Pulmonary Fibrosis Model Research Articles

Intermittent hypoxia exacerbated depressive and anxiety-like behaviors in the bleomycin-induced pulmonary fibrosis mice

Depression and anxiety are prevalent in patients with idiopathic pulmonary fibrosis (IPF). Recent researchers reveal that intermittent hypoxia (IH) increases the severity of bleomycin (BLM)-induced lung injury. However, experimental studies dealing with anxiety- and depression-like behavior in animal models of BLM-induced pulmonary fibrosis in a combination of IH are lacking, hence, this study aimed to investigate that.In this study, 80 C57BL/6J male mice were intratracheally injected with BLM or normal saline at day0 and then exposed to IH (alternating cycles of FiO2 21 % for 60 s and FiO2 10 % for 30 s, 40 cycles/hour, 8 h/day) or intermittent air (IA) for 21 days. Behavioral tests, including open field test (OFT), sucrose preference test (SPT) and tail suspension test (TST), were detected from day22 to day26. This study found that pulmonary fibrosis developed and lung inflammation were activated in BLM-induced mice, which were potentiated by IH. Significant less time in center and less frequency of entries in the centre arena in OFT were observed in BLM treated mice, and IH exposure further decreased that. Marked decreased percent of sucrose preference in SPT, and significant increased immobility time of the TST were detected in BLM treated mice and IH widen the gaps. The expression of ionized calcium-binding adaptor molecule (Iba1) was activated in the hippocampus of BLM instillation mice and IH enlarged it. Moreover, a positive correlation between hippocampal microglia activation and inflammatory factors was observed. Our results demonstrated that IH exacerbated depressive and anxiety-like behaviors in the BLM-induced pulmonary fibrosis mice. The changes in pulmonary inflammation-hippocampal microglia activation may be a potential mechanism in this phenomenon, which can be researched in future.

Meta-analysis of Ac-SDKP inhibition of Pulmonary fibrosis in animal models

Objective: To systematically study the anti-fibrotic effect of N-acetyl-seryl-as partyl-lysyl-proline (Ac-SDKP) on pulmonary fibrosis. Methods: In May 2021, a computer search was performed on CNKI, Wanfang Knowledge Service Platform, VIP.com, China Biomedical Literature Database, Pubmed, OVID and other databases. The retrieval time was from January 2008 to May 2021. Randomized controlled experiments on the inhibition of pulmonary fibrosis by Ac-SDKP were screened. The control group was the pulmonary fibrosis model group and the experimental group was the Ac-SDKP treatment group. The quality of the literature was assessed using the syrcle risk of bias assessment tool, and data were extracted. Data analysis was Performed using revman 5.4 software. Results: 18 papers were included, with a total of 428 animal models. The results of meta analysis showed that the contents of α-smooth muscle actin (α-SMA), type I collagen, type Ⅲ collagen, transforming growth factor-β (TGF-β) and Nodule area in the exPerimental group were lower than those in the control grouP. [SMD=-2.44, 95%CI (-3.71--1.17), P=0.000][SMD=-5.36, 95%CI (-7.13--3.59), P=0.000] [SMD=-3.07, 95%CI (-4.13--2.02), P<0.000][SMD=-2.88, 95%CI (-3.63--2.14), P=0.000] [SMD=-1.80, 95%CI (-2.42--1.18), P=0.000], the content of hydroxy proline in the experimental group was higher than that in the control group [SMD=7.62, 95%CI (4.90-10.33), P=0.000], all indexes included in the literature were statistically significant. Conclusion: Ac-SDKP has obvious inhibitory effect on the process of pulmonary fibrosis, and may become a new clinical drug for the treatment of pulmonary fibrosis.

Identification and validation of chemokine system-related genes in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive interstitial lung disease with limited therapeutic options. Recent studies have demonstrated that chemokines play a vital role in IPF pathogenesis. In the present study, we explored whether the gene signature associated with chemokines could be used as a reliable biological marker for patients with IPF. Chemokine-related differentially expressed genes (CR-DEGs) in IPF and control lung tissue samples were identified using data from the Gene Expression Omnibus database. A chemokine-related signature of the diagnostic model was established using the LASSO-Cox regression. In addition, unsupervised cluster analysis was conducted using consensus-clustering algorithms. The CIBERSORT algorithm was used to calculate immune cell infiltration across patient subgroups. Finally, we established a mouse model of bleomycin-induced pulmonary fibrosis and a model of fibroblasts treated with TGFβ1. Expression levels of chemokine-related signature genes were determined using real-time quantitative polymerase chain reaction (RT-qPCR). We established a chemokine-related eleven-gene signature of a diagnostic model consisting of CXCL2, CCRL2, ARRB1, XCL1, GRK5, PPBP, CCL19, CCL13, CCL11, CXCL6, and CXCL13, which could easily distinguish between IPF patients and controls. Additionally, we identified two subtypes of IPF samples based on chemokine-related gene expression. Pulmonary function parameters and stromal scores were significantly higher in subtype 1 than in subtype 2. Several immune cell types, especially plasma cells and macrophages, differ significantly between the two subtypes. RT-qPCR results showed that the expression levels of Cxcl2 and Ccl2 increased considerably in bleomycin-induced mice. Meanwhile, Arrb1, Ccrl2, Grk5, and Ppbp expression was significantly reduced. Furthermore, multiple chemokine-related genes were altered in TGFβ1 or TNFα-induced fibroblast cells. A novel chemokine-related eleven-signature of diagnostic model was developed. These genes are potential biomarkers of IPF and may play essential roles in its pathogenesis.

Comparative Aspects of Ricin Toxicity by Inhalation.

The pathogenesis of ricin toxicity following inhalation has been investigated in many animal models, including the non-human primate (predominantly the rhesus macaque), pig, rabbit and rodent. The toxicity and associated pathology described in animal models are broadly similar, but variation appears to exist. This paper reviews the published literature and some of our own unpublished data and describes some of the possible reasons for this variation. Methodological variation is evident, including method of exposure, breathing parameters during exposure, aerosol characteristics, sampling protocols, ricin cultivar, purity and challenge dose and study duration. The model species and strain used represent other significant sources of variation, including differences in macro- and microscopic anatomy, cell biology and function, and immunology. Chronic pathology of ricin toxicity by inhalation, associated with sublethal challenge or lethal challenge and treatment with medical countermeasures, has received less attention in the literature. Fibrosis may follow acute lung injury in survivors. There are advantages and disadvantages to the different models of pulmonary fibrosis. To understand their potential clinical significance, these factors need to be considered when choosing a model for chronic ricin toxicity by inhalation, including species and strain susceptibility to fibrosis, time it takes for fibrosis to develop, the nature of the fibrosis (e.g., self-limiting, progressive, persistent or resolving) and ensuring that the analysis truly represents fibrosis. Understanding the variables and comparative aspects of acute and chronic ricin toxicity by inhalation is important to enable meaningful comparison of results from different studies, and for the investigation of medical countermeasures.

RETRACTED ARTICLE: Qilongtian ameliorate bleomycin-induced pulmonary fibrosis in mice via inhibiting IL-17 signal pathway

Pulmonary fibrosis (PF) is a special type of pulmonary parenchymal disease, with chronic, progressive, fibrosis, and high mortality. There is a lack of safe, effective, and affordable treatment methods. Qilongtian (QLT) is a traditional Chinese prescription that is composed of Panax notoginseng, Earthworm, and Rhodiola, and shows the remarkable clinical curative effect of PF. However, the mechanism of QLT remains to be clarified. Therefore, we studied the effectivity of QLT in treating Bleomycin (BLM) induced PF mice. 36 C57BL/6 J mice were randomized into the control group, the model group, the low-, medium- and high-dose QLT group, and Pirfenidone group. After establishing a model of pulmonary fibrosis in mice, the control and model groups were infused with a normal saline solution, and the delivery group was infused with QLT. Pulmonary function in the mice from each group was detected. Pulmonary tissue morphologies and collagen deposition were stained by HE and Masson. The content of hydroxyproline (HYP) was detected by alkaline hydrolysis and the mRNA and protein expression of related genes in pulmonary tissues were detected by using q-PCR, ELISA, and Western blot. Our studies have shown that QLT significantly reduced the inflammatory injury, hydroxy-proline content, and collagen deposition of pulmonary tissue in BLM-induced PF mice and down-regulated the cytokine related to inflammation and fibrosis and PF expression on the mRNA and protein level in PF mice. To identify the mechanism of QLT, the Transcriptome was measured and the IL-17 signal pathway was screened out for further research. Further studies indicated that QLT reduced the mRNAs and protein levels of interleukin 17 (IL-17), c–c motif chemokine ligand 12 (CCL12), c-x-c motif chemokine ligand 5 (CXCL5), fos-like antigen 1 (FOSL1), matrix metalloproteinase-9 (MMP9), and amphiregulin (AREG), which are inflammation and fibrosis-related genes in the IL-17 signal pathway. The results indicated that the potential mechanism for QLT in the prevention of PF progression was by inhibiting inflammation resulting in the IL-17 signal pathway. Our study provides the novel scientific basis of QLT and represents new therapeutics for PF in clinical.

The mRNA-binding protein DDX3 mediates TGF-β1 upregulation of translation and promotes pulmonary fibrosis.

Pulmonary fibrosis is potentiated by a positive feedback loop involving the extracellular sialidase enzyme neuraminidase 3 (NEU3) causing release of active TGF-β1 and TGF-β1 upregulating NEU3 by increasing translation without affecting mRNA levels. In this report, we elucidate the TGF-β1 upregulation of the translation mechanism. In human lung fibroblasts, TGF-β1 increased levels of proteins, including NEU3, by increasing translation of the encoding mRNAs without significantly affecting levels of these mRNAs. A total of 180 of these mRNAs shared a common 20-nucleotide motif. Deletion of this motif from NEU3 mRNA eliminated the TGF-β1 upregulation of NEU3 translation, while insertion of this motif in 2 mRNAs insensitive to TGF-β1 caused TGF-β1 to upregulate their translation. RNA-binding proteins including DEAD box helicase 3, X-linked (DDX3), bind the RNA motif, and TGF-β1 regulates their protein levels and/or binding to the motif. We found that DDX3 was upregulated in the fibrotic lesions in patients with pulmonary fibrosis, and inhibiting DDX3 in fibroblasts reduced TGF-β1 upregulation of NEU3 levels. In the mouse bleomycin model of pulmonary fibrosis, injections of the DDX3 inhibitor RK-33 potentiated survival and reduced lung inflammation, fibrosis, and tissue levels of DDX3, TGF-β1, and NEU3. These results suggest that inhibiting an mRNA-binding protein that mediates TGF-β1 upregulation of translation can reduce pulmonary fibrosis.

An Integrative Multiomics Framework for Identification of Therapeutic Targets in Pulmonary Fibrosis.

Pulmonary fibrosis (PF) is a heterogeneous disease with a poor prognosis. Therefore, identifying additional therapeutic modalities is required to improve outcome. However, the lack of biomarkers of disease progression hampers the preclinical to clinical translational process. Here, this work assesses and identifies progressive alterations in pulmonary function, transcriptomics, and metabolomics in the mouse lung at 7, 14, 21, and 28 days after a single dose of oropharyngeal bleomycin. By integrating multi-omics data, this work identifies two central gene subnetworks associated with multiple critical pathological changes in transcriptomics and metabolomics as well as pulmonary function. This work presents a multi-omics-based framework to establish a translational link between the bleomycin-induced PF model in mice and human idiopathic pulmonary fibrosis to identify druggable targets and test therapeutic candidates. This work also indicates peripheralcannabinoid receptor 1 (CB1 R) antagonism as a rational therapeutic target for clinical translation in PF. Mouse Lung Fibrosis Atlas can be accessed freely at https://niaaa.nih.gov/mouselungfibrosisatlas.

RNF130 protects against pulmonary fibrosis through suppressing aerobic glycolysis by mediating c-myc ubiquitination.

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal disease,characterized by an excessive accumulation of extracellular matrix (ECM) proteins in response to chronic lung injury. Current evidence suggests that metabolic reprogramming is always accompanied by myofibroblast activation in IPFof whichthe underlying mechanisms remain unclear. Ring finger protein 130 (RNF130), was demonstrated involved in multiple diseases. However, whether RNF130 plays a critical role in the pathogenesis of IPF needs to be clarified. We first investigated the expression of RNF130 in pulmonary fibrosis in vivo and in vitro. We then observed the effect and explored the molecular mechanism of RNF130 on the transition of fibroblast to myofibroblast and aerobic glycolysis. Further, we assessed the effects of adeno-associated virus (AAV)-induced RNF130 overexpression in the pulmonary fibrosis model, conducting pulmonary function, assessment of collagen depositionusing the hydroxyproline assay, and biochemical and histopathological analyses. We found that RNF130 was down-regulated in lung tissues of mice with bleomycin-induced pulmonary fibrosis and lung fibroblasts treated with transforming growth factor-β1 (TGF-β1). Then we demonstrated that RNF130 inhibitedthe transition of fibroblast to myofibroblast by suppressing aerobic glycolysis. Mechanistically, we revealed that RNF130 promotedc-myc ubiquitination and degradation, while c-myc overexpression reverses the inhibitory effects of RNF130. Importantly, pulmonary function, collagen deposition and fibroblast differentiation were significantly alleviated in adeno-associated virus serotype (AAV)6-RNF130 treated mice, which further validated the contribution of RNF130/c-myc signaling axis in pulmonary fibrosis pathological process. In summary, RNF130 participates in the pathogenesis of pulmonary fibrosis by inhibiting the transition of fibroblast to myofibroblast and aerobic glycolysis through promoting c-myc ubiquitination and degradation. Targeting RNF130-c-myc axismightrepresent a promising strategy to alleviate the progression of IPF.

Synergistic effect of constituent drugs of Baibutang on improving Yin-deficiency pulmonary fibrosis in rats.

Baibutang (BBT) is an ancient prescription for the treatment of pulmonary fibrosis. Previous experiments have shown that BBT had a good therapeutic effect on pulmonary fibrosis. However, there had been no study on the synergy between drugs composed of BBT. Due to the interaction between the constituent drugs, exploring their synergy profile is of great significance for explaining the essence of BBT's efficacy in improving pulmonary fibrosis. Based on the pharmacodynamic value, this study aimed to explore a method for the evaluation of the synergy profile between constituent drugs in traditional Chinese medicine (TCM) compounds. Nine herbs of BBT were divided into Zhikeqingre (ZK), Yangyinyiqi (YY) and Lishijianpi (LS) groups. A rat model of Yin-deficiency pulmonary fibrosis induced by thyroxine-bleomycin was used to evaluate the effects of BBT and the three groups. The pathological changes of lung tissue and the changes of biomarkers associated with fibrosis, Yin-deficiency and water-fluid metabolism were detected. After standardization of pharmacodynamics value (PV), the compatibility coefficient (CC) of the three groups, the relative PV (RPV) and contribution value (CV) of each group on every index were calculated. The average CC on fibrosis indexes was 0.44, indicating that 44% of the efficacy of BBT came from the synergistic effect of the three groups. ZK group had the highest RPV (0.80) in improving fibrosis indexes such as histopathological changes, α-SMA, collagen-I and renin-angiotensin system. The average CC on Yin-deficiency indexes was 0.25, and YY group had the highest RPV (0.96) in improving deficiency indexes such as body temperature, cAMP/cGMP ratio, and PDEs, PGE2 and COX-2 levels. The average CC on water-fluid metabolism indexes was 0.15, and LS group had the highest RPV (1.52) in improving water-fluid metabolism indexes such as aquaporins, mucins, and surfactant proteins. The results also showed that 29% of the improvement effect of BBT on all indexes came from the synergistic effect of the three groups, and the contribution of ZK, YY and LS groups to the efficacy of BBT were 25%, 25% and 21%, respectively. The established semiquantitative method can clearly and simply evaluate the synergy of the three groups in BBT, which will help to promote the research on the synergy of TCM compounds and other multiple-components combinations.

Repurposing digoxin for geroprotection in patients with frailty and multimorbidity.

The geroscience hypothesis proposes biological hallmarks of ageing are modifiable. Increasing evidence supports targeting these hallmarks with therapeutics could prevent and ameliorate age-related conditions - collectively termed "geroprotector drugs". Cellular senescence is a hallmark with considerable potential to be modified with geroprotector drugs. Senotherapeutics are drugs that target cellular senescence for therapeutic benefit. Repurposing commonly used medications with secondary geroprotector properties is a strategy of interest to promote incorporation of geroprotector drugs into clinical practice. One candidate is the cardiac glycoside digoxin. Evidence in mouse models of pulmonary fibrosis, Alzheimer's disease, arthritis and atherosclerosis support digoxin as a senotherapeutic agent. Proposed senolytic mechanisms are upregulation of intrinsic apoptotic pathways and promoting intracellular acidification. Digoxin also appears to have a senomorphic mechanism - altering the T cell pool to ameliorate pro-inflammatory SASP. Despite being widely prescribed to treat atrial fibrillation and heart failure, often in multimorbid older adults, it is not known whether digoxin exerts senotherapeutic effects in humans. Further cellular and animal studies, and ultimately clinical trials with participation of pre-frail older adults, are required to identify whether digoxin has senotherapeutic effect at low dose. This paper reviews the biological mechanisms identified in preliminary cellular and animal studies that support repurposing digoxin as a geroprotector in patients with frailty and multimorbidity.

Hsa-miR-379 down-regulates Rac1/MLK3/JNK/AP-1/Collagen I cascade reaction by targeting connective tissue growth factor in human alveolar basal epithelial A549 cells

ObjectiveThis study was aimed to screen and identify miRNAs that could regulate human CTGF gene and downstream cascade reaction Rac1/MLK3/JNK/AP-1/Collagen I by bioinformatics and experimental means. MethodsTargetScan and Tarbase were used to predict miRNAs that may have regulatory effects on human CTGF gene. The dual-luciferase reporter gene assay was employed to verify the results obtained in bioinformatics. Human alveolar basal epithelial A549 cells were exposed to silica (SiO2) culture medium for 24 h to establish an in vitro model of pulmonary fibrosis, and bleomycin (BLM) of 100 ng/mL was used as a positive control. The miRNA and mRNA expression levels were determined by RT-qPCR, and the protein levels were measured by western blot in hsa-miR-379-3p overexpression group or not. ResultsA total of 9 differentially expressed miRNAs that might regulate the human CTGF gene were predicted. Hsa-miR-379-3p and hsa-miR-411-3p were selected for the subsequent experiments. The results of the dual-luciferase reporter assay showed that hsa-miR-379-3p could bind to CTGF, but hsa-miR-411-3p could not. Compared with the control group, SiO2 exposure (25 and 50 μg/mL) could significantly reduce the expression level of hsa-miR-379-3p in A549 cells. SiO2 exposure (50 μg/mL) could significantly increase the mRNA expression levels of CTGF, Collagen I, Rac1, MLK3, JNK, AP1, and VIM in A549 cells, while CDH1 level was significantly decreased. Compared with SiO2 + NC group, the mRNA expression levels of CTGF, Collagen I, Rac1, MLK3, JNK, AP1, and VIM were significantly decreased, and CDH1 level was significantly higher when hsa-miR-379-3p was overexpressed. At the same time, overexpression of hsa-miR-379-3p improved the protein levels of CTGF, Collagen I, c-Jun and phospho-c-Jun, JNK1 and phospho-JNK1 significantly compared with SiO2 + NC group. ConclusionHsa-miR-379-3p was demonstrated for the first time that could directly target and down-regulate human CTGF gene, and further affect the expression levels of key genes and proteins in Rac1/MLK3/JNK/AP-1/Collagen I cascade reaction.

Protective effects of Qing-Re-Huo-Xue formula on bleomycin-induced pulmonary fibrosis through the p53/IGFBP3 pathway

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic, progressive fibrosing lung disease with high mortality. Inflammation and epithelial mesenchymal transformation (EMT) may play an important role in the occurrence and development of IPF. Qing-Re-Huo-Xue formula (QRHXF) has been used clinically by our team for half a century and has obvious therapeutic effects on lung disease. Nevertheless, the role and mechanism of QRHXF in the treatment of IPF have never been studied.MethodsA mouse pulmonary fibrosis model was established by intratracheal injection of BLM. The effects of QRHXF on the treatment of pulmonary fibrosis were studied by pulmonary function testing, imaging examination, pathological staining, transmission electron microscopy (TEM) observation and mRNA expression. Tandem mass tag (TMT)-based quantitative proteomics was carried out to analyse the lung protein expression profiles between the control (CTL), bleomycin (BLM) and QRHXF (BLM + QRHXF) groups. Immunohistochemistry and qRT-PCR were used to verify the possible existence of drug target proteins and signalling pathways.ResultsThe results of pulmonary function, lung pathology and imaging examinations showed that QRHXF could significantly alleviate BLM-induced pulmonary fibrosis in vivo. Additionally, inflammatory cell infiltration and EMT were markedly reduced in BLM-induced PF mice administered QRHXF. Proteomics detected a total of 35 proteins, of which 17 were upregulated and 18 were downregulated. A total of 19 differentially expressed proteins (DEPs) overlapped between the BLM versus CTL groups and the BLM + QRHXF versus BLM groups. The expression of p53 and IGFBP3 was reversed in the QRHXF intervention group, which was verified by immunohistochemistry and qRT-PCR.ConclusionsQRHXF attenuated BLM-induced pulmonary fibrosis, and regulation of the p53/IGFBP3 pathway might be associated with its efficacy, which holds promise as a novel treatment strategy for pulmonary fibrosis patients.Graphical

Synthesis, hemorheological and antifibrotic activity of newly synthesized 3-acetyl-2,4,6-trimethylpyridine derivatives

Pyridine derivatives still attract the attention of many researchers for their comprehensive biological research, possible promotion to the pharmaceutical market in order to be used as effective drugs. In this article, based on the well-known and fairly accessible in terms of laboratory synthesis of the 3-acetyl derivative of collidine, we carried out several simple chemical modifications to obtain new collidine derivatives not previously described. The structure of the synthesized compounds was confirmed by 1H- and 13C NMR spectroscopy. A biological screening of the synthesized compounds for hemorheological activity was carried out on an in vitro blood hyperviscosity model. It was shown that pronounced hemorheological activity was established for compounds 3, 4 and 5. Compound 4 also showed a pronounced antifibrotic effect in an experimental model of pulmonary fibrosis induced by intratracheal administration of bleomycin to rats. In an experimental model of pulmonary fibrosis in rats, the formation of a hyperviscosity syndrome was also observed, and blood viscosity decreased when compound 4 was administered to animals.

Evaluating the Effect of Hydrogen Sulfide in the Idiopathic Pulmonary Fibrosis Model with a Fluorescent Probe.

Hydrogen sulfide (H2S), a gaseous signaling molecule, is involved in a wide range of physiological and pathological processes. H2S has been proven to play a beneficial role in lung diseases, and the relationship between perturbations in endogenous H2S synthesis and degree with idiopathic pulmonary fibrosis (IPF) has attacted increasing attention. However, the changes in endogenous lung H2S levels in the pathological progression of chronic pulmonary diseases remain unclear. To this end, we synthesized a fluorescent probe (Bcy-HS) for the selective imaging of H2S in living cells and mice. This probe was mainly used for in situ in vivo and cellular imaging as well as a systematic assessment of intrapulmonary H2S levels at different stages of IPF. In addition, we also discussed the potential of H2S supplementation in the treatment of pulmonary fibrotic diseases. Our results confirmed the key role of H2S in pulmonary fibrosis. In cellular and mice models of pulmonary fibrosis, intracellular H2S levels are reduced. However, the severity of oxidative damage and pulmonary fibrosis decreased after NaSH (H2S donor). Therefore, we concluded that increasing the H2S content in vivo may be a novel strategy for IPF treatment.

Pulmonary fibrosis model of mice induced by different administration methods of bleomycin

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic, progressive disease of the lung. How to build a typical human mimicking animal model has been a challenge. Thus, to reveal the mechanism and to make it useful for IPF clinical treatment, a different type of mice model and inspection methods are used to evaluate which one is applicable for the study of IPF.Method69 Twelve-weeks-old C57BL/6 mice were divided into 3 type groups (n = 7 for each control group, n = 8 for each BLM-induced pulmonary fibrosis groups), as intraperitoneal injection, intratracheal administration, and intravenous administration of bleomycin (BLM) to initiate lung fibrosis. Changes of the lung function measured through mice Pulmonary function test (PFT). Morphological changes in mice were observed by PET/CT, Masson and Picro-Sirius staining, Transmission electron microscopy (TEM). Biochemical changes were tested by Enzyme-linked immunosorbent assay (Elisa).ResultsPET/CT of BLM-receiving mice showed an increase in fibrotic consolidations and an increase in non-aerated lung area in BLM-treated mice compared with that in controls. TGF-b1, TNF-a, IL-6, GM-CSF in BALF and serum. PAI-1, HYP in the lung tissue of mice were significantly different in each BLM groups than those in the controls. The results of Masson staining in mice indicate that the lung tissues of all BLM received groups, the intratracheal groups, the intravenous groups, and the intraperitoneal groups have a higher degree of pulmonary septal thickening and collagen fiber consolidation compare to saline control. Picro-Sirius staining results are consistent with the results of Masson staining. Compared with the saline control group, the ratio of Col 1/Col 3 was significantly increased in each BLM group. TEM results found that in BLM group, type I alveolar epithelial cells were degenerated. Exfoliated endothelial cells were swelling, and type II alveolar epithelial cells were proliferated, the shape of the nucleus was irregular, and some tooth-like protrusions were seen.ConclusionsWith three different methods of animal model construction, high dose of each show more compliable, and BLM can successfully induce animal models of pulmonary fibrosis, however, certain differences in the fibrosis formation sites of them three, and tail vein injection of BLM induced PF model is closer to the idiopathic pulmonary interstitial fibrosis.

Application and Study of ROCK Inhibitors in Pulmonary Fibrosis: Recent Developments and Future Perspectives.

Rho-associated coiled-coil-containing kinases (ROCKs), serine/threonine protein kinases, were initially identified as downstream targets of the small GTP-binding protein Rho. Pulmonary fibrosis (PF) is a lethal disease with limited therapeutic options and a particularly poor prognosis. Interestingly, ROCK activation has been demonstrated in PF patients and in animal PF models, making it a promising target for PF treatment. Many ROCK inhibitors have been discovered, and four of these have been approved for clinical use; however, no ROCK inhibitors are approved for the treatment of PF patients. In this article, we describe ROCK signaling pathways and the structure-activity relationship, potency, selectivity, binding modes, pharmacokinetics (PKs), biological functions, and recently reported inhibitors of ROCKs in the context of PF. We will also focus our attention on the challenges to be addressed when targeting ROCKs and discuss the strategy of ROCK inhibitor use in the treatment of PF.

High-Fat Diet Related Lung Fibrosis-Epigenetic Regulation Matters

Pulmonary fibrosis (PF) is an interstitial lung disease characterized by the destruction of the pulmonary parenchyma caused by excessive extracellular matrix deposition. Despite the well-known etiological factors such as senescence, aberrant epithelial cell and fibroblast activation, and chronic inflammation, PF has recently been recognized as a metabolic disease and abnormal lipid signature was observed both in serum and bronchoalveolar lavage fluid (BALF) of PF patients and mice PF model. Clinically, observational studies suggest a significant link between high-fat diet (HFD) and PF as manifested by high intake of saturated fatty acids (SFAs) and meat increases the risk of PF and mice lung fibrosis. However, the possible mechanisms between HFD and PF remain unclear. In the current review we emphasize the diversity effects of the epigenetic dysregulation induced by HFD on the fibrotic factors such as epithelial cell injury, abnormal fibroblast activation and chronic inflammation. Finally, we discuss the potential ways for patients to improve their conditions and emphasize the prospect of targeted therapy based on epigenetic regulation for scientific researchers or drug developers.

LncRNA GAS5 suppresses TGF-β1-induced transformation of pulmonary pericytes into myofibroblasts by recruiting KDM5B and promoting H3K4me2/3 demethylation of the PDGFRα/β promoter

BackgroundIdiopathic pulmonary fibrosis (IPF) is a condition that may cause persistent pulmonary damage. The transformation of pericytes into myofibroblasts has been recognized as a key player during IPF progression. This study aimed to investigate the functions of lncRNA growth arrest-specific transcript 5 (GAS5) in myofibroblast transformation during IPF progression.MethodsWe created a mouse model of pulmonary fibrosis (PF) via intratracheal administration of bleomycin. Pericytes were challenged with exogenous transforming growth factor-β1 (TGF-β1). To determine the expression of target molecules, we employed quantitative reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemical and immunofluorescence staining. The pathological changes in the lungs were evaluated via H&E and Masson staining. Furthermore, the subcellular distribution of GAS5 was examined using FISH. Dual-luciferase reporter assay, ChIP, RNA pull-down, and RIP experiments were conducted to determine the molecular interaction.ResultsGAS5 expression decreased whereas PDGFRα/β expression increased in the lungs of IPF patients and mice with bleomycin-induced PF. The in vitro overexpression of GAS5 or silencing of PDGFRα/β inhibited the TGF-β1-induced differentiation of pericytes to myofibroblasts, as evidenced by the upregulation of pericyte markers NG2 and desmin as well as downregulation of myofibroblast markers α-SMA and collagen I. Further mechanistic analysis revealed that GAS5 recruited KDM5B to promote H3K4me2/3 demethylation, thereby suppressing PDGFRα/β expression. In addition, KDM5B overexpression inhibited pericyte–myofibroblast transformation and counteracted the promotional effect of GAS5 knockdown on pericyte–myofibroblast transformation. Lung fibrosis in mice was attenuated by GAS5 overexpression but promoted by GAS5 deficiency.ConclusionGAS5 represses pericyte–myofibroblast transformation by inhibiting PDGFRα/β expression via KDM5B-mediated H3K4me2/3 demethylation in IPF, identifying GAS5 as an intervention target for IPF.

Arctiin-encapsulated DSPE-PEG bubble-like nanoparticles inhibit alveolar epithelial type 2 cell senescence to alleviate pulmonary fibrosis via the p38/p53/p21 pathway.

Background: Idiopathic pulmonary fibrosis is a severe and deadly form of diffuse parenchymal lung disease and treatment options are few. Alveolar epithelial type 2 (AEC2) cell senescence is implicated in the pathogenies of IPF. A major bioactive compound from the traditional Chinese medicine Fructus arctii, arctiin (ARC) has robust anti-inflammatory, anti-senescence, and anti-fibrosis functions. However, the potential therapeutic effects of ARC on IPF and the underlying mechanisms involved are still unknown. Methods: First of all, ARC was identified as an active ingredient by network pharmacology analysis and enrichment analysis of F. arctii in treating IPF. We developed ARC-encapsulated DSPE-PEG bubble-like nanoparticles (ARC@DPBNPs) to increase ARC hydrophilicity and achieve high pulmonary delivery efficiency. C57BL/6 mice were used to establish a bleomycin (BLM)-induced pulmonary fibrosis model for assessing the treatment effect of ARC@DPBNPs on lung fibrosis and the anti-senescence properties of AEC2. Meanwhile, p38/p53 signaling in AEC2 was detected in IPF lungs, BLM-induced mice, and an A549 senescence model. The effects of ARC@DPBNPs on p38/p53/p21 were assessed in vivo and in vitro. Results: Pulmonary route of administration of ARC@DPBNPs protected mice against BLM-induced pulmonary fibrosis without causing significant damage to the heart, liver, spleen, or kidney. ARC@DPBNPs blocked BLM-induced AEC2 senescence in vivo and in vitro. The p38/p53/p21 signaling axis was significantly activated in the lung tissues of patients with IPF, senescent AEC2, and BLM-induced lung fibrosis. ARC@DPBNPs attenuated AEC2 senescence and pulmonary fibrosis by inhibiting the p38/p53/p21 pathway. Conclusion: Our data suggest that the p38/p53/p21 signaling axis plays a pivotal role in AEC2 senescence in pulmonary fibrosis. The p38/p53/p21 signaling axis inhibition by ARC@DPBNPs provides an innovative approach to treating pulmonary fibrosis in clinical settings.

DAB2 promotes pulmonary fibrosis and may act as an intermediate between IGF‑1R and PI3K/AKT signaling pathways.

Idiopathic pulmonary fibrosis (IPF) is a heterogeneous lung disease associated with high mortality. Disabled-2 (DAB2), an adapter protein, regulates cell-fibrinogen adhesion and fibrinogen uptake. DAB2 is differentially expressed in mouse fibrotic lungs induced by bleomycin according to a genome microarray analysis based on Gene Expression Omnibus database. However, the role of DAB2 in IPF has not been revealed. A bleomycin-induced mouse model of pulmonary fibrosis was constructed in the present study. It found that the expression of DAB2 was upregulated in bleomycin-induced fibrotic lung tissue with collagen fiber deposition and pulmonary interstitium thickening. Colocalization of DAB2 with α-smooth muscle actin (SMA) was observed in lung tissue sections. In vitro, human lung fibroblast MRC-5 cells were treated with TGF-β1 and the expression of DAB2 was increased. Knockdown of DAB2 suppressed cell proliferation and the expression of α-SMA, collagen I, collagen IV and fibronectin in TGF-β1-treated MRC-5 cells. The phosphorylation levels of PI3K and AKT were suppressed in DAB2-knockdown cells. IGF-1/IGF-1R has been reported to promote pulmonary fibrosis and activate the PI3K/Akt signaling. In the present study, the activation of IGF-1/IGF-1R signaling pathways in bleomycin-induced fibrotic lung tissues were positively associated with DAB2 expression. The phosphorylation level of IGF-1R was increased in MRC-5 cells with TGF-β1 treatment, and DAB2 expression was decreased by silencing of IGF-1R. This suggested that DAB2 might be a downstream target of the IGF-1R pathway and thus induced PI3K/AKT signaling activation and fibrogenesis. The current study demonstrated the importance of DAB2 in pulmonary fibrosis and suggested the potential of IGF-1R/DAB2/PI3K in the pathogenesis of IPF.