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Background: Although bone marrow-derived cells with high aldehyde dehydrogenase activity (ALDH br ) have shown therapeutic potential against various diseases in animal studies, clinical trials have failed to show concurrent findings. We aimed to clarify the optimal conditions for the efficacy of ALDH br cells by using a murine bleomycin-induced pulmonary fibrosis model., Methods: We intravenously transferred male or female donor C57BL/6 mice-derived ALDH br cells into recipient C57BL/6 mice under various conditions, and used mCherry-expressing mice as a donor to trace the transferred ALDH br cells., Results: Pulmonary fibrosis improved significantly when (1) female-derived, not male-derived, and (2) lineage (Lin)-negative, not lineage-positive, ALDH br cells were transferred during the (3) fibrotic, not inflammatory, phase. Consistent with the RNA-sequencing results, female-derived Lin - /ALDH br cells were more resistant to oxidative stress than male-derived cells in vitro, and transferred female-derived Lin - /ALDH br cells were more viable than male-derived cells in the fibrotic lung. The mechanism underlying the antifibrotic effects of Lin - /ALDH br cells was strongly associated with reduction of oxidative stress., Conclusions: Our results indicated that Lin - /ALDH br cell therapy could ameliorate pulmonary fibrosis by reducing oxidative stress and suggested that their efficacy was mediated by sex-related differences. Thus, sex-awareness strategies may be important for clinical application of bone marrow ALDH br cells as a therapeutic tool., (© 2024. The Author(s).)

Background: The prognosis of patients with lung cancer accompanied by interstitial pneumonia is poorer than that of patients with lung cancer but without interstitial pneumonia. Moreover, the available therapeutic interventions for lung cancer patients with interstitial pneumonia are limited. Therefore, a new treatment strategy for these patients is required. The aim of the present study was to investigate the pathophysiological relationship between interstitial pneumonia and lung cancer and explore potential therapeutic agents., Methods: A novel hybrid murine model of lung cancer with interstitial pneumonia was established via bleomycin-induced pulmonary fibrosis followed by orthotopic lung cancer cell transplantation into the lungs. Changes in tumor progression, lung fibrosis, RNA expression, cytokine levels, and tumor microenvironment in the lung cancer with interstitial pneumonia model were investigated, and therapeutic agents were examined. Additionally, clinical data and samples from patients with lung cancer accompanied by interstitial pneumonia were analyzed to explore the potential clinical significance of the findings., Results: In the lung cancer with interstitial pneumonia model, accelerated tumor growth was observed based on an altered tumor microenvironment. RNA sequencing analysis revealed upregulation of the hypoxia-inducible factor 1 signaling pathway. These findings were consistent with those obtained for human samples. Moreover, we explored whether ascorbic acid could be an alternative treatment for lung cancer with interstitial pneumonia to avoid the disadvantages of hypoxia-inducible factor 1 inhibitors. Ascorbic acid successfully downregulated the hypoxia-inducible factor 1 signaling pathway and inhibited tumor progression and lung fibrosis., Conclusions: The hypoxia-inducible factor 1 pathway is critical in lung cancer with interstitial pneumonia and could be a therapeutic target for mitigating interstitial pneumonia-mediated lung cancer progression., (© 2023. The Author(s).)

Increasing evidence indicates an interaction between the intestinal microbiota and diseases in distal organs. However, the relationship between pulmonary fibrosis and the intestinal microbiota, especially intestinal fungal microbiota, is poorly understood. Thus, this study aimed to determine the effects of changes in the intestinal fungal microbiota on the pathogenesis of pulmonary fibrosis. Mice with intestinal overgrowth of Candida albicans, which was established by oral administration of antibiotics plus C. albicans, showed accelerated bleomycin-induced pulmonary fibrosis relative to the control mice (i.e. without C. albicans treatment). In addition, the mice with intestinal overgrowth of C. albicans showed enhanced Th17-type immunity, and treatment with IL-17A-neutralizing antibody alleviated pulmonary fibrosis in these mice but not in the control mice. This result indicates that IL-17A is involved in the pathogenesis of C. albicans-exacerbated pulmonary fibrosis. Even before bleomycin treatment, the expression of Rorc, the master regulator of Th17, was already upregulated in the pulmonary lymphocytes of the mice with intestinal overgrowth of C. albicans. Subsequent administration of bleomycin triggered these Th17-skewed lymphocytes to produce IL-17A, which enhanced endothelial-mesenchymal transition. These results suggest that intestinal overgrowth of C. albicans exacerbates pulmonary fibrosis via IL-17A-mediated endothelial-mesenchymal transition. Thus, it might be a potential therapeutic target in pulmonary fibrosis. This study may serve as a basis for using intestinal fungal microbiota as novel therapeutic targets in pulmonary fibrosis. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland., (© 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.)

Background: Mesenchymal stromal cells (MSCs) are a potential therapeutic tool for pulmonary fibrosis. However, ex vivo MSC expansion using serum poses risks of harmful immune responses or unknown pathogen infections in the recipients. Therefore, MSCs cultured in serum-free media (SF-MSCs) are ideal for clinical settings; however, their efficacy in pulmonary fibrosis is unknown. Here, we investigated the effects of SF-MSCs on bleomycin-induced pulmonary inflammation and fibrosis compared to those of MSCs cultured in serum-containing media (S-MSCs)., Methods: SF-MSCs and S-MSCs were characterized in vitro using RNA sequence analysis. The in vivo kinetics and efficacy of SF-MSC therapy were investigated using a murine model of bleomycin-induced pulmonary fibrosis. For normally distributed data, Student's t test and one-way repeated measures analysis of variance followed by post hoc Tukey's test were used for comparison between two groups and multiple groups, respectively. For non-normally distributed data, Kruskal-Wallis and Mann-Whitney U tests were used for comparison between groups, using e Bonferroni's correction for multiple comparisons. All tests were two-sided, and P < 0.05 was considered statistically significant., Results: Serum-free media promoted human bone marrow-derived MSC expansion and improved lung engraftment of intravenously administered MSCs in recipient mice. SF-MSCs inhibited the reduction in serum transforming growth factor-β1 and the increase of interleukin-6 in both the serum and the bronchoalveolar lavage fluid during bleomycin-induced pulmonary fibrosis. SF-MSC administration increased the numbers of regulatory T cells (Tregs) in the blood and lungs more strongly than in S-MSC administration. Furthermore, SF-MSCs demonstrated enhanced antifibrotic effects on bleomycin-induced pulmonary fibrosis, which were diminished by antibody-mediated Treg depletion., Conclusions: SF-MSCs significantly suppressed BLM-induced pulmonary inflammation and fibrosis through enhanced induction of Tregs into the lungs and corrected the dysregulated cytokine balance. Therefore, SF-MSCs could be a useful tool for preventing pulmonary fibrosis progression without the demerits of serum use., (© 2021. The Author(s).)

Background: Aldehyde dehydrogenase (ALDH) is highly expressed in stem/progenitor cells in various tissues, and cell populations with high ALDH activity (ALDH br ) are associated with tissue repair. However, little is known about lung-resident ALDH br . This study was performed to clarify the characteristics of lung-resident ALDH br cells and to evaluate their possible use as a tool for cell therapy using a mouse model of bleomycin-induced pulmonary fibrosis., Methods: The characteristics of lung-resident/nonhematopoietic (CD45 - ) ALDH br cells were assessed in control C57BL/6 mice. The kinetics and the potential usage of CD45 - /ALDH br for cell therapy were investigated in bleomycin-induced pulmonary fibrosis. Localization of transferred CD45 - /ALDH br cells was determined using mCherry-expressing mice as donors. The effects of aging on ALDH expression were also assessed using aged mice., Results: Lung CD45 - /ALDH br showed higher proliferative and colony-forming potential than cell populations with low ALDH activity. The CD45 - /ALDH br cell population, and especially its CD45 - /ALDH br /PDGFRα + subpopulation, was significantly reduced in the lung during bleomycin-induced pulmonary fibrosis. Furthermore, mRNA expression of ALDH isoforms was significantly reduced in the fibrotic lung. When transferred in vivo into bleomycin-pretreated mice, CD45 - /ALDH br cells reached the site of injury, ameliorated pulmonary fibrosis, recovered the reduced expression of ALDH mRNA, and prolonged survival, which was associated with the upregulation of the retinol-metabolizing pathway and the suppression of profibrotic cytokines. The reduction in CD45 - /ALDH br /PDGFRα + population was more remarkable in aged mice than in young mice., Conclusions: Our results strongly suggest that the lung expression of ALDH and lung-resident CD45 - /ALDH br cells are involved in pulmonary fibrosis. The current study signified the possibility that CD45 - /ALDH br cells could find application as novel and useful cell therapy tools in pulmonary fibrosis treatment., (© 2021. The Author(s).)

Amphiregulin (AREG), an epidermal growth factor receptor ligand, is implicated in tissue repair and fibrosis, but its cellular source and role in regeneration versus fibrosis remain unclear. In this study, we hypothesize that AREG induced in bone marrow-derived CD11c(+) cells is essential for pulmonary fibrosis. Thus, the objectives were to evaluate the importance and role of AREG in pulmonary fibrosis, identify the cellular source of AREG induction, and analyze its regulation of fibroblast function and activation. The results showed that lung AREG expression was significantly induced in bleomycin-induced pulmonary fibrosis. AREG deficiency in knockout mice significantly diminished pulmonary fibrosis. Analysis of AREG expression in major lung cell types revealed induction in fibrotic lungs predominantly occurred in CD11c(+) cells. Moreover, depletion of bone marrow-derived CD11c(+) cells suppressed both induction of lung AREG expression and pulmonary fibrosis. Conversely, adoptive transfer of bone marrow-derived CD11c(+) cells from bleomycin-treated donor mice exacerbated pulmonary fibrosis, but not if the donor cells were made AREG deficient prior to transfer. CD11c(+) cell-conditioned media or coculture stimulated fibroblast proliferation, activation, and myofibroblast differentiation in an AREG-dependent manner. Furthermore, recombinant AREG induced telomerase reverse transcriptase, which appeared to be essential for the proliferative effect. Finally, AREG significantly enhanced fibroblast motility, which was associated with increased expression of α6 integrin. These findings suggested that induced AREG specifically in recruited bone marrow-derived CD11c(+) cells promoted bleomycin-induced pulmonary fibrosis by activation of fibroblast telomerase reverse transcriptase-dependent proliferation, motility, and indirectly, myofibroblast differentiation., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Rationale: Bone marrow (BM)-derived cells have been implicated in pulmonary fibrosis. However, their precise role in pathogenesis is incompletely understood., Objectives: To elucidate roles of BM-derived cells in bleomycin-induced pulmonary fibrosis, and clarify their potential relationship to lung hematopoietic progenitor cells (LHPCs)., Methods: GFP BM-chimera mice treated with or without bleomycin were used to assess the BM-derived cells., Measurements and Main Results: GFP(+) cells in the chimera lung were found to be comprised of two distinct phenotypes: GFP(hi) and GFP(low) cells. The GFP(hi), but not GFP(low), population was significantly increased after bleomycin treatment. Flow-cytometric analysis and quantitative real-time polymerase chain reaction revealed that GFP(hi) cells exhibited phenotypic characteristics of CD11c(+) dendritic cells and macrophages. GFP(hi) cell conditioned media were chemotactic for fibroblasts obtained from fibrotic but not normal lung in vitro. Moreover, adoptive transfer of GFP(hi) cells exacerbated fibrosis in recipient mice, similar to that seen on adoptive transfer of BM-derived CD11c(+) cells from donor bleomycin-treated mice. Next, we evaluated the potential of LHPCs as the source of GFP(hi) cells. Isolation of LHPCs by flow sorting revealed enrichment in cKit(+)/Sca1(-)/Lin(-) cells, most of which were GFP(+) indicating their BM origin. The number of LHPCs increased rapidly after bleomycin treatment. Furthermore, stem cell factor induced LHPC proliferation, whereas granulocyte-macrophage-colony stimulating factor induced differentiation to GFP(hi) cells., Conclusions: BM-derived LHPCs with a novel phenotype could differentiate into GFP(hi) cells, which enhanced pulmonary fibrosis. Targeting this mobilized LHPCs might represent a novel therapeutic approach in chronic fibrotic lung diseases.

Stem cell factor (SCF) and its receptor c-Kit have been implicated in tissue remodelling and fibrosis. Alveolar fibroblasts from patients with diffuse interstitial fibrosis secrete more SCF. However, its precise role remains unclear. In this study the potential role of the SCF-c-Kit axis in pulmonary fibrosis was examined. Fibrosis was induced by intratracheal instillation of bleomycin (BLM), which caused increased SCF levels in plasma, bronchoalveolar lavage fluid (BALF) and lung tissue, as well as increased expression by lung fibroblasts. These changes were accompanied by increased numbers of bone marrow-derived c-Kit(+) cells in the lung, with corresponding depletion in bone marrow. Both recombinant SCF and lung extracts from BLM-treated animals induced bone-marrow cell migration, which was blocked by c-Kit inhibitor. The migrated cells promoted myofibroblast differentiation when co-cultured with fibroblasts, suggesting a paracrine pathogenic role. Interestingly, lung fibroblast cultures contained a subpopulation of cells that expressed functionally active c-Kit, which were significantly greater and more responsive to SCF induction when isolated from fibrotic lungs, including those from patients with idiopathic pulmonary fibrosis (IPF). This c-Kit(+) subpopulation was αSMA-negative and expressed lower levels of collagen I but significantly higher levels of TGFβ than c-Kit-negative cells. SCF deficiency achieved by intratracheal treatment with neutralizing anti-SCF antibody or by use of Kitl(Sl)/Kitl(Sl-d) mutant mice in vivo resulted in significant reduction in pulmonary fibrosis. Taken together, the SCF-c-Kit pathway was activated in BLM-injured lung and might play a direct role in pulmonary fibrosis by the recruitment of bone marrow progenitor cells capable of promoting lung myofibroblast differentiation., (Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

The CCAAT/enhancer-binding protein β (C/EBPβ) regulates a variety of factors and cellular responses associated with pulmonary fibrosis. To distinguish its role in the mesenchyme from that in other compartments, the effects of mesenchymal-specific deletion of C/EBPβ on pulmonary fibrosis was examined. Crossing of mice with the floxed C/EBPβ gene with α2(I) collagen enhancer-CreER(T)-bearing mice successfully generated progeny with a conditional knockout (CKO) of C/EBPβ in collagen I-expressing ("mesenchymal") cells only on treatment with tamoxifen (C/EBPβ CKO). When treated with an endotracheal bleomycin injection, C/EBPβ CKO mice showed significant attenuation of pulmonary fibrosis relative to control C/EBPβ-intact mice. C/EBPβ CKO mice also had reduced myofibroblasts in the lung. However, no significant differences in inflammatory/immune cell influx were noted in the mutant mice relative to the control mice. DNA microarray and real-time PCR analyses identified a series of myofibroblast differentiation regulators as novel target genes of C/EBPβ. Interestingly, C/EBPβ deficiency caused a marked induction of matrix metalloproteinase 12 expression, suggesting its potential role as a repressor, which could account for the noted reduction in fibrosis in the C/EBPβ-deficient mice. Thus, these findings indicate an essential role for C/EBPβ in the mesenchymal compartment in pulmonary fibrosis that is independent of its effects on inflammation or immune cell infiltration., (Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Found in inflammatory zone (FIZZ) 2, also known as resistin-like molecule (RELM)-β, belongs to a novel cysteine-rich secreted protein family named FIZZ/RELM. Its function is unclear, but a closely related family member, FIZZ1, has profibrotic activities. The human ortholog of rodent FIZZ1 has not been identified, but human FIZZ2 has significant sequence homology to both rodent FIZZ2 (59%) and FIZZ1 (50%). Given the greater homology to rodent FIZZ2, analyzing the role of FIZZ2 in a rodent model of bleomycin-induced pulmonary fibrosis would be of greater potential relevance to human fibrotic lung disease. The results showed that FIZZ2 was highly induced in lungs of rodents with bleomycin-induced pulmonary fibrosis and of human patients with idiopathic pulmonary fibrosis. FIZZ2 expression was induced in rodent and human lung epithelial cells by Th2 cytokines, which was mediated via STAT6 signaling. The FIZZ2 induction in murine lungs was found to be essential for pulmonary fibrosis, as FIZZ2 deficiency significantly suppressed pulmonary fibrosis and associated enhanced extracellular matrix and cytokine gene expression. In vitro analysis indicated that FIZZ2 could stimulate type I collagen and α-smooth muscle actin expression in lung fibroblasts. Furthermore, FIZZ2 was shown to have chemoattractant activity for bone marrow (BM) cells, especially BM-derived CD11c(+) dendritic cells. Notably, lung recruitment of BM-derived cells was impaired in FIZZ2 knockout mice. These findings suggest that FIZZ2 is a Th2-associated multifunctional mediator with potentially important roles in the pathogenesis of fibrotic lung diseases.

Background: Suppressor of cytokine signaling (SOCS) proteins are inhibitors of cytokine signaling. Our previous study suggested that SOCS1 regulates collagen synthesis by lung fibroblasts, suggesting a role of SOCS1 in the pathophysiology of pulmonary fibrosis., Objectives: We sought to investigate the role of SOCS1 in pulmonary inflammation and fibrosis in vivo., Methods: SOCS1-haplodeficient mice treated with bleomycin (BLM) were evaluated for pulmonary inflammation and fibrosis compared with wild-type mice. The human study group was composed of 18 patients with interstitial lung disease. Lung specimens obtained by means of open lung biopsy were investigated to determine whether the severity of fibrosis was associated with decreased SOCS1 expression. Finally, we further analyzed the effect of exogenous SOCS1 on BLM-induced lung injury based on adenoviral SOCS1 gene transfer to the lung., Results: SOCS1-haplodeficient mice treated with BLM showed markedly enhanced pulmonary inflammation and fibrosis compared with wild-type mice. Using human lung specimens, we found that SOCS1 mRNA levels inversely correlated with duration of the disease. SOCS1 expression was significantly less in lung tissue from patients with idiopathic pulmonary fibrosis (IPF) compared with that in non-IPF patients. Moreover, SOCS1 expression was significantly less in severe fibrotic lesions (lower lobe) than in less fibrotic lesions (upper lobe). Adenoviral SOCS1 gene transfer to murine lungs significantly decreased lymphocytic inflammation, pulmonary fibrosis, and mortality because of BLM-induced lung injury. Exogenous SOCS1 inhibited expression of various cytokines, including TNF-alpha, which might play a key role., Conclusions: These results suggest that SOCS1 might act as a suppressor for pulmonary fibrosis. SOCS1 might be a target of IPF treatment.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and often fatal pulmonary disorder, and its pathology is characterized by parenchymal fibrosis. To investigate the characteristics of fibroblasts in IPF, we obtained eight fibroblast cell lines from lungs with IPF and eight lines from normal lungs. We found that the fibroblasts from IPF spontaneously produced higher amounts of type I collagen and had lower expression levels of SOCS1 than fibroblasts from normal lung. By using mouse fibroblasts, we demonstrated the causal relationship between them: the deficiency of SOCS1 in fibroblasts resulted in increased collagen production, whereas overexpression of SOCS1 suppressed collagen production. IFN-gamma suppressed spontaneous collagen production even in SOCS1-deficient fibroblasts, indicating that IFN-gamma inhibition is SOCS1-independent. In contrast, IFN-gamma suppressed the increase of collagen production induced by IL-4 in wild type fibroblasts but not SOCS1-deficient fibroblasts, suggesting IFN-gamma acted exclusively via SOCS1 in this case. Following IFN-gamma stimulation, the amount of SOCS1 mRNA expressed by IPF fibroblasts was comparable to that of normal fibroblasts. Thus, the extent of SOCS1 increase after stimulation by IFN-gamma was significantly higher in IPF fibroblasts. The extent to which IFN-gamma inhibited collagen production was also larger in IPF fibroblasts than in normal fibroblasts. These results suggest that the exaggerated production of collagen observed in fibroblasts from IPF is causally related to the diminished expression of SOCS1, and IPF fibroblasts are more susceptible to IFN-gamma because of decreased expression of SOCS1.

Idiopathic pulmonary fibrosis is a chronic, fibrosing interstitial pneumonia that presents with various clinical courses and progression ranging from rapid to slow. To identify novel biomarkers that can support the diagnosis and/or prognostic prediction of idiopathic pulmonary fibrosis, we performed gene expression analysis, and the mRNA of interleukin-18 binding protein was increasingly expressed in patients with idiopathic pulmonary fibrosis compared with healthy controls. Therefore, we hypothesized that the interleukin-18 binding protein can serve as a diagnostic and/or prognostic biomarker for idiopathic pulmonary fibrosis. We investigated the expression of interleukin-18 binding protein in lung tissue, bronchoalveolar lavage fluid, and serum. Additionally, the correlation between interleukin-18 binding protein expression levels and the extent of fibrosis was investigated using mouse models of lung fibrosis induced by subcutaneous bleomycin injections. Serum interleukin-18 binding protein levels were significantly higher in idiopathic pulmonary fibrosis patients (5.06 ng/mL, interquartile range [IQR]: 4.20–6.35) than in healthy volunteers (3.31 ng/mL, IQR: 2.84–3.99) (p < 0.001). Multivariate logistic regression models revealed that the correlation between serum interleukin-18 binding protein levels and idiopathic pulmonary fibrosis was statistically independent after adjustment for age, sex, and smoking status. Multivariate Cox proportional hazard models revealed that serum interleukin-18 binding protein levels were predictive of idiopathic pulmonary fibrosis disease prognosis independent of other covariate factors (hazard ratio: 1.655, 95% confidence interval: 1.224–2.237, p = 0.001). We also demonstrated a significant positive correlation between lung hydroxyproline expression levels and interleukin-18 binding protein levels in bronchoalveolar lavage fluid from bleomycin-treated mice (Spearman r = 0.509, p = 0.004). These results indicate the utility of interleukin-18 binding protein as a novel prognostic biomarker for idiopathic pulmonary fibrosis. [ABSTRACT FROM AUTHOR]

Rationale: The impact of lung insult on the bone marrow (BM) and subsequent disease is unknown.Objectives: To study alterations in the BM in response to lung injury/fibrosis and examine their impact on subsequent lung insult.Methods: BM cells from control or bleomycin-treated donor mice were transplanted into naive mice, which were subsequently evaluated for bleomycin-induced pulmonary fibrosis. In addition, the effect of prior bleomycin treatment on subsequent fibrosis was examined in wild-type and B7H3-knockout mice. Samples from patients with idiopathic pulmonary fibrosis were analyzed for potential clinical relevance of the findings.Measurements and Main Results: Recipient mice transplanted with BM from bleomycin-pretreated donors showed significant exacerbation of subsequent fibrosis with increased B7H3+ cell numbers and a T-helper cell type 2-skewed phenotype. Pretreatment with a minimally fibrogenic/nonfibrogenic dose of bleomycin also caused exacerbation, but not in B7H3-deficient mice. Exacerbation was not observed if the mice received naive BM cell transplant after the initial bleomycin pretreatment. Soluble B7H3 stimulated BM Ly6Chi monocytic cell expansion in vitro and caused similar expansion in the lung in vivo. Notably, soluble B7H3 was elevated in plasma of patients with idiopathic pulmonary fibrosis and in BAL fluid in those with acute exacerbation. Finally, ST2 deficiency diminished the bleomycin-induced B7H3 and IL-13 upregulation, suggesting a role for type 2 innate lymphoid cells.Conclusions: Pulmonary fibrosis caused significant alterations in BM with expansion and activation of monocytic cells, which enhanced fibrosis when transplanted to naive recipients with potential mediation by a novel role for B7H3 in the pathophysiology of pulmonary fibrosis in both mice and humans. [ABSTRACT FROM AUTHOR]

Although radiotherapy is important in the treatment of malignant thoracic tumors, it has harmful effects on healthy tissues. We previously showed that suplatast tosilate, an anti-allergic agent, scavenged reactive oxygen species (ROS), including hydroxyl radicals. Because ROS-mediated oxidative stress is involved in radiation-induced lung injury, we hypothesized that suplatast tosilate could reduce radiation-induced lung injury via suppression of oxidative stress. Murine alveolar epithelial cells were irradiated with or without a medium containing suplatast tosilate in vitro to determine whether the agent had cytoprotective effects against radiation-induced injury. On the other hand, the thoracic region of C57BL/6 mice was exposed to a single irradiation dose of 15 Gy and the effects of suplatast tosilate were determined by a histological evaluation and assessment of the following parameters: cell number and inflammatory cytokine levels in bronchoalveolar lavage fluid, and oxidative stress markers and hydroxyproline content in pulmonary tissues. Suplatast tosilate protected murine alveolar epithelial cells in vitro from irradiation-induced inhibition of cell proliferation, which was accompanied by the suppression of intracellular ROS and DNA double-strand breaks induced by irradiation. Oxidative stress markers and the levels of inflammatory and fibrogenic cytokines were upregulated in irradiated murine lungs in vivo. Suplatast tosilate suppressed both oxidative stress markers and the levels of cytokines, which resulted in reduced pulmonary fibrosis and clearly improved the survival rate after irradiation. These findings demonstrate that suplatast tosilate could be a useful lung-protective agent that acts via suppression of oxidative stress associated with thoracic radiotherapy. Image 1 • Suplatast tosilate (ST) protects lungs against radiation-induced lung injury. • ST prolongs the survival of mice exposed to thoracic irradiation. • ST suppresses radiation-induced pulmonary oxidative stress. • Cytoprotective effects of ST against radiation-induced injury were demonstrated. [ABSTRACT FROM AUTHOR]

Plasminogen activator inhibitor‐1 (PAI‐1) promotes pulmonary fibrosis through increasing myofibroblast (MF) characteristics, expressing alpha‐smooth muscle actin (α‐SMA) in fibroblasts. Fibroblasts in the tumour stroma are called cancer‐associated fibroblasts (CAFs). Some CAFs have MF characteristics and substantially promote tumour progression and chemotherapy resistance. This study determined whether inhibition of PAI‐1 suppressed MF characteristics of CAFs and limited chemotherapy resistance in lung cancer. To investigate cellular PAI‐1 expression and its correlation with α‐SMA expression of CAFs, 34 patients' paraffin‐embedded lung adenocarcinoma tissue sections were immunohistochemically stained for PAI‐1 and α‐SMA. Immunohistochemical analysis of lung adenocarcinoma tissues showed that PAI‐1 expression was correlated with that of α‐SMA (r = 0.71, p < 0.001). Furthermore, in vitro, α‐SMA expression of CAFs was limited by PAI‐1 inhibition, and apoptosis of CAFs was increased. In addition, the effectiveness of cisplatin on lung cancer cells co‐cultured with CAFs was increased by suppressing α‐SMA expression using PAI‐1 inhibitor. In lung adenocarcinoma tissues, PAI‐1 expression was associated with T factor and TNM stage. Our data suggest that inhibition of PAI‐1 increased the chemotherapeutic effect on lung cancer through suppressing the MF characteristics of CAFs. Hence, PAI‐1 might be a promising therapeutic target for patients with chemotherapeutic‐resistant lung cancer with CAFs. [ABSTRACT FROM AUTHOR]

Radiation-induced pulmonary fibrosis is a serious complication. Plasminogen activator inhibitor-1 (PAI-1) has been indicated to be a key factor in the progression of pulmonary fibrosis. In the present study, the effect of PAI-1 deficiency on radiation-induced pulmonary fibrosis was analyzed. Wild-type (WT) and PAI-1-deficient (PAI-1-/-) mice were treated with thoracic irradiation of 15 Gy to induce pulmonary fibrosis. Analyses of bronchoalveolar lavage (BAL) fluids were performed 0, 4, 12, 18, and 24 weeks after irradiation. The degree of pulmonary fibrosis was assessed according to the histology of lung tissues and hydroxyproline contents. The results demonstrated that the irradiation of WT mice increased PAI-1 expression in the lungs after 18 weeks and established lung fibrosis at 24 weeks. The number of total cells and transforming growth factor-β levels in BAL fluid were significantly lower at 24 weeks after irradiation in PAI-1-/- mice compared with WT mice. Furthermore, histological examination revealed that the extent of pulmonary fibrosis was attenuated in PAI-1-/- mice compared with that in WT mice. Hydroxyproline content was also significantly lower in PAI-1-/- mice compared with WT mice at 24 weeks after irradiation. In conclusion, PAI-1 serves an important role in the development of radiation-induced pulmonary fibrosis and may represent a novel therapeutic target for pulmonary fibrosis. [ABSTRACT FROM AUTHOR]

Transforming growth factor-β (TGF-β) is central during the pathogenesis of pulmonary fibrosis, in which the plasminogen activator inhibitor-1 (PAI-1) also has an established role. TGF-β is also known to be the strongest inducer of PAI-1. To investigate the link between PAI-1 and TGF-β in fibrotic processes, we evaluated the effect of SK-216, a PAI-1-specific inhibitor, in TGF-β-dependent epithelial-mesenchymal transition (EMT) and fibroblast to myofibroblast differentiation. In human alveolar epithelial A549 cells, treatment with TGF-β induced EMT, whereas co-treatment with SK-216 attenuated the occurrence of EMT. The inhibition of TGF-β-induced EMT by SK-216 was also confirmed in the experiment using murine epithelial LA-4 cells. Blocking EMT by SK-216 inhibited TGF-β-induced endogenous production of PAI-1 and TGF-β in A549 cells as well. These effects of SK-216 were not likely mediated by suppressing either Smad or ERK pathways. Using human lung fibroblast MRC-5 cells, we demonstrated that SK-216 inhibited TGF-β-dependent differentiation of fibroblasts to myofibroblasts. We also observed this inhibition by SK-216 in human primary lung fibroblasts. Following these in vitro results, we tested oral administration of SK-216 into mice injected intratracheally with bleomycin. We found that SK-216 reduced the degree of bleomycin-induced pulmonary fibrosis in mice. Although the precise mechanisms underlying the link between TGF-β and PAI-1 regarding fibrotic process were not determined, PAI-1 seems to act as a potent downstream effector on the pro-fibrotic property of TGF-β. In addition, inhibition of PAI-1 activity by a PAI-1 inhibitor exerts an antifibrotic effect even in vivo. These data suggest that targeting PAI-1 as a downstream effector of TGF-β could be a promising therapeutic strategy for pulmonary fibrosis. [ABSTRACT FROM AUTHOR]

Found in inflammatory zone (FIZZ) 2, also known as resistin-like molecule (RELM)-β, belongs to a novel cysteine-rich secreted protein family named FIZZ/RELM. Its function is unclear, but a closely related family member, FIZZ1, has profibrotic activities. The human ortholog of rodent FIZZ1 has not been identified, but human FIZZ2 has significant sequence homology to both rodent FIZZ2 (59%) and FIZZ1 (50%). Given the greater homology to rodent FIZZ2, analyzing the role of FIZZ2 in a rodent model of bleomycin-induced pulmonary fibrosis would be of greater potential relevance to human fibrotic lung disease. The results showed that FIZZ2 was highly induced in lungs of rodents with bleomycin-induced pulmonary fibrosis and of human patients with idiopathic pulmonary fibrosis. FIZZ2 expression was induced in rodent and human lung epithelial cells by Th2 cytokines, which was mediated via STAT6 signaling. The FIZZ2 induction in murine lungs was found to be essential for pulmonary fibrosis, as FIZZ2 deficiency significantly suppressed pulmonary fibrosis and associated enhanced extracellular matrix and cytokine gene expression. In vitro analysis indicated that FIZZ2 could stimulate type I collagen and a-smooth muscle actin expression in lung fibroblasts. Furthermore, FIZZ2 was shown to have chemoattractant activity for bone marrow (BM) cells, especially BM-derived CD11c+ dendritic cells. Notably, lung recruitment of BM-derived cells was impaired in FIZZ2 knockout mice. These findings suggest that FIZZ2 is a Th2-associated multifunctional mediator with potentially important roles in the pathogenesis of fibrotic lung diseases. [ABSTRACT FROM AUTHOR]