Idiopathic Pulmonary Fibrosis Mice Research Articles

Structural optimization of Moracin M as novel selective phosphodiesterase 4 inhibitors for the treatment of idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and high mortality lung disease. Although the antifibrotic drugs pirfenidone and nintedanib could slow the rate of lung function decline, the usual course of the condition is inexorably to respiratory failure and death. Therefore, new approaches and novel therapeutic drugs for the treatment of IPF are urgently needed. And the selective PDE4 inhibitor has in vivo and in vitro anti-fibrotic effects in IPF models. But the clinical application of most PDE4 inhibitors are limited by their unexpected and severe side effects such as nausea, vomiting, and diarrhea. Herein, structure-based optimizations of the natural product Moracin M resulted in a novel a novel series of 2-arylbenzofurans as potent PDE4 inhibitors. The most potent inhibitor L13 has an IC50 of 36 ± 7 nM with remarkable selectivity across the PDE families and administration of L13·citrate (10.0 mg/kg) exhibited comparable anti-pulmonary fibrosis effects to pirfenidone (300 mg/kg) in a bleomycin-induced IPF mice model, indicate that L13 is a potential lead for the treatment of IPF.

Involvement of E3 ubiquitin ligase NEDD4-mediated YY1 ubiquitination in alleviating idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic and lethal lung disease characterized by progressive lung scarring. This study aims to elucidate the role of the E3 ubiquitin ligase NEDD4 in the ubiquitination of YY1 and its subsequent impact on TAB1 transcription, revealing a possible molecular mechanism in the development of IPF. Through bioinformatics analysis and both in vitro and in vivo experiments, we observed differential expression levels of NEDD4 and YY1 between normal and IPF samples, identifying NEDD4 as an upstream E3 ubiquitin ligase of YY1. Furthermore, binding sites for the transcription factor YY1 on the promoter region of TAB1 were discovered, indicating a direct interaction. In vitro experiments using HEPF cells showed that NEDD4 mediates the ubiquitination and degradation of YY1, leading to suppressed TAB1 transcription, thereby inhibiting cell proliferation and fibrogenesis. These findings were corroborated by in vivo experiments in an IPF mouse model, where the ubiquitination pathway facilitated by NEDD4 attenuated IPF progression through the downregulation of YY1 and TAB1 transcription. These results suggest that NEDD4 plays a crucial role in the development of IPF by modulating YY1 ubiquitination and TAB1 transcription, providing new insights into potential therapeutic targets for treating IPF.

Protection of Qingfei Xieding prescription from idiopathic pulmonary fibrosis by regulating renin-angiotensin and ferroptosis in MLE-12 cells.

Idiopathic pulmonary fibrosis (IPF) is a lifelong lung disease, but there is no specific drug for treatment. Qingfei Xieding prescription (QF) is active in the treatment of lung diseases. More comprehensive mechanisms over how QF exhibits anti-pulmonary fibrosis need to be elucidated. TGF-β was used to construct a pulmonary fibrosis cell model in vitro. Bleomycin was applied to induce a lung tissue fibrosis model in mice in vivo. Flow cytometry was used to detect cellular ROS and lipid oxidation levels. Cell substructure was observed by Transmission Electron Microscopy. ELISA was used to determine the levels of inflammatory factors. HE staining, Masson staining and immunohistochemistry were performed to evaluate the degree of fibrosis. Western Blot assay was used to determine the protein expressions of different molecules. In TGF-β-exposed lung epithelial MLE-12 cell model, α-SMA and Collagen I were significantly elevated and cell viability was reduced. QF treatment restored the cell viability decreased by exogenous TGF-β. Ferroptosis inducer Erastin administration could reverse the beneficial effects such as lipid oxidation and ROS reduction caused by QF treatment. QF was proven to inhibit ferroptosis and alleviated the process of IPF by activating ACE2 signal axis. In bleomycin induced IPF mice model, QF altered lung coefficient, body weight and the expression of inflammatory factors, which were prevented by ferroptosis activator Erastin. QF was demonstrated to affect the ACE2-ERK signaling axis in vivo. QF alleviated idiopathic pulmonary fibrosis by regulating renin-angiotensin through blocking ferroptosis. This research offers evidence for the potentiality of QF in clinical application for IPF therapy.

Co-Delivery of Astaxanthin and siTGF-β1 via Ionizable Liposome Nanoparticles for Improved Idiopathic Pulmonary Fibrosis Therapy.

Alleviating the injury of type II alveolar epithelial cells (AEC 2s) and inhibiting the activation and differentiation of fibroblasts are significant for improving the therapeutic effect of idiopathic pulmonary fibrosis (IPF). To this aim, ionizable liposome nanoparticles (ASNPs) coloaded with antioxidant drug astaxanthin (AST) and small interfering RNA targeting transforming growth factor β1 (siTGF-β1) were developed for enhanced IPF therapy. ASNPs showed high loading and intracellular delivery efficiency for AST and siTGF-β1. After the injection of ASNPs in an IPF mice model, the loaded AST largely scavenged reactive oxygen species (ROS) in the diseased lung to reduce AEC2 apoptosis, thereby ensuring the integrity of the alveolar epithelium. Meanwhile, siTGF-β1, delivered by ASNPs, significantly silenced the expression of TGF-β1 in fibroblasts, inhibiting the differentiation of fibroblasts into myofibroblasts as well as reducing the excessive deposition of extracellular matrix (ECM). The combined use of the two drugs exhibited an excellent synergistic antifibrotic effect and was conducive to minimizing alveolar epithelial damage. This work provides a codelivery strategy of AST and siTGF-β1, which shows great promise for the treatment of IPF by simultaneously reducing alveolar epithelial damage and inhibiting fibroblast activation.

ESL attenuates BLM-induced IPF in mice: Dual mediation of the TLR4/NF-κB and TGF-β1/PI3K/Akt/FOXO3a pathways

Idiopathic pulmonary fibrosis (IPF) is a persistent and advanced pulmonary ailment. The roles of innate immunity and adaptive immunity are pivotal in the evolution of IPF. An ill-adjusted interaction between epithelial cells and immune cells is responsible for initiating the epithelial-mesenchymal transition (EMT) process and sustaining chronic inflammation, thereby fostering fibrosis progression. The intricacy of IPF pathogenesis has hindered the availability of efficacious agents. Elephantopus scaber Linn. (ESL) is a canonical Chinese medicine with significant immunoregulatory effects, and its aqueous extract has been proven to attenuate IPF symptoms in bleomycin (BLM)-induced mice. However, the underlying mechanism through which ESL relieves IPF remains unclear. To validate whether ESL reverses IPF by mediating the immune response and EMT. Ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS) and UPLC were used to identify the components and determine the concentrations of the specific compounds in the ESL. Network pharmacology and molecular docking were applied to predict the potential mechanism underlying the anti-IPF effect of ESL. BLM-induced IPF mice were used to validate the anti-IPF effect of ESL, and lung tissue was collected to test putative pathways involved in inflammation and EMT via immunohistochemistry (ICH), real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Sixty-one compounds were identified, and thirteen main ingredients were quantified in the ESL. In silico experiments predicted that the IPF-mediated reversal of adverse effects by ESL would be related to interruption of the Toll-like receptor 4 (TLR4)/nuclear factor-k-gene binding (NF-ĸB) inflammatory pathway and the transforming growth factor-beta l (TGF-β1)/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/forkhead box O3 (FOXO3a) fibrosis pathway. In vivo experiments showed that ESL alleviates BLM-induced lung inflammation and fibrosis by reducing neutrophil aggregation and fibroblast foci, similar to the effects of the positive control drug pirfenidone (PFD). ESL markedly inhibited the transcription of TNF-α, IL-1β, and IL-6, which are downstream genes of the NF-κB signaling pathway. Furthermore, the protein levels of TLR4 and p-NF-κB were correspondingly inhibited in response to ESL treatment. Additionally, ESL reverses BLM-induced changes in the expression of EMT-related biological characteristic indicators (collagen I [COLIA1], E-cadherin, and alpha smooth muscle actin [α-SMA]) at the messenger ribonucleic acid (mRNA) level and markedly inhibits the expression of EMT-related upstream proteins (TGF-β1, p-PI3K, p-Akt, and p-FOXO3a). Our research suggested that ESL attenuates BLM-induced IPF through mediating the EMT process via the TGF-β1/PI3K/Akt/FOXO3a signaling pathway and inhibiting inflammation through the TLR4/NF-κB signaling pathway, highlighting that ESL can serve as an immunoregulator for relieving the abnormal immune response and reversing the EMT in IPF.

Mesenchymal stem cells-derived exosomes carrying microRNA-30b confer protection against pulmonary fibrosis by downregulating Runx1 via Spred2.

Idiopathic pulmonary fibrosis (IPF) is a chronic pulmonary fibrosis disease that is fatal. Mesenchymal stem cells (MSCs)-secreted exosomes (exos) have been linked to improving PF. Moreover, exosomal microRNAs (miRs) can control the growth of numerous diseases, including lung disorders. Our bioinformatics analysis showed that miR-30b was downregulated in tissue samples from surgical remnants of biopsies or lungs explanted from patients with IPF who underwent pulmonary transplantation. This suggests that miR-30b plays an important role in both the pathogenesis and treatment of IPF. Herein, this research was designed to ascertain the mechanism of MSCs-exos-packaged miR-30b in alleviating PF. The serum was harvested from idiopathic PF (IPF) patients with interstitial pneumonia caused by dermatomyositis and the MLE12 lung epithelial cell fibrosis model was built with TGF-β1 (10ng/mL), followed by miR-30b expression determination. TGF-β1-stimulated MLE12 cells were co-incubated with exos from MSCs with or without Spred2 or Runx1 overexpression, followed by measurement of cell viability and apoptosis. After establishing the IPF mouse model with bleomycin and injecting exos and/or silencing and overexpressing adenovirus vectors, fibrosis evaluation was conducted. In mice and cells, the expression of TGF-β1, TNF-α, and IL-1β was tested via ELISA, and the levels of E-cad, ZO-1, α-SMA, and collagen type I via western blot analysis. The promoters of miR-30b, Runx1, and Spred2 were investigated. miR-30b was downregulated in the serum of IPF patients and TGF-β1-stimulated MLE12 cells. Mechanistically, miR-30b inhibited Spred2 transcription by negatively targeting Runx1. MSCs-exos or MSCs-exo-miR-30b decreased the apoptosis, inflammation, and fibrosis while increasing their viability in TGF-β1-stimulated MLE12 cells, which was annulled by overexpressing Runx1 or Spred2. Exo-miR-30b decreased Runx1 expression to downregulate Spred2, reducing fibrosis and inflammation in IPF mice. Our results indicated that MSCs-exos-encapsulated miR-30b had a potential function to inhibit PF and part of its function may be achieved by targeting RUNX1 to reduce the Spred2 transcription level. Moreover, this work offered evidence and therapeutic targets for therapeutic strategies for managing clinical PF in patients.

SGK1 aggravates idiopathic pulmonary fibrosis by triggering H3k27ac-mediated macrophage reprogramming and disturbing immune homeostasis.

Idiopathic pulmonary fibrosis (IPF) is characterized by fibrotic matrix deposition and irreversible aberrant tissue remodeling. Their mechanisms of action are associated with the activation of macrophages and a disturbed immune environment. We aim to determine how these activated macrophages influenced the pathogenesis of pulmonary fibrosis. We found the fibrotic areas of IPF patients contained more serum and glucocorticoid-induced kinase 1 (SGK1)-positive and M2-type macrophages. Similarly, bleomycin (BLM)+LPS significantly triggered high expression of SGK1 in the IPF mice, accompanied by destroyed lung structure and function, increased fibrosis markers and disturbed immune microenvironment. Mechanistically, SGK1 markedly promoted the reprogramming of M2-type macrophages in fibrotic lungs by triggering glycogen synthase kinase 3beta (GSK3β)-tat-interacting protein 60 (TIP60)- histone-3 lysine-27 acetylation (H3K27ac) signalings, which further released chemokine (C-C motif) ligand 9 (CCL9) to attract Th17 cells and delivered TGF-β to fibroblasts for synergistically destroying immune microenvironment, which was largely reversed by macrophage depletion in mice. We took macrophages as the entry point to deeply analyze IPF pathogenesis and further provided insights for the development of novel drugs represented by SGK1.

Induced pluripotent stem cells modulate the Wnt pathway in the bleomycin-induced model of idiopathic pulmonary fibrosis

BackgroundThe Wnt signaling pathway has been implicated in the pathogenesis of fibrotic disorders and malignancies. Hence, we aimed to assess the potential of the induced pluripotent stem cells (IPS) in modulating the expression of the cardinal genes of the Wnt pathway in a mouse model of idiopathic pulmonary fibrosis (IPF).MethodsC57Bl/6 mice were randomly divided into three groups of Control, Bleomycin (BLM), and BLM + IPS; the BLM mice received intratracheal instillation of bleomycin, BLM + IPS mice received tail vein injection of IPS cells 48 h post instillation of the BLM; The Control group received Phosphate-buffered saline instead. After 3 weeks, the mice were sacrificed and Histologic assessments including hydroxy proline assay, Hematoxylin and Eosin, and Masson-trichrome staining were performed. The expression of the genes for Wnt, β-Catenin, Lef, Dkk1, and Bmp4 was assessed utilizing specific primers and SYBR green master mix.ResultsHistologic assessments revealed that the fibrotic lesions and inflammation were significantly alleviated in the BLM + IPS group. Besides, the gene expression analyses demonstrated the upregulation of Wnt, β-Catenin, and LEF along with the significant downregulation of the Bmp4 and DKK1 in response to bleomycin treatment; subsequently, it was found that the treatment of the IPF mice with IPS cells results in the downregulation of the Wnt, β-Catenin, and Lef, as well as upregulation of the Dkk1, but not the Bmp4 gene (P values < 0.05).ConclusionThe current study highlights the therapeutic potential of the IPS cells on the IPF mouse model in terms of regulating the aberrant expression of the factors contributing to the Wnt signaling pathway.

Inhalation of taraxasterol loaded mixed micelles for the treatment of idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal lung disease characterized by pulmonary inflammation, oxidative stress, and excessive extracellular matrix (ECM) deposition. Current anti-fibrotic drugs for IPF treatment in the clinic lack selectivity and demonstrate unsatisfactory efficacy, highlighting the urgent necessity for a novel therapeutic strategy. Taraxasterol (TA), which has biological activities against lung injury induced by various factors, is a potential anti-IPF drug due to its anti-inflammatory, antioxidant and lung-protective effects. However, the protective effect of TA on IPF has not been confirmed, and its clinical application is limited due to its poor aqueous solubility. In this study, we demonstrated that TA could inhibit epithelial-mesenchymal transition (EMT) and migration of A549 cells by inhibiting the transforming growth factor-β1 (TGF-β1)/Smad signaling pathway. To improve the aqueous solubility and pulmonary administration performance of TA, we prepared TA loaded methoxy poly(ethylene glycol)-poly(d,l-lactide) (mPEG-PLA)/d-α-tocopheryl polyethylene glycol succinate (TPGS) mixed polymeric micelles (TA-PM). Then a MicroSprayerⓇ Aerosolizer was used to deliver TA-PM once every two days for three weeks to evaluate their therapeutic effects on bleomycin (BLM)-induced IPF mice. Our results demonstrated that inhaled TA-PM significantly inhibited BLM-induced inflammation, oxidative stress and fibrosis in lung tissue. Furthermore, TA-PM exhibited high pulmonary deposition and retention by pulmonary administration, along with a favorable safety profile. Overall, this study emphasizes the potential of inhaled TA-PM as a promising treatment for IPF, providing a new opportunity for their clinical application.

Untargeted Metabolomics Reveals Alterations of Rhythmic Pulmonary Metabolism in IPF.

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive condition characterized by the impairment of alveolar epithelial cells. Despite continued research efforts, the effective therapeutic medication is still absent due to an incomplete understanding of the underlying etiology. It has been shown that rhythmic alterations are of significant importance in the pathophysiology of IPF. However, a comprehensive understanding of how metabolite level changes with circadian rhythms in individuals with IPF is lacking. Here, we constructed an extensive metabolite database by utilizing an unbiased reference system culturing with 13C or 15N labeled nutrients. Using LC-MS analysis via ESI and APCI ion sources, 1300 potential water-soluble metabolites were characterized and applied to evaluate the metabolic changes with rhythm in the lung from both wild-type mice and mice with IPF. The metabolites, such as glycerophospholipids and amino acids, in WT mice exhibited notable rhythmic oscillations. The concentrations of phospholipids reached the highest during the fast state, while those of amino acids reached their peak during fed state. Similar diurnal variations in the metabolite rhythm of amino acids and phospholipids were also observed in IPF mice. Although the rhythmic oscillation of metabolites in the urea cycle remained unchanged, there was a significant up-regulation in their levels in the lungs of IPF mice. 15N-ammonia in vivo isotope tracing further showed an increase in urea cycle activity in the lungs of mice with IPF, which may compensate for the reduced efficiency of the hepatic urea cycle. In sum, our metabolomics database and method provide evidence of the periodic changes in lung metabolites, thereby offering valuable insights to advance our understanding of metabolic reprogramming in the context of IPF.

Non-peptidic immunoproteasome β5i-selective inhibitor as potential treatment for idiopathic pulmonary fibrosis: Virtual screening, hit evolution and lead identification

The immunoproteasome has emerged as a potential therapeutic target for idiopathic pulmonary fibrosis (IPF). We report herein our efforts to discover novel non-peptidic immunoproteasome inhibitors as potential treatment for IPF. A structure-based virtual screening was initially performed and the hit compound VS-7 with an IC50 of 9.437 μM against β5i was identified. Hit evolution based on the interaction mode of VS-7 proceeded, and a potent β5i inhibitor 54 (IC50 = 8.463 nM) with favorable subunit-selective profiles was obtained. Compound 54 also imposed significant effects on the release of TNF-α and IL-6, the transcriptional activity of NF-κB, as well as TGF-β1 induced fibroblast proliferation, activation and collagen synthesis. Notably, when administered at 30 mg/kg in a bleomycin-induced IPF mouse model, compound 54 showed anti-fibrotic effects comparable to the clinical drug nintedanib. The results suggest that selective inhibition of immunoproteasome could be an effective approach to treat IPF.

MICALL2 participates in the regulation of epithelial-mesenchymal transition in alveolar epithelial cells – Potential roles in pulmonary fibrosis

Epithelial-mesenchymal transition (EMT) of alveolar epithelial cells is a vital process in idiopathic pulmonary fibrosis (IPF), which results in the accumulation of fibroblasts and myofibroblasts and excessive extracellular matrix deposition. Based on RNA sequencing analysis and GEO dataset reanalysis, we screened out MICALL2, a gene upregulated in the lungs of IPF mice and alveolar epithelial type II (ATII) cells from IPF patients, and aimed to explore its role in IPF. We validated the expression of MICALL2 in bleomycin (BLM)-induced IPF mice and TGF-β1-stimulated ATII cells (primary murine ATII cells and A549 cells), and explored the role of MICALL2 in IPF by knockdown of MICALL2 in BLM-induced mice and TGF-β1-stimulated ATII cells. We found that MICALL2 was upregulated in the lungs of BLM-induced mice and TGF-β1-stimulated ATII cells. MICALL2-deficient mice had reduced fibrogenesis and restrained EMT upon BLM challenge. Knockdown of MICALL2 restrained the EMT process, in vitro, through impeding β‐catenin nuclear translocation. Mechanistically, we demonstrated that NPAS2 is directly bound to the promoter of MICALL2. Altogether, our data revealed transactivation of MICALL2 induced by NPAS2, contributing to activation of the Wnt/β-catenin pathway in ATII cells, thus leading to the EMT process and subsequent pulmonary fibrosis. Interfering with MICALL2 may represent an innovative therapeutic target to mitigate the extent of IPF.

A Manganese(Ⅱ) complex as high-order multiphoton fluorescent probe for detecting nitric oxide content change in idiopathic pulmonary fibrosis mice

Nitric oxide (NO) is an important gas signaling molecule, is present in the alveoli in correlation with the extent of idiopathic pulmonary fibrosis (IPF) patients. In this work, a high-order multiphoton fluorescent probe FS-MnCl2 is designed and synthesized for detecting NO level in mice model of IPF. FS-MnCl2 has the advantage of low detection limit (11 nM) and resistance to photobleaching in vitro. The three-photon absorption cross section is greatly enhanced after FS-MnCl2 recognizes NO. Moreover, the fluorescence intensity of FS-MnCl2-NO is stronger than that of FS-MnCl2 in the presence of RNA. Biological applications suggest that FS-MnCl2 can monitor changes in NO levels in living cells and IPF mice. This work provides a new strategy for reasonably designing Mn(Ⅱ) complex that can respond to NO for predicting IPF at an early stage.

Zingiber officinale-Derived Extracellular Vesicles Attenuate Bleomycin-Induced Pulmonary Fibrosis Trough Antioxidant, Anti-Inflammatory and Protease Activity in a Mouse Model.

Idiopathic pulmonary fibrosis (IPF) is the most frequent and severe idiopathic interstitial pneumonia. It is a chronic and progressive disease with a poor prognosis and is a major cause of morbidity and mortality. This disease has no cure; therefore, there is a clinical need to search for alternative treatments with greater efficacy. In this study, we aimed to evaluate the effect of extracellular vesicles (EVs) from Zingiber officinale (EVZO) in a murine model of bleomycin (BLM)-induced IPF administered through an osmotic minipump. EVZO had an average size of 373 nm and a spherical morphology, as identified by scanning electron microscopy. Label-free proteomic analysis of EVZOs was performed by liquid chromatography coupled to mass spectrometry, and 20 proteins were identified. In addition, we demonstrated the protease activity of EVZO by gelatin-degrading zymography assay and the superoxide dismutase (SOD) activity of EVZO by an enzymatic assay. In the BLM-induced IPF mouse model, nasal administration of 50 μg of EVZO induced recovery of alveolar space size and decreased cellular infiltrate, collagen deposition, and expression of α-SMA-positive cells. Additionally, EVZO inhibited inflammatory markers such as iNOS and COX-2, lipid peroxidation, and apoptotic cells. These results show that EVZO may represent a novel natural delivery mechanism to treat IPF.

Evaluation of lung protection of Sanghuangporus sanghuang through TLR4/NF-κB/MAPK, keap1/Nrf2/HO-1, CaMKK/AMPK/Sirt1, and TGF-β/SMAD3 signaling pathways mediating apoptosis and autophagy

Idiopathic pulmonary fibrosis (IPF) is a type of interstitial pneumonia characterized by chronic and progressive fibrosis with an unknown etiology. Previous pharmacological studies have shown that Sanghuangporus sanghuang possesses various beneficial properties including immunomodulatory, hepatoprotective, antitumor, antidiabetic, anti-inflammatory, and neuroprotective effects. This study used a bleomycin (BLM)-induced IPF mouse model to illustrate the possible benefits of SS in ameliorating IPF. BLM was administered on day 1 to establish a pulmonary fibrosis mouse model, and SS was administered through oral gavage for 21 d. Hematoxylin and eosin (H&E) and Masson’s trichrome staining results showed that SS significantly reduced tissue damage and decreased fibrosis expression. We observed that SS treatment resulted in a substantial lowering in the level of pro-inflammatory cytokines like TGF-β, TNF-α, IL-1β, and IL-6 as well as MPO. In addition, we observed a notable increase in glutathione (GSH) levels. Western blot analysis of SS showed that it reduces inflammatory factors (TWEAK, iNOS, and COX-2), MAPK (JNK, p-ERK, and p-38), fibrosis-related molecules (TGF-β, SMAD3, fibronectin, collagen, α-SMA, MMP2, and MMP9), apoptosis (p53, p21, and Bax), and autophagy (Beclin-1, LC3A/B-I/II, and p62), and notably increases caspase 3, Bcl-2, and antioxidant (Catalase, GPx3, and SOD-1) levels. SS alleviates IPF by regulating the TLR4/NF-κB/MAPK, Keap1/Nrf2/HO-1, CaMKK/AMPK/Sirt1, and TGF-β/SMAD3 pathways. These results suggest that SS has a pharmacological activity that protects the lungs and has the potential to improve pulmonary fibrosis.

Early Detection and Staging of Lung Fibrosis Enabled by Collagen-Targeted MRI Protein Contrast Agent.

Chronic lung diseases, such as idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD), are major leading causes of death worldwide and are generally associated with poor prognoses. The heterogeneous distribution of collagen, mainly type I collagen associated with excessive collagen deposition, plays a pivotal role in the progressive remodeling of the lung parenchyma to chronic exertional dyspnea for both IPF and COPD. To address the pressing need for noninvasive early diagnosis and drug treatment monitoring of pulmonary fibrosis, we report the development of human collagen-targeted protein MRI contrast agent (hProCA32.collagen) to specifically bind to collagen I overexpressed in multiple lung diseases. When compared to clinically approved Gd3+ contrast agents, hProCA32.collagen exhibits significantly better r1 and r2 relaxivity values, strong metal binding affinity and selectivity, and transmetalation resistance. Here, we report the robust detection of early and late-stage lung fibrosis with stage-dependent MRI signal-to-noise ratio (SNR) increase, with good sensitivity and specificity, using a progressive bleomycin-induced IPF mouse model. Spatial heterogeneous mapping of usual interstitial pneumonia (UIP) patterns with key features closely mimicking human IPF, including cystic clustering, honeycombing, and traction bronchiectasis, were noninvasively detected by multiple MR imaging techniques and verified by histological correlation. We further report the detection of fibrosis in the lung airway of an electronic cigarette-induced COPD mouse model, using hProCA32.collagen-enabled precision MRI (pMRI), and validated by histological analysis. The developed hProCA32.collagen is expected to have strong translational potential for the noninvasive detection and staging of lung diseases, and facilitating effective treatment to halt further chronic lung disease progression.

Insights into Overlappings of Fibrosis and Cancer: Exploring the Tumor-related Cardinal Genes in Idiopathic Pulmonary Fibrosis.

The pathogenesis of idiopathic pulmonary fibrosis (IPF) is quite similar to that of cancer pathogenesis, and several pathways appear to be involved in both disorders. The mammalian target of the rapamycin (mTOR) pathway harbors several established oncogenes and tumor suppressors. The same signaling molecules and growth factors, such as vascular endothelial growth factor (VEGF), contributing to cancer development and progression play a part in fibroblast proliferation, myofibroblast differentiation, and the production of extracellular matrix in IPF development as well. The expression of candidate genes acting upstream and downstream of mTORC1, as well as Vegf and low-density lipoprotein receptor related protein 1(Lrp1), was assessed using specific primers and quantitative polymerase chain reaction (qPCR) within the lung tissues of bleomycin (BLM)-induced IPF mouse models. Lung fibrosis was evaluated by histological examinations and hydroxyproline colorimetric assay. BLM-exposed mice developed lung injuries characterized by inflammatory manifestations and fibrotic features, along with higher levels of collagen and hydroxyproline. Gene expression analyses indicated a significant elevation of regulatory associated protein of mTOR (Raptor), Ras homolog enriched in brain (Rheb), S6 kinase 1, and Eukaryotic translation initiation factor 4E-binding protein 1 (4Ebp1), as well as a significant reduction of Vegfa, Tuberous sclerosis complex (Tsc2), and Lrp1; no changes were observed in the Tsc1 mRNA level. Our findings support the elevation of S6K1 and 4EBP1 in response to the TSC/RHEB/mTORC1 axis, which profoundly encourages the development and establishment of IPF and cancer. In addition, this study suggests a possible preventive role for VEGF-A and LRP1 in the development of IPF.

Geniposide improves bleomycin-induced pulmonary fibrosis by inhibiting NLRP3 inflammasome activation and modulating metabolism

Geniposide (Gen) is a natural compound derived from Gardenia jasminoides that has anti-inflammatory, antioxidative stress, antiapoptotic, and antitumour properties. However, its mechanisms of action in idiopathic pulmonary fibrosis (IPF) have not been well studied. Recent studies have found that the progression of IPF is influenced by host metabolism. This study investigated the antifibrotic effects and metabolic modulation of Gen. Gen significantly ameliorated pathological injury in the IPF mouse model. Gen reduced the expression of proinflammatory cytokines and inhibited oxidative stress levels. Gen also inhibited NLRP3 inflammasome activation. Untargeted metabolomics analysis revealed that metabolites were affected in the lung. These metabolites were associated with the regulation of arachidonic acid metabolism, histidine metabolism, and tryptophan metabolism. Gen reduced the inflammatory response and oxidative stress and inhibited NLRP3 inflammasome activation in IPF mice. Mechanistically, Gen-mediated treatment of IPF may be linked to the regulation of host metabolism.

Astragalus polysaccharide attenuates bleomycin-induced pulmonary fibrosis by inhibiting TLR4/ NF-κB signaling pathway and regulating gut microbiota

PurposeAstragalus polysaccharide (APS) is a naturally-occurring compound derived from Astragalus membranaceus with anti-inflammatory and antioxidant properties. However, its beneficial effects and mechanisms on pulmonary fibrosis are unknown. Gut microbiota impact lung diseases via the gut-lung axis. Herein, we investigated APS progression to intervene in pulmonary fibrosis via the toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB) signaling pathway and gut microbiota homeostasis regulation. MethodsWe used bleomycin (BLM) to construct an idiopathic pulmonary fibrosis (IPF) mouse model and assessed the pathology with Masson, hematoxylin-eosin (HE), and Sirius red staining. Enzyme-linked immunosorbent assay (ELISA) kits were employed to evaluate the inflammatory cytokine levels. Western blot evaluated TLR4/NF-κB signaling pathway expression. TUNEL staining to detect apoptosis. Mice feces samples were gathered for 16S rRNA gene sequencing. ResultsOur findings revealed that APS ameliorated the extent of damage and collagen deposition in lung tissues, reduced inflammatory cytokines TNF-α, IL-6, and IL-1β levels, and decreased apoptosis. APS might attenuate the inflammatory response through TLR4/NF-κB signaling pathway inhibition. Meanwhile, the IPF mice model exhibited dysregulation of gut microbiota, and these changes were restored after APS intervention. APS may increase the proportion of probiotics, decrease that of harmful bacteria, and balance the gut microbiota via regulating metabolic pathways. ConclusionAPS ameliorated lung tissue injury in the IPF mice model, inhibited TLR4/NF-κB signaling pathway, suppressed inflammatory cytokines activation, and reduced apoptosis. Moreover, APS regulated the metabolism of gut microbiota besides beneficial bacteria content elevation.

A mitochondrial-targeted near-infrared fluorescent probe for visualizing the fluctuation of hypochlorite acid in idiopathic pulmonary fibrosis mice

Idiopathic pulmonary fibrosis (IPF) is a chronic inflammatory disease destroying lungs irreversibly with high mortality rates. There are challenges in diagnosing IPF and treating it at an early stage. Mounting evidence suggests that hypochlorous acid (HClO) can help in diagnosing inflammation and relevant conditions. Pulmonary fibrosis is linked to the mitochondrial oxidative stress where excessive HClO production is a key molecular mechanism. Measuring mitochondrial HClO levels assists in the investigations of how the mitochondrial oxidative stress affects IPF. Herein, NIR-PTZ-HClO was developed and optimized as a probe for detecting fluctuations in HClO concentrations of cells and mice models through near-infrared (NIR) fluorescence. The probe featured large Stokes shift of 150 nm, NIR turn-on signal at 650 nm, high sensitivity (45-fold) and quick HClO detection (2 s). The probe is selective for HClO in the presence of range of other analytes. NIR-PTZ-HClO visualized both endogenous and exogenous HClO in living cells (RAW264.7, H460 and A549). The probe monitored HClO in mice models with IPF and moreover the HClO profile could be tracked during the IPF process. The probe also detected precipitous decrease in HClO levels in IPF mice treated with OFEV. NIR-PTZ-HClO probe has thus the potential for earlier diagnosis of lung fibrosis, thereby improving the treatment efficacy.