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Generation of Distal Airway Epithelium from Multipotent Human Foregut Stem Cells.

Hannan NR, Sampaziotis F, Segeritz CP, Hanley NA, Vallier L - Stem Cells Dev. (2015)

Bottom Line: Human foregut stem cells (hFSCs) represent an advantageous progenitor cell type that can be used to amplify large quantities of cells for regenerative medicine applications and can be derived from any human pluripotent stem cell line.This culture system can be applied to hFSCs carrying the CFTR mutation Δf508, enabling the development of an in vitro model for cystic fibrosis.This is the first demonstration that multipotent endoderm stem cells can differentiate not only into both liver and pancreatic cells but also into lung endoderm.

View Article: PubMed Central - PubMed

Affiliation: 1 Anne McLaren Laboratory for Regenerative Medicine, Department of Surgery, Wellcome Trust-Medical Research Council Stem Cell Institute, University of Cambridge , Cambridge, United Kingdom .

ABSTRACT
Collectively, lung diseases are one of the largest causes of premature death worldwide and represent a major focus in the field of regenerative medicine. Despite significant progress, only few stem cell platforms are currently available for cell-based therapy, disease modeling, and drug screening in the context of pulmonary disorders. Human foregut stem cells (hFSCs) represent an advantageous progenitor cell type that can be used to amplify large quantities of cells for regenerative medicine applications and can be derived from any human pluripotent stem cell line. Here, we further demonstrate the application of hFSCs by generating a near homogeneous population of early pulmonary endoderm cells coexpressing NKX2.1 and FOXP2. These progenitors are then able to form cells that are representative of distal airway epithelium that express NKX2.1, GATA6, and cystic fibrosis transmembrane conductance regulator (CFTR) and secrete SFTPC. This culture system can be applied to hFSCs carrying the CFTR mutation Δf508, enabling the development of an in vitro model for cystic fibrosis. This platform is compatible with drug screening and functional validations of small molecules, which can reverse the phenotype associated with CFTR mutation. This is the first demonstration that multipotent endoderm stem cells can differentiate not only into both liver and pancreatic cells but also into lung endoderm. Furthermore, our study establishes a new approach for the generation of functional lung cells that can be used for disease modeling as well as for drug screening and the study of lung development.

No MeSH data available.


Related in: MedlinePlus

The CFTR-Δf508 mis-folded receptor can be rescued to a functional state using the small molecule VX809. (A) Chloride influx and efflux trace in mature airway epithelial cells expressing the wild-type CFTR (red trace), the ΔF508 mutation in the CFTR (blue trace), or the ΔF508 mutation in the CFTR but treated with the small molecule VX809 (green trace). (B) Confocal microscopy showing expression of CFTR in matured airway epithelium from wild-type cells, cells with the ΔF508 mutation in the CFTR, and cells with the ΔF508 mutation in the CFTR but treated with the small molecule VX809. White bars=100 μM. Addition of Cl− or NO3− indicated with solid arrowheads.
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f4: The CFTR-Δf508 mis-folded receptor can be rescued to a functional state using the small molecule VX809. (A) Chloride influx and efflux trace in mature airway epithelial cells expressing the wild-type CFTR (red trace), the ΔF508 mutation in the CFTR (blue trace), or the ΔF508 mutation in the CFTR but treated with the small molecule VX809 (green trace). (B) Confocal microscopy showing expression of CFTR in matured airway epithelium from wild-type cells, cells with the ΔF508 mutation in the CFTR, and cells with the ΔF508 mutation in the CFTR but treated with the small molecule VX809. White bars=100 μM. Addition of Cl− or NO3− indicated with solid arrowheads.

Mentions: Finally, to characterize the potential of our system as a platform for validating novel therapeutic agents, we decided to assess the efficiency of the small molecule VX809 for restoring CFTR function in CF-hFSCs-derived AECs. Indeed, VX809 has been shown in several studies to correct folding defects in the mature CFTR protein allowing reduced degradation, increased trafficking to the cell surface, and restoration of chloride transport. CF-hFSCs-derived lung cells were incubated for 48 h with or without VX809 and then loaded with MQAE for detection of chloride transport. CF-hFSCs-derived AECs grown in the presence of VX809 showed a dramatic improvement in the capacity to transport chloride, confirming the efficiency of this drug to correct CF in vitro (Fig. 4A). Furthermore, confocal microscopy revealed that CFTR was able to accumulate to greater levels on VX809 treatment (Fig. 4B). Taken together, these data confirm the mode of action of VX809 and thus demonstrate that hFSCs-derived distal AECs could be used to predict the activity of small molecules to correct lung diseases in vitro.


Generation of Distal Airway Epithelium from Multipotent Human Foregut Stem Cells.

Hannan NR, Sampaziotis F, Segeritz CP, Hanley NA, Vallier L - Stem Cells Dev. (2015)

The CFTR-Δf508 mis-folded receptor can be rescued to a functional state using the small molecule VX809. (A) Chloride influx and efflux trace in mature airway epithelial cells expressing the wild-type CFTR (red trace), the ΔF508 mutation in the CFTR (blue trace), or the ΔF508 mutation in the CFTR but treated with the small molecule VX809 (green trace). (B) Confocal microscopy showing expression of CFTR in matured airway epithelium from wild-type cells, cells with the ΔF508 mutation in the CFTR, and cells with the ΔF508 mutation in the CFTR but treated with the small molecule VX809. White bars=100 μM. Addition of Cl− or NO3− indicated with solid arrowheads.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4499787&req=5

f4: The CFTR-Δf508 mis-folded receptor can be rescued to a functional state using the small molecule VX809. (A) Chloride influx and efflux trace in mature airway epithelial cells expressing the wild-type CFTR (red trace), the ΔF508 mutation in the CFTR (blue trace), or the ΔF508 mutation in the CFTR but treated with the small molecule VX809 (green trace). (B) Confocal microscopy showing expression of CFTR in matured airway epithelium from wild-type cells, cells with the ΔF508 mutation in the CFTR, and cells with the ΔF508 mutation in the CFTR but treated with the small molecule VX809. White bars=100 μM. Addition of Cl− or NO3− indicated with solid arrowheads.
Mentions: Finally, to characterize the potential of our system as a platform for validating novel therapeutic agents, we decided to assess the efficiency of the small molecule VX809 for restoring CFTR function in CF-hFSCs-derived AECs. Indeed, VX809 has been shown in several studies to correct folding defects in the mature CFTR protein allowing reduced degradation, increased trafficking to the cell surface, and restoration of chloride transport. CF-hFSCs-derived lung cells were incubated for 48 h with or without VX809 and then loaded with MQAE for detection of chloride transport. CF-hFSCs-derived AECs grown in the presence of VX809 showed a dramatic improvement in the capacity to transport chloride, confirming the efficiency of this drug to correct CF in vitro (Fig. 4A). Furthermore, confocal microscopy revealed that CFTR was able to accumulate to greater levels on VX809 treatment (Fig. 4B). Taken together, these data confirm the mode of action of VX809 and thus demonstrate that hFSCs-derived distal AECs could be used to predict the activity of small molecules to correct lung diseases in vitro.

Bottom Line: Human foregut stem cells (hFSCs) represent an advantageous progenitor cell type that can be used to amplify large quantities of cells for regenerative medicine applications and can be derived from any human pluripotent stem cell line.This culture system can be applied to hFSCs carrying the CFTR mutation Δf508, enabling the development of an in vitro model for cystic fibrosis.This is the first demonstration that multipotent endoderm stem cells can differentiate not only into both liver and pancreatic cells but also into lung endoderm.

View Article: PubMed Central - PubMed

Affiliation: 1 Anne McLaren Laboratory for Regenerative Medicine, Department of Surgery, Wellcome Trust-Medical Research Council Stem Cell Institute, University of Cambridge , Cambridge, United Kingdom .

ABSTRACT
Collectively, lung diseases are one of the largest causes of premature death worldwide and represent a major focus in the field of regenerative medicine. Despite significant progress, only few stem cell platforms are currently available for cell-based therapy, disease modeling, and drug screening in the context of pulmonary disorders. Human foregut stem cells (hFSCs) represent an advantageous progenitor cell type that can be used to amplify large quantities of cells for regenerative medicine applications and can be derived from any human pluripotent stem cell line. Here, we further demonstrate the application of hFSCs by generating a near homogeneous population of early pulmonary endoderm cells coexpressing NKX2.1 and FOXP2. These progenitors are then able to form cells that are representative of distal airway epithelium that express NKX2.1, GATA6, and cystic fibrosis transmembrane conductance regulator (CFTR) and secrete SFTPC. This culture system can be applied to hFSCs carrying the CFTR mutation Δf508, enabling the development of an in vitro model for cystic fibrosis. This platform is compatible with drug screening and functional validations of small molecules, which can reverse the phenotype associated with CFTR mutation. This is the first demonstration that multipotent endoderm stem cells can differentiate not only into both liver and pancreatic cells but also into lung endoderm. Furthermore, our study establishes a new approach for the generation of functional lung cells that can be used for disease modeling as well as for drug screening and the study of lung development.

No MeSH data available.


Related in: MedlinePlus