Bortezomib restrains M2 polarization and reduces CXCL16-associated CXCR6+CD4 T cell chemotaxis in bleomycin-induced pulmonary fibrosis.

BACKGROUND: The development of pulmonary fibrosis involves a cascade of events, in which inflammation mediated by immune cells plays a pivotal role. Chemotherapeutic drugs have been shown to have dual effects on fibrosis, with bleomycin exacerbating pulmonary fibrosis and bortezomib alleviating tissue fibrotic processes. Understanding the intricate interplay between chemotherapeutic drugs, immune responses, and pulmonary fibrosis is likely to serve as the foundation for crafting tailored therapeutic strategies. METHODS: A model of bleomycin-induced pulmonary fibrosis was established, followed by treatment with bortezomib. Tissue samples were collected for analysis of immune cell subsets and functional assessment by flow cytometry and in vitro cell experiments. Additionally, multi-omics analysis was conducted to further elucidate the expression of chemokines and chemokine receptors, as well as the characteristics of cell populations. RESULTS: Here, we observed that the expression of CXCL16 and CXCR6 was elevated in the lung tissue of a pulmonary fibrosis model. In the context of pulmonary fibrosis or TGF-ß1 stimulation in vitro, macrophages exhibited an M2-polarized phenotype and secreted more CXCL16 than those of the control group. Moreover, flow cytometry revealed increased expression levels of CD69 and CXCR6 in pulmonary CD4 T cells during fibrosis progression. The administration of bortezomib alleviated bleomycin-induced pulmonary fibrosis, accompanied by reduced ratio of M2-polarized macrophages and decreased accumulation of CD4 T cells expressing CXCR6. CONCLUSIONS: Our findings provide insights into the key immune players involved in bleomycin-induced pulmonary fibrosis and offer preclinical evidence supporting the repurposing strategy and combination approaches to reduce lung fibrosis.

Inhibition of OGG1 ameliorates pulmonary fibrosis via preventing M2 macrophage polarization and activating PINK1-mediated mitophagy.

BACKGROUND: 8-Oxoguanine DNA glycosylase (OGG1), a well-known DNA repair enzyme, has been demonstrated to promote lung fibrosis, while the specific regulatory mechanism of OGG1 during pulmonary fibrosis remains unclarified. METHODS: A bleomycin (BLM)-induced mouse pulmonary fibrosis model was established, and TH5487 (the small molecule OGG1 inhibitor) and Mitochondrial division inhibitor 1 (Mdivi-1) were used for administration. Histopathological injury of the lung tissues was assessed. The profibrotic factors and oxidative stress-related factors were examined using the commercial kits. Western blot was used to examine protein expression and immunofluorescence analysis was conducted to assess macrophages polarization and autophagy. The conditional medium from M2 macrophages was harvested and added to HFL-1 cells for culture to simulate the immune microenvironment around fibroblasts during pulmonary fibrosis. Subsequently, the loss- and gain-of function experiments were conducted to further confirm the molecular mechanism of OGG1/PINK1. RESULTS: In BLM-induced pulmonary fibrosis, OGG1 was upregulated while PINK1/Parkin was downregulated. Macrophages were activated and polarized to M2 phenotype. TH5487 administration effectively mitigated pulmonary fibrosis, M2 macrophage polarization, oxidative stress and mitochondrial dysfunction while promoted PINK1/Parkin-mediated mitophagy in lung tissues of BLM-induced mice, which was partly hindered by Mdivi-1. PINK1 overexpression restricted M2 macrophages-induced oxidative stress, mitochondrial dysfunction and mitophagy inactivation in lung fibroblast cells, and OGG1 knockdown could promote PINK1/Parkin expression and alleviate M2 macrophages-induced mitochondrial dysfunction in HFL-1 cells. CONCLUSION: OGG1 inhibition protects against pulmonary fibrosis, which is partly via activating PINK1/Parkin-mediated mitophagy and retarding M2 macrophage polarization, providing a therapeutic target for pulmonary fibrosis.

NR2F2 alleviates pulmonary fibrosis by inhibition of epithelial cell senescence.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, fatal, and aging-associated interstitial lung disease with a poor prognosis and limited treatment options, while the pathogenesis remains elusive. In this study, we found that the expression of nuclear receptor subfamily 2 group F member 2 (NR2F2), a member of the steroid thyroid hormone superfamily of nuclear receptors, was reduced in both IPF and bleomycin-induced fibrotic lungs, markedly in bleomycin-induced senescent epithelial cells. Inhibition of NR2F2 expression increased the expression of senescence markers such as p21 and p16 in lung epithelial cells, and activated fibroblasts through epithelial-mesenchymal crosstalk, inversely overexpression of NR2F2 alleviated bleomycin-induced epithelial cell senescence and inhibited fibroblast activation. Subsequent mechanistic studies revealed that overexpression of NR2F2 alleviated DNA damage in lung epithelial cells and inhibited cell senescence. Adenovirus-mediated Nr2f2 overexpression attenuated bleomycin-induced lung fibrosis and cell senescence in mice. In summary, these data demonstrate that NR2F2 is involved in lung epithelial cell senescence, and targeting NR2F2 may be a promising therapeutic approach against lung cell senescence and fibrosis.

Bleomycin induces senescence and repression of DNA repair via downregulation of Rad51.

BACKGROUND: Bleomycin, a potent antitumor agent, is limited in clinical use due to the potential for fatal pulmonary toxicity. The accelerated DNA damage and senescence in alveolar epithelial cells (AECs) is considered a key factor in the development of lung pathology. Understanding the mechanisms for bleomycin-induced lung injury is crucial for mitigating its adverse effects. METHODS: Human lung epithelial (A549) cells were exposed to bleomycin and subsequently assessed for cellular senescence, DNA damage, and double-strand break (DSB) repair. The impact of Rad51 overexpression on DSB repair and senescence in AECs was evaluated in vitro. Additionally, bleomycin was intratracheally administered in C57BL/6 mice to establish a pulmonary fibrosis model. RESULTS: Bleomycin exposure induced dose- and time-dependent accumulation of senescence hallmarks and DNA lesions in AECs. These effects are probably due to the inhibition of Rad51 expression, consequently suppressing homologous recombination (HR) repair. Mechanistic studies revealed that bleomycin-mediated transcriptional inhibition of Rad51 might primarily result from E2F1 depletion. Furthermore, the genetic supplement of Rad51 substantially mitigated bleomycin-mediated effects on DSB repair and senescence in AECs. Notably, decreased Rad51 expression was also observed in the bleomycin-induced mouse pulmonary fibrosis model. CONCLUSIONS: Our works suggest that the inhibition of Rad51 plays a pivotal role in bleomycin-induced AECs senescence and lung injury, offering potential strategies to alleviate the pulmonary toxicity of bleomycin.

Selenite selectively kills lung fibroblasts to treat bleomycin-induced pulmonary fibrosis.

BACKGROUND: Interstitial lung disease (ILD) treatment is a critical unmet need. Selenium is an essential trace element for human life and an antioxidant that activates glutathione, but the gap between its necessity and its toxicity is small and requires special attention. Whether selenium can be used in the treatment of ILD remains unclear. METHODS: We investigated the prophylactic and therapeutic effects of selenite, a selenium derivative, in ILD using a murine model of bleomycin-induced idiopathic pulmonary fibrosis (IPF). We further elucidated the underlying mechanism using in vitro cell models and examined their relevance in human tissue specimens. The therapeutic effect of selenite in bleomycin-administered mice was assessed by respiratory function and histochemical changes. Selenite-induced apoptosis and reactive oxygen species (ROS) production in murine lung fibroblasts were measured. RESULTS: Selenite, administered 1 day (inflammation phase) or 8 days (fibrotic phase) after bleomycin, prevented and treated deterioration of lung function and pulmonary fibrosis in mice. Mechanistically, selenite inhibited the proliferation and induced apoptosis of murine lung fibroblasts after bleomycin treatment both in vitro and in vivo. In addition, selenite upregulated glutathione reductase (GR) and thioredoxin reductase (TrxR) in murine lung fibroblasts, but not in lung epithelial cells, upon bleomycin treatment. GR and TrxR inhibition eliminates the therapeutic effects of selenite. Furthermore, we found that GR and TrxR were upregulated in the human lung fibroblasts of IPF patient samples. CONCLUSIONS: Selenite induces ROS production and apoptosis in murine lung fibroblasts through GR and TrxR upregulation, thereby providing a therapeutic effect in bleomycin-induced IPF.

F127-SE-tLAP thermosensitive hydrogel alleviates bleomycin-induced skin fibrosis via TGF-ß/Smad pathway.

BACKGROUND: Skin fibrosis affects the normal function of the skin. TGF-ß1 is a key cytokine that affects organ fibrosis. The latency-associated peptide (LAP) is essential for TGF-ß1 activation. We previously constructed and prepared truncated LAP (tLAP), and confirmed that tLAP inhibited liver fibrosis by affecting TGF-ß1. SPACE peptide has both transdermal and transmembrane functions. SPACE promotes the delivery of macromolecules through the stratum corneum into the dermis. This study aimed to alleviate skin fibrosis through the delivery of tLAP by SPACE. METHODS: The SPACE-tLAP (SE-tLAP) recombinant plasmid was constructed. SE-tLAP was purified by nickel affinity chromatography. The effects of SE-tLAP on the proliferation, migration, and expression of fibrosis-related and inflammatory factors were evaluated in TGF-ß1-induced NIH-3T3 cells. F127-SE-tLAP hydrogel was constructed by using F127 as a carrier to load SE-tLAP polypeptide. The degradation, drug release, and biocompatibility of F127-SE-tLAP were evaluated. Bleomycin was used to induce skin fibrosis in mice. HE, Masson, and immunohistochemistry were used to observe the skin histological characteristics. RESULTS: SE-tLAP inhibited the proliferation, migration, and expression of fibrosis-related and inflammatory factors in NIH-3T3 cells. F127-SE-tLAP significantly reduced ECM production, collagen deposition, and fibrotic pathological changes, thereby alleviating skin fibrosis. CONCLUSION: F127-SE-tLAP could increase the transdermal delivery of LAP, reduce the production and deposition of ECM, inhibit the formation of dermal collagen fibers, and alleviate the progression of skin fibrosis. It may provide a new idea for the therapy of skin fibrosis.

Favipiravir ameliorates bleomycin-induced pulmonary fibrosis by reprogramming M1/M2 macrophage polarization.

Corona Virus Disease 2019 (COVID-19) is an infectious disease that seriously endangers human life and health. The pathological anatomy results of patients who died of the COVID-19 showed that there was an excessive inflammatory response in the lungs. It is also known that most of the COVID-19 infected patients will cause different degrees of lung damage after infection, and may have pulmonary fibrosis remaining after cure. Macrophages are a type of immune cell population with pluripotency and plasticity. In the early and late stages of infection, the dynamic changes of the balance and function of M1/M2 alveolar macrophages have a significant impact on the inflammatory response of the lungs. In the early stage of pulmonary fibrosis inflammation, the increase in the proportion of M1 type is beneficial to clear pathogenic microorganisms and promote the progress of inflammation; in the later stage of fibrosis, the increase in the number of M2 type macrophages can inhibit the inflammatory response and promote the degradation of fibrosis. As a potential treatment drug for new coronavirus pneumonia, favipiravir is in the process of continuously carried out relevant clinical trials. This study aims to discuss whether the antiviral drug favipiravir can suppress inflammation and immune response by regulating the M1/M2 type of macrophages, thereby alleviating fibrosis. We established a bleomycin-induced pulmonary fibrosis model, using IL-4/13 and LPS/IFN-γ cell stimulating factor to induce macrophage M1 and M2 polarization models, respectively. Our study shows that favipiravir exerts anti-fibrotic effects mainly by reprogramming M1/M2 macrophages polarization, that is, enhancing the expression of anti-fibrotic M1 type, reducing the expression of M2 type pro-fibrotic factors and reprogramming it to anti-fibrotic phenotype. Aspects of pharmacological mechanisms, favipiravir inhibits the activation of JAK2-STAT6 and JAK2-PI3K-AKT signaling by targeting JAK2 protein, thereby inhibiting pro-fibrotic M2 macrophages polarization and M2-induced myofibroblast activation. In summary, favipiravir can reduce the progression of pulmonary fibrosis, we hope to provide a certain reference for the treatment of pulmonary fibrosis.

L-arginine mitigates bleomycin-induced pulmonary fibrosis in rats through regulation of HO-1/PPAR-γ/ß-catenin axis.

Pulmonary fibrosis is a chronic and progressively deteriorating lung condition that can be replicated in laboratory animals by administering bleomycin, a chemotherapeutic antibiotic known for its lung fibrosis-inducing side effects. L-arginine, a semi-essential amino acid, is recognized for its diverse biological functions, including its potential to counteract fibrosis. This study aimed to evaluate the antifibrotic properties of L-arginine on bleomycin-induced pulmonary fibrosis in rats. The administration of a single intratracheal dose of bleomycin resulted in visible and microscopic damage to lung tissues, an uptick in oxidative stress markers, and an elevation in inflammatory, apoptotic, and fibrotic indicators. A seven-day treatment with L-arginine post-bleomycin exposure markedly improved the gross and histological architecture of the lungs, prevented the rise of malondialdehyde and carbonyl content, and enhanced total antioxidant capacity alongside the activities of antioxidant enzymes. Also, L-arginine attenuated the expression of the pro-fibrotic factors, transforming growth factor-ß and lactate dehydrogenase in bronchoalveolar lavage fluid. In the lung tissue, L-arginine reduced collagen deposition, hydroxyproline concentration, and mucus production, along with decreasing expression of α-smooth muscle actin, tumor necrosis factor-α, caspase-3, matrix metalloproteinase-9, and ß-catenin. Moreover, it boosted levels of nitric oxide and upregulated the expression of peroxisome proliferator-activated receptor-γ (PPAR-γ), heme oxygenase-1 (HO-1), and E-cadherin and downregulating the expression of ß-catenin. These findings suggest that L-arginine has preventive activities against bleomycin-induced pulmonary fibrosis. This effect can be attributed to the increased production of nitric oxide, which modulates the HO-1/PPAR-γ/ß-catenin axis.

Qingkailing granule alleviates pulmonary fibrosis by inhibiting PI3K/AKT and SRC/STAT3 signaling pathways.

Pulmonary fibrosis (PF) poses a significant challenge with limited treatment options and a high mortality rate of approximately 45 %. Qingkailing Granule (QKL), derived from the Angong Niuhuang Pill, shows promise in addressing pulmonary conditions. Using a comprehensive approach, combining network pharmacology analysis with experimental validation, this study explores the therapeutic effects and mechanisms of QKL against PF for the first time. In vivo, QKL reduced collagen deposition and suppressed proinflammatory cytokines in a bleomycin-induced PF mouse model. In vitro studies demonstrated QKL's efficacy in protecting cells from bleomycin-induced injury and reducing collagen accumulation and cell migration in TGF-ß1-induced pulmonary fibrosis cell models. Network pharmacology analysis revealed potential mechanisms, confirmed by western blotting, involving the modulation of PI3K/AKT and SRC/STAT3 signaling pathways. Molecular docking simulations highlighted interactions between QKL's active compounds and key proteins, showing inhibitory effects on epithelial damage and fibrosis. Collectively, these findings underscore the therapeutic potential of QKL in alleviating pulmonary inflammation and fibrosis through the downregulation of PI3K/AKT and SRC/STAT3 signaling pathways, with a pivotal role attributed to its active compounds.

Reduced Corticosteroid Exposure Is Safe and Does Not Reduce Disease Control among Hodgkin Lymphoma Patients Treated with Escalated BEACOPP (eBEACOPP).

Background and Objectives: eBEACOPP is the most effective chemotherapy regimen for younger patients with early unfavorable (EU) and advanced-stage (AS) Hodgkin lymphoma (HL), albeit with significant toxicities. The 14-day/cycle prednisone course contributes to side effects, including osteoarticular events like avascular bone necrosis (AVN). Our center has been using eBEACOPP since 2009 for AS and 2014 for EU patients. In 2016, we reduced prednisone treatment to 7-10 days to lessen AVN risk. We analyzed the effects of this approach. Materials and Methods: We retrospectively collected data on patients who received at least two cycles of eBEACOPP for first-line HL treatment. Results: A total of 162 patients (33 EU, 129 AS) were included. Their median age was 31 (range 19-59 years), and 88 were males. A total of 94 patients received full corticosteroid courses, and 68 received reduced corticosteroid courses. The overall response rate (ORR) was 98%. Different corticosteroid dosings had no significant effect on ORR, febrile neutropenia episodes, or hospital admissions. After a median follow-up (mFU) of 58 months, the 5yPFS for the entire cohort was 98% vs. 95% for the standard course vs. the short corticosteroids course, respectively (p = 0.37), while the 5yOS was 98% vs. 99% for the standard course vs. short corticosteroids course, respectively (p = 0.87). In AS patients intended to be treated with six eBEACOPP cycles, 5yPFS and 5yOS were 100% vs. 97% and 100% vs. 99% for standard vs. short corticosteroid courses, respectively (p = 0.56 and p = 0.17). In EU patients, 5yPFS was 97% (standard) vs. 95% (short) (p = 0.98) and 5yOS 100% vs. 93.3% (p = 0.87). Osteoarticular events were numerically lower in patients receiving the shorter prednisone course, both in the whole cohort and in the subgroup of patients treated with six cycles of eBEACOPP, but this difference failed to reach statistical significance. Conclusions: eBEACOPP provides excellent and durable first-line disease control. Shortening the corticosteroid course does not compromise efficacy, potentially reducing toxicity. However, longer follow-ups and larger studies are needed for confirmation.

Prevention of bleomycin-induced pulmonary fibrosis by vaccination with the Tocilizumab mimotope.

Mimotope, a kind of peptide vaccine, is developed to bind natural receptor and inhibit the downstream signaling. We have demonstrated that the vaccination of Tocilizumab mimotopes could alleviate the renal fibrosis by interfering with both IL-6 and ferroptosis signaling. However, the effect of the vaccination of Tocilizumab mimotopes on the fibroblast was not investigated in previous study. Thus, we sought to explore the changes in the fibroblast induced by the Tocilizumab mimotopes vaccination. Bleomycin instillation was performed to construct the pulmonary fibrosis model after the immunization of Tocilizumab mimotopes. Lung histological analysis showed that the Tocilizumab mimotopes could significantly reduce the maladaptive repairment and abnormal remodeling. Immunoblotting assay and fluorescence staining showed that Immunization with the Tocilizumab mimotopes reduces the accumulation of fibrosis-related proteins. High level of lipid peroxidation product was observed in the animal model, while the Tocilizumab mimotopes vaccination could reduce the generation of lipid peroxidation product. Mechanism analysis further showed that Nrf-2 signaling, but not GPX-4 and FSP-1 signaling, was upregulated, and reduced the lipid peroxidation. Our results revealed that in the BLM-induced pulmonary fibrosis, high level of lipid peroxidation product was significantly accumulation in the lung tissues, which might lead to the occurrence of ferroptosis. The IL-6 pathway block therapy could inhibit lipid peroxidation product generation in the lung tissues by upregulating the Nrf-2 signaling, and further alleviate the pulmonary fibrosis.

The XPO1 inhibitor selinexor ameliorates bleomycin-induced pulmonary fibrosis in mice via GBP5/NLRP3 inflammasome signaling.

Pulmonary fibrosis is an irreversible and progressive lung disease with limited treatments available. Selinexor (Sel), an orally available, small-molecule, selective inhibitor of XPO1, exhibits notable antitumor, anti-inflammatory and antiviral activities. However, its potential role in treating pulmonary fibrosis is unknown. C57BL/6J mice were used to establish a pulmonary fibrosis model by intratracheal administration of bleomycin (BLM). Subsequently, Sel was administered intraperitoneally. Our data demonstrated that Sel administration ameliorated BLM-induced pulmonary fibrosis by increasing mouse body weights; reducing H&E staining, Masson staining scores, and shadows in mouse lung computed tomography (CT) images, decreasing the total cell and neutrophil counts in the lung and bronchoalveolar lavage fluid (BALF); and decreasing the levels of TGF-ß1. We next confirmed that Sel reduced the deposition of extracellular matrix (ECM) components in the lungs of BLM-induced pulmonary fibrosis mice. We showed that collagen I, alpha-smooth muscle actin (α-SMA), and hydroxyproline levels and the mRNA levels of Col1a1, Eln, Fn1, Ctgf, and Fgf2 were reduced. Mechanistically, tandem mass tags (TMT)- based quantitative proteomics analysis revealed a significant increase in GBP5 in the lungs of BLM mice but a decrease in that of BLM + Sel mice; this phenomenon was confirmed by western blotting and RT-qPCR. NLRP3 inflammasome signaling was significantly enriched in both the BLM group and BLM + Sel group based on GO and KEGG analyses of differentially expressed proteins between the groups. Furthermore, Sel reduced the expression of NLRP3, cleaved caspase 1, and ASC in vivo and in vitro, and decreased the levels of IL-1ß, IL-18, and IFN-r in lung tissue and BALF. SiRNA-GBP5 inhibited NLRP3 signaling in vitro, and overexpression of GBP5 inhibited the protective effect of Sel against BLM-induced cellular injury. Taken together, our findings indicate that Sel ameliorates BLM-induced pulmonary fibrosis by targeting GBP5 via NLRP3 inflammasome signaling. Thus, the XPO1 inhibitor - Sel might be a potential therapeutic agent for pulmonary fibrosis.

CD109 Attenuates Bleomycin-induced Pulmonary Fibrosis by Inhibiting TGF-ß Signaling.

Pulmonary fibrosis is a fatal condition characterized by fibroblast and myofibroblast proliferation and collagen deposition. TGF-ß plays a pivotal role in the development of pulmonary fibrosis. Therefore, modulation of TGF-ß signaling is a promising therapeutic strategy for treating pulmonary fibrosis. To date, however, interventions targeting TGF-ß have not shown consistent efficacy. CD109 is a GPI-anchored glycoprotein that binds to TGF-ß receptor I and negatively regulates TGF-ß signaling. However, no studies have examined the role and therapeutic potential of CD109 in pulmonary fibrosis. The purpose of this study was to determine the role and therapeutic value of CD109 in bleomycin-induced pulmonary fibrosis. CD109-transgenic mice overexpressing CD109 exhibited significantly attenuated pulmonary fibrosis, preserved lung function, and reduced lung fibroblasts and myofibroblasts compared with wild-type (WT) mice. CD109-/- mice exhibited pulmonary fibrosis comparable to WT mice. CD109 expression was induced in variety types of cells, including lung fibroblasts and macrophages, upon bleomycin exposure. Recombinant CD109 protein inhibited TGF-ß signaling and significantly decreased ACTA2 expression in human fetal lung fibroblast cells in vitro. Administration of recombinant CD109 protein markedly reduced pulmonary fibrosis in bleomycin-treated WT mice in vivo. Our results suggest that CD109 is not essential for the development of pulmonary fibrosis, but excess CD109 protein can inhibit pulmonary fibrosis development, possibly through suppression of TGF-ß signaling. CD109 is a novel therapeutic candidate for treating pulmonary fibrosis.

Phœnix dactylifera, L. seed oil alleviates Bleomycin-induced pulmonary fibrosis and oxidative stress in Wistar rats.

OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is the most serious form of interstitial lung disease. We aimed to investigate the effect of Phœnix dactylifera, L. seed oil (DSO) on a murine model of IPF induced by bleomycin (BLM). METHODS: Male Wistar rats were treated with a single intra-tracheal injection of BLM (4 mg/kg) and a daily intraperitoneal injection of DSO (75, 150 and 300 mg/kg) for 4 weeks. RESULTS: Our phytochemical results showed that DSO has an important antioxidant activity with a high content of polyphenols and flavonoids. High-Performance Liquid Chromatography (HPLC) and Gas chromatography/mass spectrometry (GC-MS) analysis revealed a high amount of oleic and lauric acids and a large quantity of vitamins. Histological examination showed a significant reduction in fibrosis score and collagen bands in the group of rats treated with 75 mg/kg of DSO compared to the BLM group. DSO (75 mg/kg) reversed also the increase in catalase and malondialdehyde (MDA) levels while higher doses (150 and 300 mg/kg) are ineffective against the deleterious effects of BLM. We revealed also that DSO has no renal or hepatic cytotoxic effects. CONCLUSION: DSO can play antioxidant and antifibrotic effects on rat models of pulmonary fibrosis at the lowest dose administered.

High S100A9 level predicts poor survival, and the S100A9 inhibitor paquinimod is a candidate for treating idiopathic pulmonary fibrosis.

BACKGROUND: S100A9 is a damage-associated molecular pattern protein that may play an important role in the inflammatory response and fibrotic processes. Paquinimod is an immunomodulatory compound that prevents S100A9 activity. Its safety and pharmacokinetics have been confirmed in human clinical trials. In this study, we investigated the effects of paquinimod in preventing the development of lung fibrosis in vivo and examined the prognostic values of circulatory and lung S100A9 levels in patients with idiopathic pulmonary fibrosis (IPF). METHODS: The expression and localisation of S100A9 and the preventive effect of S100A9 inhibition on fibrosis development were investigated in a mouse model of bleomycin-induced pulmonary fibrosis. In this retrospective cohort study, the S100A9 levels in the serum and bronchoalveolar lavage fluid (BALF) samples from 76 and 55 patients with IPF, respectively, were examined for associations with patient survival. RESULTS: S100A9 expression was increased in the mouse lungs, especially in the inflammatory cells and fibrotic interstitium, after bleomycin administration. Treatment with paquinimod ameliorated fibrotic pathological changes and significantly reduced hydroxyproline content in the lung tissues of mice with bleomycin-induced pulmonary fibrosis. Additionally, we found that paquinimod reduced the number of lymphocytes and neutrophils in BALF and suppressed endothelial-mesenchymal transition in vivo. Kaplan-Meier curve analysis and univariate and multivariate Cox hazard proportion analyses revealed that high levels of S100A9 in the serum and BALF were significantly associated with poor prognoses in patients with IPF (Kaplan-Meier curve analysis: p=0.037 (serum) and 0.019 (BALF); multivariate Cox hazard proportion analysis: HR=3.88, 95% CI=1.06 to 14.21, p=0.041 (serum); HR=2.73, 95% CI=1.05 to 7.10, p=0.039 (BALF)). CONCLUSIONS: The present results indicate that increased S100A9 expression is associated with IPF progression and that the S100A9 inhibitor paquinimod is a potential treatment for IPF.

Nicotinamide phosphoribose transferase facilitates macrophage-mediated pulmonary fibrosis through the Sirt1-Smad7 pathway in mice.

Pulmonary fibrosis is a challenging lung disease characterized by a bleak prognosis. A pivotal element in the progression of this disease is the dysregulated recruitment of macrophages. Nicotinamide phosphoribose transferase (NAMPT), secreted by alveolar epithelial cells and inflammatory cells, has been previously identified to influence macrophage inflammation in acute lung injury through the nicotinamide adenine dinucleotide (NAD) rescue synthesis pathway. Nonetheless, the exact role of NAMPT in the regulation of lung fibrosis is yet to be elucidated. In our research, we employed bleomycin (BLM) to induce pulmonary fibrosis in Namptflox/flox;Cx3cr1CreER mice, using Namptflox/flox mice as controls. Our findings revealed an augmented expression of NAMPT concurrent with a marked increase in the secretion of NAD and inflammatory cytokines such as IL-6, TNF-α, and TGF-ß1 post-BLM treatment. Furthermore, an upsurge in NAMPT-positive macrophages was observed in the lungs of BLM-treated Namptflox/flox mice. Notably, a conditional knockout of NAMPT (NAMPT cKO) in lung macrophages curtailed the BLM-induced inflammatory responses and significantly mitigated pulmonary fibrosis. This was associated with diminished phospho-Sirt1 (p-Sirt1) expression levels and a concomitant rise in mothers against decapentaplegic homolog 7 (Smad7) expression in BLM-treated mouse lungs and murine RAW 264.7 macrophage cells. Collectively, our data suggests that NAMPT exacerbates macrophage-driven inflammation and pulmonary fibrosis via the Sirt1-Smad7 pathway, positioning NAMPT as a promising therapeutic target for pulmonary fibrosis intervention.

Ivermectin ameliorates bleomycin-induced lung fibrosis in male rats by inhibiting the inflammation and oxidative stress.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a pulmonary fibrotic disease characterized by a poor prognosis, which its pathogenesis involves the accumulation of abnormal fibrous tissue, inflammation, and oxidative stress. Ivermectin, a positive allosteric modulator of GABAA receptor, exerts anti-inflammatory and antioxidant properties in preclinical studies. The present study investigates the potential protective effects of ivermectin treatment in rats against bleomycin-induced IPF. MATERIALS AND METHODS: The present study involved 42 male Wistar rats, which were divided into five groups: control (without induction of IPF), bleomycin (IPF-induced by bleomycin 2.5 mg/kg, by intratracheal administration), and three fibrosis groups receiving ivermectin (0.5, 1, and 3 mg/kg). lung tissues were harvested for measurement of oxidative stress [via myeloperoxidase (MPO), superoxide dismutase (SOD), glutathione (GSH)] and inflammatory markers (tumor necrosis factor-α [TNF-α], interleukin-1ß [IL-1ß], and transforming growth factor-ß [TGF-ß]). Histological assessments of tissue damage were performed using hematoxylin-eosin (H&E) and Masson's trichrome staining methods. RESULTS: The induction of fibrosis via bleomycin was found to increase levels of MPO as well as TNF-α, IL-1ß, and TGF-ß while decrease SOD activity and GSH level. Treatment with ivermectin at a dosage of 3 mg/kg was able to reverse the effects of bleomycin-induced fibrosis on these markers. In addition, results from H&E and Masson's trichrome staining showed that ivermectin treatment at this same dose reduced tissue damage and pulmonary fibrosis. CONCLUSION: The data obtained from this study indicate that ivermectin may have therapeutic benefits for IPF, likely due to its ability to reduce inflammation and mitigate oxidative stress-induced toxicity.

Amifostine ameliorates bleomycin-induced murine pulmonary fibrosis via NAD+/SIRT1/AMPK pathway-mediated effects on mitochondrial function and cellular metabolism.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a devastating chronic lung disease characterized by irreversible scarring of the lung parenchyma. Despite various interventions aimed at mitigating several different molecular aspects of the disease, only two drugs with limited clinical efficacy have so far been approved for IPF therapy. OBJECTIVE: We investigated the therapeutic efficacy of amifostine, a detoxifying drug clinically used for radiation-caused cytotoxicity, in bleomycin-induced murine pulmonary fibrosis. METHODS: C57BL6/J mice were intratracheally instilled with 3 U/kg of bleomycin. Three doses of amifostine (WR-2721, 200 mg/kg) were administered intraperitoneally on days 1, 3, and 5 after the bleomycin challenge. Bronchoalveolar lavage fluid (BALF) was collected on day 7 and day 21 for the assessment of lung inflammation, metabolites, and fibrotic injury. Human fibroblasts were treated in vitro with transforming growth factor beta 1 (TGF-ß1), followed by amifostine (WR-1065, 1-4 µg/mL) treatment. The effects of TGF-ß1 and amifostine on the mitochondrial production of reactive oxygen species (ROS) were assessed by live cell imaging of MitoSOX. Cellular metabolism was assessed by the extracellular acidification rate (ECAR), the oxygen consumption rate (OCR), and the concentrations of various energy-related metabolites as measured by mass spectrum (MS). Western blot analysis was performed to investigate the effect of amifostine on sirtuin 1 (SIRT1) and adenosine monophosphate activated kinase (AMPK). RESULTS: Three doses of amifostine significantly attenuated lung inflammation and pulmonary fibrosis. Pretreatment and post-treatment of human fibroblast cells with amifostine blocked TGF-ß1-induced mitochondrial ROS production and mitochondrial dysfunction in human fibroblast cells. Further, treatment of fibroblasts with TGF-ß1 shifted energy metabolism away from mitochondrial oxidative phosphorylation (OXPHOS) and towards glycolysis, as observed by an altered metabolite profile including a decreased ratio of NAD + /NADH and increased lactate concentration. Treatment with amifostine significantly restored energy metabolism and activated SIRT1, which in turn activated AMPK. The activation of AMPK was required to mediate the effects of amifostine on mitochondrial homeostasis and pulmonary fibrosis. This study provides evidence that repurposing of the clinically used drug amifostine may have therapeutic applications for IPF treatment. CONCLUSION: Amifostine inhibits bleomycin-induced pulmonary fibrosis by restoring mitochondrial function and cellular metabolism.

The Impact of Bleomycin Deficit on Survival in Hodgkin's Lymphoma Patients: A Retrospective Study.

PURPOSE: Hodgkin's lymphoma is currently treated with a chemotherapy protocol consisting of doxorubicin, bleomycin, vinblastine, and dacarbazine. Due to Brazil facing a bleomycin shortage in 2017, and this drug's high toxicity, this retrospective study evaluates the effect that the absence of bleomycin had on treatment response and overall survival of Hodgkin's lymphoma patients. METHODS: The medical records of 126 HL patients treated between 2007 and 2021 were reviewed and their data collected, followed by grouping into ABVD and AVD groups according to bleomycin use. Data concerning the patient's characteristics, cancer type, and treatment plan were analyzed with proportion tests, Kaplan-Meier curves. univariate Cox regression, and χ2 tests. RESULTS: No discernible differences were found in this study between the overall survival and recurrence rate of patients treated with bleomycin compared to those without. Additionally, there was an increased risk of death in each subsequent cycle of chemotherapy of the complete ABVD protocol, demonstrating a risk of toxicity. Among the variables analyzed, hypertension and the presence of B symptoms were also associated with an increased risk of death, while the use of radiotherapy significantly improved survival. CONCLUSION: The results of this study suggest that bleomycin did not impact the outcome of Hodgkin's lymphoma treatment. Moreover, the increased risk of death associated with its toxicity during each cycle of treatment raises concerns about its role as an essential component of the gold standard for Hodgkin's lymphoma treatment. Therefore, further research and consideration are needed to reassess the use of bleomycin in Hodgkin's lymphoma treatment protocols.

Menstrual blood-derived stem cells exosomal miR-let-7 to ameliorate pulmonary fibrosis through inhibiting ferroptosis by Sp3/HDAC2/Nrf2 signaling pathway.

Idiopathic pulmonary fibrosis (IPF) is a serious, lifelong lung disease with high morbidity and high mortality. Menstrual blood-derived stem cells (MenSCs) derived exosomes (MenSCs-Exo) emerge as an attractive tool for the treatment of acute lung injury and fibrosis-related diseases. However, more comprehensive mechanism over how MenSCs derived exosomes exhibits anti-pulmonary fibrosis needs to be elucidated. In this study, TGF-ß was used to construct cell fibrosis model, and bleomycin (BLM) was applied to induce lung tissue fibrosis mice model. BLM- and TGF-ß1-induced cellular reactive oxygen species (ROS), mitochondrial DNA (mtDNA) damage, and lung epithelial cell apoptosis were alleviated by MenSCs-Exo treatment in vivo and in vitro. Besides, it was found that MenSCs-Exo delivered miR-let-7 into MLE-12 cells/lung epithelial cell and the reduction of miR-let-7 blocked the improvement produced by MenSCs-Exo. Mechanistically, miR-let-7 directly bound to Sp3 and negatively regulated its expression. Sp3 elevation promoted the expression of ferroptosis-related protein and mitochondrial DNA (mtDNA) damage markers via recruiting HDAC2, thereby inactivating keap1/Nrf2 signal cascade, which were confirmed in BLM-induced pulmonary fibrosis mice model under the combination therapy of the MenSCs-Exo and let-7 inhibitor. Collectively, MenSCs derived exosomes could transmit miR-let-7 into MLE-12 cells to inhibit the expression of Sp3, thereby weakening the recruitment effect of Sp3 on HDAC2, lifting the deacetylation restriction of HDAC2 on Nrf2, and enhancing the Nrf2 pathway. These changes further declined ferroptosis and delayed the pathological process of oxidative damage and lung epithelial cell apoptosis in PF.