism by which the IFN signaling pathway blocks RNA virus to escape immune elimination.

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Regorafenib-Attenuated, Bleomycin-Induced Pulmonary Fibrosis by Inhibiting the TGF-β1 Signaling Pathway

International Journal of Molecular Sciences ◽

10.3390/ijms22041985 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1985

Author(s):

Xiaohe Li ◽

Ling Ma ◽

Kai Huang ◽

Yuli Wei ◽

Shida Long ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Signaling Pathway ◽

Transforming Growth Factor ◽

Kinase Inhibitor ◽

In Vivo Experiments ◽

Age Related ◽

And Migration ◽

Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a fatal and age-related pulmonary disease. Nintedanib is a receptor tyrosine kinase inhibitor, and one of the only two listed drugs against IPF. Regorafenib is a novel, orally active, multi-kinase inhibitor that has similar targets to nintedanib and is applied to treat colorectal cancer and gastrointestinal stromal tumors in patients. In this study, we first identified that regorafenib could alleviate bleomycin-induced pulmonary fibrosis in mice. The in vivo experiments indicated that regorafenib suppresses collagen accumulation and myofibroblast activation. Further in vitro mechanism studies showed that regorafenib inhibits the activation and migration of myofibroblasts and extracellular matrix production, mainly through suppressing the transforming growth factor (TGF)-β1/Smad and non-Smad signaling pathways. In vitro studies have also indicated that regorafenib could augment autophagy in myofibroblasts by suppressing TGF-β1/mTOR (mechanistic target of rapamycin) signaling, and could promote apoptosis in myofibroblasts. In conclusion, regorafenib attenuates bleomycin-induced pulmonary fibrosis by suppressing the TGF-β1 signaling pathway.

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Acceleration of TAA-Induced Liver Fibrosis by Stress Exposure Is Associated with Upregulation of Nerve Growth Factor and Glycopattern Deviations

International Journal of Molecular Sciences ◽

10.3390/ijms22105055 ◽

2021 ◽

Vol 22 (10) ◽

pp. 5055

Author(s):

Catalina Atorrasagasti ◽

Flavia Piccioni ◽

Sophia Borowski ◽

Irene Tirado-González ◽

Nancy Freitag ◽

...

Keyword(s):

Nerve Growth Factor ◽

Growth Factor ◽

Liver Fibrosis ◽

Signaling Pathway ◽

Liver Damage ◽

Neurotrophin Receptor ◽

P75 Neurotrophin Receptor ◽

Nerve Growth ◽

Fibrotic Process ◽

Ngf Signaling

Liver fibrosis results from many chronic injuries and may often progress to cirrhosis and hepatocellular carcinoma (HCC). In fact, up to 90% of HCC arise in a cirrhotic liver. Conversely, stress is implicated in liver damage, worsening disease outcome. Hence, stress could play a role in disrupting liver homeostasis, a concept that has not been fully explored. Here, in a murine model of TAA-induced liver fibrosis we identified nerve growth factor (NGF) to be a crucial regulator of the stress-induced fibrogenesis signaling pathway as it activates its receptor p75 neurotrophin receptor (p75NTR), increasing liver damage. Additionally, blocking the NGF decreased liver fibrosis whereas treatment with recombinant NGF accelerated the fibrotic process to a similar extent than stress challenge. We further show that the fibrogenesis induced by stress is characterized by specific changes in the hepatoglycocode (increased β1,6GlcNAc-branched complex N-glycans and decreased core 1 O-glycans expression) which are also observed in patients with advanced fibrosis compared to patients with a low level of fibrosis. Our study facilitates an understanding of stress-induced liver injury and identify NGF signaling pathway in early stages of the disease, which contributes to the established fibrogenesis.

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