Lung epithelial cell damage and dysfunctional repair play a role in the development of lung disease. Effective repair likely requires the normal functioning of alveolar stem/progenitor cells. For ...example, we have shown in a mouse model of bronchopulmonary dysplasia (BPD) that mesenchymal stem cells (MSC) protect against hyperoxic lung injury at least in part by increasing the number of Epcam
Sca-1
distal lung epithelial cells. These cells are capable of differentiating into both small airway (CCSP
) and alveolar (SPC
) epithelial cells in three-dimensional (3D) organoid cultures. To further understand the interactions between MSC and distal lung epithelial cells, we added MSC to lung progenitor 3D cultures. MSC stimulated Epcam
Sca-1
derived organoid formation, increased alveolar differentiation and decreased self-renewal. MSC-conditioned media was sufficient to promote alveolar organoid formation, demonstrating that soluble factors secreted by MSC are likely responsible for the response. This work provides strong evidence of a direct effect of MSC-secreted factors on lung progenitor cell differentiation.
Coronavirus disease 2019 (COVID-19) is the latest respiratory pandemic caused by severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2). Although infection initiates in the proximal ...airways, severe and sometimes fatal symptoms of the disease are caused by infection of the alveolar type 2 (AT2) cells of the distal lung and associated inflammation. In this study, we develop primary human lung epithelial infection models to understand initial responses of proximal and distal lung epithelium to SARS-CoV-2 infection. Differentiated air-liquid interface (ALI) cultures of proximal airway epithelium and alveosphere cultures of distal lung AT2 cells are readily infected by SARS-CoV-2, leading to an epithelial cell-autonomous proinflammatory response with increased expression of interferon signaling genes. Studies to validate the efficacy of selected candidate COVID-19 drugs confirm that remdesivir strongly suppresses viral infection/replication. We provide a relevant platform for study of COVID-19 pathobiology and for rapid drug screening against SARS-CoV-2 and emergent respiratory pathogens.
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•Human alveospheres are composed of renewing AT2 cells and AT1-like cells•Alveolar epithelial cells are efficiently infected by SARS-CoV-2 in vitro•Interferon signaling is activated in SARS-CoV-2-infected alveolar epithelial cells•Lung organoid models provide a platform for drug discovery and disease modeling
In vitro models of human lung epithelium, including diverse cell types of the proximo-distal axis, are critical for modeling infection. Mulay et al. show that alveospheres, with epithelial type 2- and type 1-like cells, are infected by SARS-CoV-2, initiating an interferon response, and serve as a platform for screening antiviral drugs.
The repair process of damaged tissue involves the coordinated activities of several cell types in response to local and systemic signals. Following acute tissue injury, infiltrating inflammatory ...cells and resident stem cells orchestrate their activities to restore tissue homeostasis. However, during chronic tissue damage, such as in muscular dystrophies, the inflammatory-cell infiltration and fibroblast activation persists, while the reparative capacity of stem cells (satellite cells) is attenuated. Abnormal dystrophic muscle repair and its end stage, fibrosis, represent the final common pathway of virtually all chronic neurodegenerative muscular diseases. As our understanding of the pathogenesis of muscle fibrosis has progressed, it has become evident that the muscle provides a useful model for the regulation of tissue repair by the local microenvironment, showing interplay among muscle-specific stem cells, inflammatory cells, fibroblasts and extracellular matrix components of the mammalian wound-healing response. This article reviews the emerging findings of the mechanisms that underlie normal versus aberrant muscle-tissue repair.
Fibrosis is the aberrant deposition of extracellular matrix (ECM) components during tissue healing leading to loss of its architecture and function. Fibrotic diseases are often associated with ...chronic pathologies and occur in a large variety of vital organs and tissues, including skeletal muscle. In human muscle, fibrosis is most readily associated with the severe muscle wasting disorder Duchenne muscular dystrophy (DMD), caused by loss of dystrophin gene function. In DMD, skeletal muscle degenerates and is infiltrated by inflammatory cells and the functions of the muscle stem cells (satellite cells) become impeded and fibrogenic cells hyperproliferate and are overactivated, leading to the substitution of skeletal muscle with nonfunctional fibrotic tissue. Here, we review new developments in our understanding of the mechanisms leading to fibrosis in DMD and several recent advances towards reverting it, as potential treatments to attenuate disease progression.
Recent genomic and scRNA-seq analyses of melanoma demonstrated a lack of recurrent genetic drivers of metastasis, while identifying common transcriptional states correlating with invasion or drug ...resistance. To test whether transcriptional adaptation can drive melanoma progression, we made use of a zebrafish mitfa:
;
model, in which malignant progression is characterized by minimal genetic evolution. We undertook an overexpression-screen of 80 epigenetic/transcriptional regulators and found neural crest-mesenchyme developmental regulator SATB2 to accelerate aggressive melanoma development. Its overexpression induces invadopodia formation and invasion in zebrafish tumors and human melanoma cell lines. SATB2 binds and activates neural crest-regulators, including
and
. The transcriptional program induced by SATB2 overlaps with known MITF
AXL
and AQP1
NGFR1
drug-resistant states and functionally drives enhanced tumor propagation and resistance to Vemurafenib in vivo. In summary, we show that melanoma transcriptional rewiring by SATB2 to a neural crest mesenchyme-like program can drive invasion and drug resistance in autochthonous tumors.
Inactivation of SMARCA4/BRG1, the core ATPase subunit of mammalian SWI/SNF complexes, occurs at very high frequencies in non-small cell lung cancers (NSCLC). There are no targeted therapies for this ...subset of lung cancers, nor is it known how mutations in
contribute to lung cancer progression. Using a combination of gain- and loss-of-function approaches, we demonstrate that deletion of BRG1 in lung cancer leads to activation of replication stress responses. Single-molecule assessment of replication fork dynamics in BRG1-deficient cells revealed increased origin firing mediated by the prelicensing protein, CDC6. Quantitative mass spectrometry and coimmunoprecipitation assays showed that BRG1-containing SWI/SNF complexes interact with RPA complexes. Finally, BRG1-deficient lung cancers were sensitive to pharmacologic inhibition of ATR. These findings provide novel mechanistic insight into BRG1-mutant lung cancers and suggest that their dependency on ATR can be leveraged therapeutically and potentially expanded to BRG1-mutant cancers in other tissues. SIGNIFICANCE: These findings indicate that inhibition of ATR is a promising therapy for the 10% of non-small cell lung cancer patients harboring mutations in SMARCA4/BRG1. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/18/3841/F1.large.jpg.
Preservation of cell identity is necessary for homeostasis of most adult tissues. This process is challenged every time a tissue undergoes regeneration after stress or injury. In the lethal Duchenne ...muscular dystrophy (DMD), skeletal muscle regenerative capacity declines gradually as fibrosis increases. Using genetically engineered tracing mice, we demonstrate that, in dystrophic muscle, specialized cells of muscular, endothelial, and hematopoietic origins gain plasticity toward a fibrogenic fate via a TGFβ-mediated pathway. This results in loss of cellular identity and normal function, with deleterious consequences for regeneration. Furthermore, this fibrogenic process involves acquisition of a mesenchymal progenitor multipotent status, illustrating a link between fibrogenesis and gain of progenitor cell functions. As this plasticity also was observed in DMD patients, we propose that mesenchymal transitions impair regeneration and worsen diseases with a fibrotic component.
•Regeneration declines as fibrosis increases in Duchenne muscular dystrophy (DMD)•Myogenic, endothelial, and hematopoietic cells acquire fibrogenic plasticity in DMD•This fibrogenic plasticity in DMD is induced by TGFβ and blunts muscle regeneration•Fibrogenesis occurs through an intermediate mesenchymal progenitor multipotent state
The cellular pathways driving fibrosis in dystrophic skeletal muscle are largely unknown. Muñoz-Cánoves and colleagues found that a significant number of muscle stem cells, endothelial cells, and hematopoietic cells gain plasticity as Duchenne muscular dystrophy (DMD) progresses, and convert into fibrotic collagen-producing cells, with negative consequences on regeneration. An association between fibrogenesis and mesenchymal transitions in DMD also is demonstrated.
Fibrosis, an excessive collagen accumulation, results in scar formation, impairing function of vital organs and tissues. Fibrosis is a hallmark of muscular dystrophies, including the lethal Duchenne ...muscular dystrophy (DMD), which remains incurable. Substitution of muscle by fibrotic tissue also complicates gene/cell therapies for DMD. Yet, no optimal models to study muscle fibrosis are available. In the widely used mdx mouse model for DMD, extensive fibrosis develops in the diaphragm only at advanced adulthood, and at about two years of age in the 'easy-to-access' limb muscles, thus precluding fibrosis research and the testing of novel therapies.
We developed distinct experimental strategies, ranging from chronic exercise to increasing muscle damage on limb muscles of young mdx mice, by myotoxin injection, surgically induced trauma (laceration or denervation) or intramuscular delivery of profibrotic growth factors (such as TGFβ). We also extended these approaches to muscle of normal non-dystrophic mice.
These strategies resulted in advanced and enhanced muscle fibrosis in young mdx mice, which persisted over time, and correlated with reduced muscle force, thus mimicking the severe DMD phenotype. Furthermore, increased fibrosis was also obtained by combining these procedures in muscles of normal mice, mirroring aberrant repair after severe trauma.
We have developed new and improved experimental strategies to accelerate and enhance muscle fibrosis in vivo. These strategies will allow rapidly assessing fibrosis in the easily accessible limb muscles of young mdx mice, without necessarily having to use old animals. The extension of these fibrogenic regimes to the muscle of non-dystrophic wild-type mice will allow fibrosis assessment in a wide array of pre-existing transgenic mouse lines, which in turn will facilitate understanding the mechanisms of fibrogenesis. These strategies should improve our ability to combat fibrosis-driven dystrophy progression and aberrant regeneration.
Dysfunction of alveolar epithelial type 2 cells (AEC2s), the facultative progenitors of lung alveoli, is implicated in pulmonary disease pathogenesis, highlighting the importance of human in vitro ...models. However, AEC2-like cells in culture have yet to be directly compared to their in vivo counterparts at single-cell resolution. Here, we performed head-to-head comparisons among the transcriptomes of primary (1°) adult human AEC2s, their cultured progeny, and human induced pluripotent stem cell-derived AEC2s (iAEC2s). We found each population occupied a distinct transcriptomic space with cultured AEC2s (1° and iAEC2s) exhibiting similarities to and differences from freshly purified 1° cells. Across each cell type, we found an inverse relationship between proliferative and maturation states, with preculture 1° AEC2s being most quiescent/mature and iAEC2s being most proliferative/least mature. Cultures of either type of human AEC2s did not generate detectable alveolar type 1 cells in these defined conditions; however, a subset of iAEC2s cocultured with fibroblasts acquired a transitional cell state described in mice and humans to arise during fibrosis or following injury. Hence, we provide direct comparisons of the transcriptomic programs of 1° and engineered AEC2s, 2 in vitro models that can be harnessed to study human lung health and disease.
Regeneration of muscle fibers, lost during pathological muscle degeneration or after injuries, is sustained by the production of new myofibers by means of the satellite cells. Survival of the ...satellite cells is a critical requirement for efficient muscle reconstitution. Necdin, a member of the MAGE proteins family, is expressed in satellite cell-derived myogenic precursors during perinatal growth and in the adult upon activation during muscle regeneration, where it plays an important role both in myoblast differentiation and survival. We show here that necdin exerts its pro-survival activity by counteracting the action of the pro-apoptotic protein Cell Cycle Apoptosis Regulatory Protein (CCAR1/CARP1) that we have identified as a new molecular interactor of necdin by two-hybrid screening. Necdin is responsible for the maintenance of CCAR1 protein levels, by implementing its ubiquitination and degradation through the proteasome. Taken together, these data shed new light on the molecular mechanism of necdin anti-apoptotic activity in myogenesis.
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