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The 9th biennial conference titled “Stem Cells, Cell Therapies, and Bioengineering in Lung Biology and Diseases” was hosted virtually, due to the ongoing COVID-19 pandemic, in collaboration with the University of Vermont Larner College of Medicine, the National Heart, Lung, and Blood Institute, the Alpha-1 Foundation, the Cystic Fibrosis Foundation, and the International Society for Cell & Gene Therapy. The event was held from July 12th through 15th, 2021 with a pre-conference workshop held on July 9th. As in previous years, the objectives remained to review and discuss the status of active research areas involving stem cells (SCs), cellular therapeutics, and bioengineering as they relate to the human lung. Topics included 1) technological advancements in the in situ analysis of lung tissues, 2) new insights into stem cell signaling and plasticity in lung remodeling and regeneration, 3) the impact of extracellular matrix in stem cell regulation and airway engineering in lung regeneration, 4) differentiating and delivering stem cell therapeutics to the lung, 5) regeneration in response to viral infection, and 6) ethical development of cell-based treatments for lung diseases. This selection of topics represents some of the most dynamic and current research areas in lung biology. The virtual workshop included active discussion on state-of-the-art methods relating to the core features of the 2021 conference, including in situ proteomics, lung-on-chip, induced pluripotent stem cell (iPSC)-airway differentiation, and light sheet microscopy. The conference concluded with an open discussion to suggest funding priorities and recommendations for future research directions in basic and translational lung biology.

PDF file - 381K, The phenotype of MDSC in the lung and spleen from lung cancer challenged mice (S1); Reduced tumor burden in mice from Gem and SODmim+Gem therapy groups (S2); Combination therapy reduced infiltration of neutrophils but not macrophages in tumor (S3); Combination therapy was efficient in reducing tumor growth in catalase deficient mice by decreasing MDSC infiltration and improving the memory CD8+ T cell response (S4); Persistent memory CD8+ T cell subsets expand following re-encounter with i.v. challenged LLC cells (S5); Adoptively transferred memory cells persist and reduce MDSC infiltration and tumor burden in LLC-rechallenged recipient mice (S6); Adoptively transferred memory cells expand upon encounter with tumor cells (S7).

PDF file - 18K, Antibodies used for characterization of Immune Cell Phenotypes.

Increased apoptosis sensitivity of alveolar type 2 (ATII) cells and increased apoptosis resistance of (myo)fibroblasts, the apoptosis paradox, contributes to the pathogenesis of idiopathic pulmonary fibrosis (IPF). The mechanism underlying the apoptosis paradox in IPF lungs, however, is unclear. Aging is the greatest risk factor for IPF. In this study, we show, for the first time, that ATII cells from old mice are more sensitive, whereas fibroblasts from old mice are more resistant, to apoptotic challenges, compared with the corresponding cells from young mice. The expression of plasminogen activator inhibitor 1 (PAI-1), an important profibrogenic mediator, was significantly increased in both ATII cells and lung fibroblasts from aged mice. In vitro studies using PAI-1 siRNA and active PAI-1 protein indicated that PAI-1 promoted ATII cell apoptosis but protected fibroblasts from apoptosis, likely through dichotomous regulation of p53 expression. Deletion of PAI-1 in adult mice led to a reduction in p53, p21, and Bax protein expression, as well as apoptosis sensitivity in ATII cells, and their increase in the lung fibroblasts, as indicated by in vivo studies. This increase was associated with an attenuation of lung fibrosis after bleomycin challenge. Since PAI-1 is up-regulated in both ATII cells and fibroblasts in IPF, the results suggest that increased PAI-1 may underlie the apoptosis paradox of ATII cells and fibroblasts in IPF lungs.

Mitochondrial dysfunction has been associated with age-related diseases, including idiopathic pulmonary fibrosis (IPF). We provide evidence that implicates chronic elevation of the mitochondrial anion carrier protein, uncoupling protein-2 (UCP2), in increased generation of reactive oxygen species, altered redox state and cellular bioenergetics, impaired fatty acid oxidation, and induction of myofibroblast senescence. This pro-oxidant senescence reprogramming occurs in concert with conventional actions of UCP2 as an uncoupler of oxidative phosphorylation with dissipation of the mitochondrial membrane potential. UCP2 is highly expressed in human IPF lung myofibroblasts and in aged fibroblasts. In an aging murine model of lung fibrosis, the in vivo silencing of UCP2 induces fibrosis regression. These studies indicate a pro-fibrotic function of UCP2 in chronic lung disease and support its therapeutic targeting in age-related diseases associated with impaired tissue regeneration and organ fibrosis.

Rationale and objectivesThe role of human lung megakaryocytes in homeostasis and their dynamics in disease states remain unknown. We sought to investigate whether megakaryocyte/platelet gene signatures are altered in IPF.MethodsWe analyzed publicly available transcriptome datasets of lung tissue, bronchoalveolar lavage (BAL) cells, and peripheral whole blood from IPF patients and healthy controls. Enrichment of megakaryocyte and platelet gene signatures in those datasets were estimated using xCell, a novel computational method. Furthermore, we analyzed whether mean platelet volume (MPV) and platelet counts in peripheral blood are associated with lung transplant-free survival in our IPF cohort.ResultsIn lung tissue, megakaryocyte gene signature enrichment scores were significantly lower in IPF than in controls. In BAL cells, platelet gene signature enrichment scores were significantly lower in IPF than in controls, and lower platelet scores were associated with lower lung transplant-free survival in IPF. In contrast, in blood, megakaryocyte scores were significantly higher in IPF than in controls, and higher megakaryocyte scores were associated with lower disease progression-free survival in IPF. Furthermore, higher MPV was associated with lower transplant-free survival in our IPF cohort, independent of age, sex, forced vital capacity (FVC), and diffusing capacity of the lung for carbon monoxide (DLCO).ConclusionsIn IPF, megakaryocyte/platelet gene signatures were altered in a compartment-specific manner. Moreover, those signatures and MPV in blood were associated with important clinical outcomes such as transplant-free survival. These findings provide new insights into altered megakaryocyte/platelet biogenesis in IPF and suggest the potential utility of megakaryocyte/platelet-based biomarkers in IPF.

Senescence of alveolar type II (ATII) cells, progenitors of the alveolar epithelium, is a pathological feature and contributes importantly to the pathogenesis of idiopathic pulmonary fibrosis. Despite recognition of the importance of ATII cell senescence in idiopathic pulmonary fibrosis pathogenesis, how ATII cell senescence is regulated and how senescent ATII cells contribute to lung fibrogenesis remain unclear. In this study, we show that TGF-β1 (transforming growth factor-β1), a most ubiquitous and potent profibrotic cytokine, induces plasminogen activator inhibitor-1 (PAI-1), a cell senescence and fibrosis mediator, and p16 as well as senescence, but not apoptosis, in primary mouse ATII cells. We also found that senescent ATII cells secrete various cytokines and chemokines, including IL-4 and IL-13, which stimulate the expression of genes associated with a profibrotic phenotype in alveolar macrophages. Similar responses were also observed in TGF-β1-treated rat ATII (L2) and rat macrophage NR8383 cells. Deletion of PAI-1 or inhibition of PAI-1 activity with a small molecule PAI-1 inhibitor, however, blocks TGF-β1-induced senescence as well as a senescence-associated secretory phenotype in ATII and L2 cells and, consequently, the stimulatory effects of the conditioned medium from senescent ATII/L2 cells on macrophages. Moreover, we show that silencing p16 ameliorates PAI-1 protein-induced ATII cell senescence and secretion of profibrotic mediators. Our data suggest that PAI-1 mediates TGF-β1-induced ATII cell senescence and secretion of profibrotic mediators through inducing p16, and they also suggest that senescent ATII cells contribute to lung fibrogenesis in part by activating alveolar macrophages through secreting profibrotic and proinflammatory mediators.

Metabolic and bioenergetic plasticity of immune cells is essential for optimal responses to bacterial infections. AMPK and Parkin ubiquitin ligase are known to regulate mitochondrial quality control mitophagy that prevents unwanted inflammatory responses. However, it is not known if this evolutionarily conserved mechanism has been coopted by the host immune defense to eradicate bacterial pathogens and influence post-sepsis immunosuppression. Parkin, AMPK levels, and the effects of AMPK activators were investigated in human leukocytes from sepsis survivors as well as wild type and Park2−/− murine macrophages. In vivo, the impact of AMPK and Parkin was determined in mice subjected to polymicrobial intra-abdominal sepsis and secondary lung bacterial infections. Mice were treated with metformin during established immunosuppression. We showed that bacteria and mitochondria share mechanisms of autophagic killing/clearance triggered by sentinel events that involve depolarization of mitochondria and recruitment of Parkin in macrophages. Parkin-deficient mice/macrophages fail to form phagolysosomes and kill bacteria. This impairment of host defense is seen in the context of sepsis-induced immunosuppression with decreased levels of Parkin. AMPK activators, including metformin, stimulate Parkin-independent autophagy and bacterial killing in leukocytes from post-shock patients and in lungs of sepsis-immunosuppressed mice. Our results support a dual role of Parkin and AMPK in the clearance of dysfunctional mitochondria and killing of pathogenic bacteria, and explain the immunosuppressive phenotype associated Parkin and AMPK deficiency. AMPK activation appeared to be a crucial therapeutic target for the macrophage immunosuppressive phenotype and to reduce severity of secondary bacterial lung infections and respiratory failure.

Fibrosis is a leading cause of morbidity and mortality worldwide. Although fibrosis may involve different organ systems, transforming growth factor-β (TGFβ) has been established as a master regulator of fibrosis across organs. Pirfenidone and Nintedanib are the only currently-approved drugs to treat fibrosis, specifically idiopathic pulmonary fibrosis, but their mechanisms of action remain poorly understood. To identify novel drug targets and uncover potential mechanisms by which these drugs attenuate fibrosis, we performed an integrative 'omics analysis of transcriptomic and proteomic responses to TGFβ1-stimulated lung fibroblasts. Significant findings were annotated as associated with pirfenidone and nintedanib treatment in silico via Coremine. Integrative 'omics identified a co-expressed transcriptomic and proteomic module significantly correlated with TGFβ1 treatment that was enriched (FDR-p = 0.04) with genes associated with pirfenidone and nintedanib treatment. While a subset of genes in this module have been implicated in fibrogenesis, several novel TGFβ1 signaling targets were identified. Specifically, four genes (BASP1, HSD17B6, CDH11, and TNS1) have been associated with pirfenidone, while five genes (CLINT1, CADM1, MTDH, SYDE1, and MCTS1) have been associated with nintedanib, and MYDGF has been implicated with treatment using both drugs. Using the Clue Drug Repurposing Hub, succinic acid was highlighted as a metabolite regulated by the protein encoded by HSD17B6. This study provides new insights into the anti-fibrotic actions of pirfenidone and nintedanib and identifies novel targets for future mechanistic studies.

Multicellular organisms maintain structure and function of tissues/organs through emergent, self-organizing behavior. In this report, we demonstrate a critical role for lung mesenchymal stromal cell (L-MSC) aging in determining the capacity to form three-dimensional organoids or ‘alveolospheres’ with type 2 alveolar epithelial cells (AEC2s). In contrast to L-MSCs from aged mice, young L-MSCs support the efficient formation of alveolospheres when co-cultured with young or aged AEC2s. Aged L-MSCs demonstrated features of cellular senescence, altered bioenergetics, and a senescence-associated secretory profile (SASP). The reactive oxygen species generating enzyme, NADPH oxidase 4 (Nox4), was highly activated in aged L-MSCs and Nox4 downregulation was sufficient to, at least partially, reverse this age-related energy deficit, while restoring the self-organizing capacity of alveolospheres. Together, these data indicate a critical role for cellular bioenergetics and redox homeostasis in an organoid model of self-organization and support the concept of thermodynamic entropy in aging biology., eLife digest Many tissues in the body are capable of regenerating by replacing defective or worn-out cells with new ones. This process relies heavily on stem cells, which are precursor cells that lack a set role in the body and can develop into different types of cells under the right conditions. Tissues often have their own pool of stem cells that they use to replenish damaged cells. But as we age, this regeneration process becomes less effective. Many of our organs, such as the lungs, are lined with epithelial cells. These cells form a protective barrier, controlling what substances get in and out of the tissue. Alveoli are parts of the lungs that allow oxygen and carbon dioxide to move between the blood and the air in the lungs. And alveoli rely on an effective epithelial cell lining to work properly. To replenish these epithelial cells, alveoli have pockets, in which a type of epithelial cell, known as AEC2, lives. These cells can serve as stem cells, developing into a different type of cell under the right conditions. To work properly, AEC2 cells require close interactions with another type of cell called L-MSC, which supports the maintenance of other cells and also has the ability to differentiate into several other cell types. Both cell types can be found close together in these stem cell pockets. So far, it has been unclear how aging affects how these cells work together to replenish the epithelial lining of the alveoli. To investigate, Chanda et al. probed AEC2s and L-MSCs in the alveoli of young and old mice. The researchers collected both cell types from young (2-3 months) and aged (22-24 months) mice. Various combinations of these cells were grown to form 3D structures, mimicking how the cells grow in the lungs. Young L-MSCs formed normal 3D structures with both young and aged AEC2 cells. But aged L-MSCs developed abnormal, loose structures with AEC2 cells (both young and old cells). Aged L-MSCs were found to have higher levels of an enzyme (called Nox4) that produces oxidants and other ‘pro-aging’ factors, compared to young L-MSCs. However, reducing Nox4 levels in aged L-MSCs allowed these cells to form normal 3D structures with young AEC2 cells, but not aged AEC2 cells. These findings highlight the varying effects specific stem cells have, and how their behaviour is affected by pro-aging factors. Moreover, the pro-aging enzyme Nox4 shows potential as a therapeutic target – downregulating its activity may reverse critical effects of aging in cells.

Aging is a risk factor for progressive fibrotic disorders involving diverse organ systems, including the lung. Idiopathic pulmonary fibrosis, an age-associated degenerative lung disorder, is characterized by persistence of apoptosis-resistant myofibroblasts. Here we demonstrate that sirtuin 3 (SIRT3), a mitochondrial deacetylase, is downregulated in the lungs of humans with idiopathic pulmonary fibrosis and in mice subjected to lung injury. Overexpression of Sirt3 cDNA via airway delivery restored the capacity for fibrosis resolution in aged mice, in association with activation of the forkhead box transcription factor FoxO3a in fibroblasts, upregulation of pro-apoptotic members of the Bcl2 family and recovery of apoptosis susceptibility. While transforming growth factor-β1 reduced levels of SIRT3 and FOXO3A in lung fibroblasts, cell non-autonomous effects involving macrophage-secreted products were necessary for SIRT3-mediated activation of FOXO3A. Together, these findings reveal a novel role of SIRT3 in pro-resolution macrophage functions that restore susceptibility to apoptosis in fibroblasts via a FOXO3A-dependent mechanism. Using a mouse model of lung fibrosis, the authors find that overexpression of SIRT3 in myofibroblasts lowers their threshold for apoptosis, allowing their clearance from tissues and restoration of the lost capacity for fibrosis resolution in aged mice.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive fibrosing interstitial pneumonia that mainly affects the elderly. Several reports have demonstrated that aging is involved in the underlying pathogenic mechanisms of IPF. α-Klotho (KL) has been well characterized as an “age-suppressing” hormone and can provide protection against cellular senescence and oxidative stress. In this study, KL levels were assessed in human plasma and primary lung fibroblasts from patients with idiopathic pulmonary fibrosis (IPF-FB) and in lung tissue from mice exposed to bleomycin, which showed significant downregulation when compared with controls. Conversely, transgenic mice overexpressing KL were protected against bleomycin-induced lung fibrosis. Treatment of human lung fibroblasts with recombinant KL alone was not sufficient to inhibit transforming growth factor-β (TGF-β)-induced collagen deposition and inflammatory marker expression. Interestingly, fibroblast growth factor 23 (FGF23), a proinflammatory circulating protein for which KL is a coreceptor, was upregulated in IPF and bleomycin lungs. To our surprise, FGF23 and KL coadministration led to a significant reduction in fibrosis and inflammation in IPF-FB; FGF23 administration alone or in combination with KL stimulated KL upregulation. We conclude that in IPF downregulation of KL may contribute to fibrosis and inflammation and FGF23 may act as a compensatory antifibrotic and anti-inflammatory mediator via inhibition of TGF-β signaling. Upon restoration of KL levels, the combination of FGF23 and KL leads to resolution of inflammation and fibrosis. Altogether, these data provide novel insight into the FGF23/KL axis and its antifibrotic/anti-inflammatory properties, which opens new avenues for potential therapies in aging-related diseases like IPF.

Idiopathic pulmonary fibrosis (IPF) is one of many clinical syndromes that are associated with aging, and is increasing in both incidence and prevalence with the rapid rise in aging populations world-wide. There is accumulating data on how the biology of aging may influence the susceptibility to lung fibrosis in the elderly. In this review, we explore some of the known "hallmarks of aging," including telomere attrition, genomic instability, epigenetic alterations, loss of proteostasis, cellular senescence and mitochondrial dysfunction in the pathobiology of IPF. Additionally, we discuss age-associated alterations in extracellular matrix that may contribute to the development and/or progression of IPF.

Idiopathic pulmonary fibrosis (IPF) is a progressive and terminal lung disease with no known cure. IPF is a disease of aging, with median age of diagnosis over 65 years. Median survival is between 3 and 5 years after diagnosis. IPF is characterized primarily by excessive deposition of extracellular matrix (ECM) proteins by activated lung fibroblasts and myofibroblasts, resulting in reduced gas exchange and impaired pulmonary function. Growing evidence supports the concept of a pro-fibrotic environment orchestrated by underlying factors such as genetic predisposition, chronic injury and aging, oxidative stress, and impaired regenerative responses may account for disease development and persistence. Currently, two FDA approved drugs have limited efficacy in the treatment of IPF. Many of the genes and gene networks associated with lung development are induced or activated in IPF. In this review, we analyze current knowledge in the field, gained from both basic and clinical research, to provide new insights into the disease process, and potential approaches to treatment of pulmonary fibrosis.

Fibrosis is a dynamic process with the potential for reversibility and restoration of near-normal tissue architecture and organ function. Herein, we review mechanisms for resolution of organ fibrosis, in particular that involving the lung, with an emphasis on the critical roles of myofibroblast apoptosis and clearance of deposited matrix.

Our understanding of myofibroblast biology has advanced over the past two decades, and the seemingly antagonistic roles of these cells in both normal tissue repair and fibrotic diseases are better reconciled. An age-related loss of cellular plasticity that results in impaired capacity for de-differentiation and apoptosis susceptibility may predispose individuals to non-resolving and progressive fibrotic disorders involving diverse organ systems.

Tumor-stromal interactions within the tumor microenvironment (TME) influence lung cancer progression and response to therapeutic interventions, yet traditional in vitro studies fail to replicate the complexity of these interactions. Herein, we developed three-dimensional (3D) lung tumor models that mimic the human TME and demonstrate tumor-stromal crosstalk mediated by extracellular vesicles (EVs). EVs released by tumor cells, independent of p53 status, and fibroblasts within the TME mediate immunomodulatory effects; specifically, monocyte/macrophage polarization to a tumor-promoting M2 phenotype within this 3D-TME. Additionally, immune checkpoint inhibition in a 3D model that included T cells showed an inhibition of tumor growth and reduced hypoxia within the TME. Thus, perfused 3D tumor models incorporating diverse cell types provide novel insights into EV-mediated tumor-immune interactions and immune-modulation for existing and emerging cancer therapies.

The mechanisms by which environmental exposures contribute to the pathogenesis of lung fibrosis are unclear. Here, we demonstrate an increase in cadmium (Cd) and carbon black (CB), common components of cigarette smoke (CS) and environmental particulate matter (PM), in lung tissue from subjects with idiopathic pulmonary fibrosis (IPF). Cd concentrations were directly proportional to citrullinated vimentin (Cit-Vim) amounts in lung tissue of subjects with IPF. Cit-Vim amounts were higher in subjects with IPF, especially smokers, which correlated with lung function and were associated with disease manifestations. Cd/CB induced the secretion of Cit-Vim in an Akt1- and peptidylarginine deiminase 2 (PAD2)–dependent manner. Cit-Vim mediated fibroblast invasion in a 3D ex vivo model of human pulmospheres that resulted in higher expression of CD26, collagen, and α-SMA. Cit-Vim activated NF-κB in a TLR4-dependent fashion and induced the production of active TGF-β1, CTGF, and IL-8 along with higher surface expression of TLR4 in lung fibroblasts. To corroborate ex vivo findings, mice treated with Cit-Vim, but not Vim, independently developed a similar pattern of fibrotic tissue remodeling, which was TLR4 dependent. Moreover, wild-type mice, but not PAD2(−/−) and TLR4 mutant (MUT) mice, exposed to Cd/CB generated high amounts of Cit-Vim, in both plasma and bronchoalveolar lavage fluid, and developed lung fibrosis in a stereotypic manner. Together, these studies support a role for Cit-Vim as a damage-associated molecular pattern molecule (DAMP) that is generated by lung macrophages in response to environmental Cd/CB exposure. Furthermore, PAD2 might represent a promising target to attenuate Cd/CB-induced fibrosis.

Aging is associated with the development of human idiopathic pulmonary fibrosis (IPF), a lethal fibrotic lung disease. In this study, the investigators identify a novel molecular target with therapeutic potential against age-related persistent lung fibrosis in mice., Aging is a strong risk factor and an independent prognostic factor for progressive human idiopathic pulmonary fibrosis (IPF). Aged mice develop nonresolving pulmonary fibrosis following lung injury. In this study, we found that mouse double minute 4 homolog (MDM4) is highly expressed in the fibrotic lesions of human IPF and experimental pulmonary fibrosis in aged mice. We identified MDM4 as a matrix stiffness–regulated endogenous inhibitor of p53. Reducing matrix stiffness down-regulates MDM4 expression, resulting in p53 activation in primary lung myofibroblasts isolated from IPF patients. Gain of p53 function activates a gene program that sensitizes lung myofibroblasts to apoptosis and promotes the clearance of apoptotic myofibroblasts by macrophages. Destiffening of the fibrotic lung matrix by targeting nonenzymatic cross-linking or genetic ablation of Mdm4 in lung (myo)fibroblasts activates the Mdm4–p53 pathway and promotes lung fibrosis resolution in aged mice. These findings suggest that mechanosensitive MDM4 is a molecular target with promising therapeutic potential against persistent lung fibrosis associated with aging., Graphical Abstract

Multicellular organisms maintain structure and function of tissues/organs through emergent, self-organizing behavior. In this report, we demonstrate a critical role for lung mesenchymal stromal cell (L-MSC) aging in determining the capacity to form 3-dimentional organoids or “alveolospheres “ with type 2 alveolar epithelial cells (AEC2s). In contrast to L-MSCs from aged mice, young L-MSCs support the efficient formation of alveolospheres when co-cultured with young or aged AEC2s. Aged L-MSCs demonstrated features of cellular senescence, altered bioenergetics, and a senescence-associated secretory profile (SASP). The reactive oxygen species generating enzyme, NADPH oxidase 4 (Nox4), was highly activated in aged L-MSCs and Nox4 downregulation was sufficient to, at least partially, reverse this age-related energy deficit, while restoring the self-organizing capacity of alveolospheres. Together, these data indicate a critical role for cellular bioenergetics and redox homeostasis in an organoid model of self-organization, and supports the concept of thermodynamic entropy in aging biology.

Our understanding of myofibroblast biology has advanced over the past two decades, and the seemingly antagonistic roles of these cells in both normal tissue repair and fibrotic diseases are better reconciled. An age-related loss of cellular plasticity that results in impaired capacity for de-differentiation and apoptosis susceptibility may predispose individuals to non-resolving and progressive fibrotic disorders involving diverse organ systems.

The oxidation of tyrosine residues to generate o,o'-dityrosine cross-links in extracellular proteins is necessary for the proper function of the extracellular matrix (ECM) in various contexts in invertebrates. Tyrosine oxidation is also required for the biosynthesis of thyroid hormone in vertebrates, and there is evidence for oxidative cross-linking reactions occurring in extracellular proteins secreted by myofibroblasts. The ECM protein fibronectin circulates in the blood as a globular protein that dimerizes through disulfide bridges generated by cysteine oxidation. We found that cellular (fibrillar) fibronectin on the surface of transforming growth factor-β1 (TGF-β1)-activated human myofibroblasts underwent multimerization by o,o'-dityrosine cross-linking under reducing conditions that disrupt disulfide bridges, but soluble fibronectin did not. This reaction on tyrosine residues required both the TGF-β1-dependent production of hydrogen peroxide and the presence of myeloperoxidase (MPO) derived from inflammatory cells, which are active participants in wound healing and fibrogenic processes. Oxidative cross-linking of matrix fibronectin attenuated both epithelial and fibroblast migration and conferred resistance to proteolysis by multiple proteases. The abundance of circulating o,o'-dityrosine-modified fibronectin was increased in a murine model of lung fibrosis and in human subjects with interstitial lung disease compared to that in control healthy subjects. These studies indicate that tyrosine can undergo stable, covalent linkages in fibrillar fibronectin under inflammatory conditions and that this modification affects the migratory behavior of cells on such modified matrices, suggesting that this modification may play a role in both physiologic and pathophysiologic tissue repair.

Tissue regeneration capacity declines with aging in association with heightened oxidative stress. Expression of the oxidant-generating enzyme, NADPH oxidase 4 (Nox4), is elevated in aged mice with diminished capacity for fibrosis resolution. Bromodomain-containing protein 4 (Brd4) is a member of the bromodomain and extraterminal (BET) family of proteins that function as epigenetic “readers” of acetylated lysine groups on histones. In this study, we explored the role of Brd4 and its interaction with the p300 acetyltransferase in the regulation of Nox4 and the in vivo efficacy of a BET inhibitor to reverse established age-associated lung fibrosis. BET inhibition interferes with the association of Brd4, p300, and acetylated histone H4K16 with the Nox4 promoter in lung fibroblasts stimulated with the profibrotic cytokine, TGF-β1. A number of BET inhibitors, including I-BET-762, JQ1, and OTX015, downregulate Nox4 gene expression and activity. Aged mice with established and persistent lung fibrosis recover capacity for fibrosis resolution with OTX015 treatment. This study implicates epigenetic regulation of Nox4 by Brd4 and p300 and supports BET/Brd4 inhibition as an effective strategy for the treatment of age-related fibrotic lung disease.

Aging is associated with an increased risk of a number of clinical syndromes, including chronic lung disease. There have been significant advances in our understanding of the biology of aging leading to the elucidation of the so-called “hallmarks of aging.” The cause-effect relationships between various hallmarks such as dysregulated nutrient sensing, mitochondrial dysfunction, and cellular senescence are not well understood. Here, I discuss the evidence for alterations in energy/metabolic sensing pathways in the degenerative chronic lung disease called idiopathic pulmonary fibrosis (IPF). The pathobiological mechanisms by which this defect may contribute to age-related susceptibility to IPF and potentially other diseases of the elderly are also discussed.