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Beta cells within single human islets originate from multiple progenitors.

Scharfmann R, Xiao X, Heimberg H, Mallet J, Ravassard P - PLoS ONE (2008)

Bottom Line: By performing gene transfer at low multiplicity of infection, we created a chimeric graft with a subpopulation of human beta cells expressing GFP and found both GFP-positive and GFP-negative beta cells within single islets.The detection of both labeled and unlabeled beta cells in single islets demonstrates that beta cells present in a human islet are derived from multiple progenitors thus providing the first dynamic analysis of human islet formation during development.This human transgenic-like tool can be widely used to elucidate dynamic genetic processes in human tissue formation.

View Article: PubMed Central - PubMed

Affiliation: University Paris-Descartes, Faculty of Medicine, INSERM, Necker Hospital, U845, Paris, France.

ABSTRACT

Background: In both humans and rodents, glucose homeostasis is controlled by micro-organs called islets of Langerhans composed of beta cells, associated with other endocrine cell types. Most of our understanding of islet cell differentiation and morphogenesis is derived from rodent developmental studies. However, little is known about human islet formation. The lack of adequate experimental models has restricted the study of human pancreatic development to the histological analysis of different stages of pancreatic development. Our objective was to develop a new experimental model to (i) transfer genes into developing human pancreatic cells and (ii) validate gene transfer by defining the clonality of developing human islets.

Methods and findings: In this study, a unique model was developed combining ex vivo organogenesis from human fetal pancreatic tissue and cell type-specific lentivirus-mediated gene transfer. Human pancreatic progenitors were transduced with lentiviruses expressing GFP under the control of an insulin promoter and grafted to severe combined immunodeficient mice, allowing human beta cell differentiation and islet morphogenesis. By performing gene transfer at low multiplicity of infection, we created a chimeric graft with a subpopulation of human beta cells expressing GFP and found both GFP-positive and GFP-negative beta cells within single islets.

Conclusion: The detection of both labeled and unlabeled beta cells in single islets demonstrates that beta cells present in a human islet are derived from multiple progenitors thus providing the first dynamic analysis of human islet formation during development. This human transgenic-like tool can be widely used to elucidate dynamic genetic processes in human tissue formation.

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Related in: MedlinePlus

Lentivirus-mediated gene transfer in human fetal pancreas.Human fetal pancreases were partially dissociated, transduced with lentiviruses expressing GFP under the control of the CMV promoter, grafted into scid mice and analyzed 10 days later. A double staining for Pdx1 (red) and GFP (green); B represent enlargements of A. Dotted arrows: infected fibroblastic-like cells. Arrows: Pdx1-positive cells. Arrow head: Pdx1–negative cells. C double staining for insulin (red) and GFP. Scale bars: A, C: 25 µm, B: 10 µm.
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pone-0003559-g003: Lentivirus-mediated gene transfer in human fetal pancreas.Human fetal pancreases were partially dissociated, transduced with lentiviruses expressing GFP under the control of the CMV promoter, grafted into scid mice and analyzed 10 days later. A double staining for Pdx1 (red) and GFP (green); B represent enlargements of A. Dotted arrows: infected fibroblastic-like cells. Arrows: Pdx1-positive cells. Arrow head: Pdx1–negative cells. C double staining for insulin (red) and GFP. Scale bars: A, C: 25 µm, B: 10 µm.

Mentions: To trace human fetal pancreatic progenitor cells, we first transduced partially dissociated fetal pancreases with replication-defective lentiviruses expressing eGFP under the control of the ubiquitous human cytomegalovirus (CMV) promoter, at low multiplicity of infection (MOI). Transduced pancreases were transplanted under the kidney capsule of scid mice. After 10 days, we observed eGFP-expressing fibroblastic-like cells (Fig 3 A, arrows). The rare scattered insulin-positive cells were found negative for eGFP (Fig. 3C). Epithelial Pdx1-positive (Fig 3 B, arrow) and Pdx1-negative (Fig 3 B, arrowhead) cells were observed in these grafts. As expected with low MOI, only 0.6% of Pdx1-positive cells expressed eGFP (123 out of 20,000 Pdx1-positive cells counted). Such low percentage enables us to determine whether human islet beta cells are derived from one or several progenitors.


Beta cells within single human islets originate from multiple progenitors.

Scharfmann R, Xiao X, Heimberg H, Mallet J, Ravassard P - PLoS ONE (2008)

Lentivirus-mediated gene transfer in human fetal pancreas.Human fetal pancreases were partially dissociated, transduced with lentiviruses expressing GFP under the control of the CMV promoter, grafted into scid mice and analyzed 10 days later. A double staining for Pdx1 (red) and GFP (green); B represent enlargements of A. Dotted arrows: infected fibroblastic-like cells. Arrows: Pdx1-positive cells. Arrow head: Pdx1–negative cells. C double staining for insulin (red) and GFP. Scale bars: A, C: 25 µm, B: 10 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2571119&req=5

pone-0003559-g003: Lentivirus-mediated gene transfer in human fetal pancreas.Human fetal pancreases were partially dissociated, transduced with lentiviruses expressing GFP under the control of the CMV promoter, grafted into scid mice and analyzed 10 days later. A double staining for Pdx1 (red) and GFP (green); B represent enlargements of A. Dotted arrows: infected fibroblastic-like cells. Arrows: Pdx1-positive cells. Arrow head: Pdx1–negative cells. C double staining for insulin (red) and GFP. Scale bars: A, C: 25 µm, B: 10 µm.
Mentions: To trace human fetal pancreatic progenitor cells, we first transduced partially dissociated fetal pancreases with replication-defective lentiviruses expressing eGFP under the control of the ubiquitous human cytomegalovirus (CMV) promoter, at low multiplicity of infection (MOI). Transduced pancreases were transplanted under the kidney capsule of scid mice. After 10 days, we observed eGFP-expressing fibroblastic-like cells (Fig 3 A, arrows). The rare scattered insulin-positive cells were found negative for eGFP (Fig. 3C). Epithelial Pdx1-positive (Fig 3 B, arrow) and Pdx1-negative (Fig 3 B, arrowhead) cells were observed in these grafts. As expected with low MOI, only 0.6% of Pdx1-positive cells expressed eGFP (123 out of 20,000 Pdx1-positive cells counted). Such low percentage enables us to determine whether human islet beta cells are derived from one or several progenitors.

Bottom Line: By performing gene transfer at low multiplicity of infection, we created a chimeric graft with a subpopulation of human beta cells expressing GFP and found both GFP-positive and GFP-negative beta cells within single islets.The detection of both labeled and unlabeled beta cells in single islets demonstrates that beta cells present in a human islet are derived from multiple progenitors thus providing the first dynamic analysis of human islet formation during development.This human transgenic-like tool can be widely used to elucidate dynamic genetic processes in human tissue formation.

View Article: PubMed Central - PubMed

Affiliation: University Paris-Descartes, Faculty of Medicine, INSERM, Necker Hospital, U845, Paris, France.

ABSTRACT

Background: In both humans and rodents, glucose homeostasis is controlled by micro-organs called islets of Langerhans composed of beta cells, associated with other endocrine cell types. Most of our understanding of islet cell differentiation and morphogenesis is derived from rodent developmental studies. However, little is known about human islet formation. The lack of adequate experimental models has restricted the study of human pancreatic development to the histological analysis of different stages of pancreatic development. Our objective was to develop a new experimental model to (i) transfer genes into developing human pancreatic cells and (ii) validate gene transfer by defining the clonality of developing human islets.

Methods and findings: In this study, a unique model was developed combining ex vivo organogenesis from human fetal pancreatic tissue and cell type-specific lentivirus-mediated gene transfer. Human pancreatic progenitors were transduced with lentiviruses expressing GFP under the control of an insulin promoter and grafted to severe combined immunodeficient mice, allowing human beta cell differentiation and islet morphogenesis. By performing gene transfer at low multiplicity of infection, we created a chimeric graft with a subpopulation of human beta cells expressing GFP and found both GFP-positive and GFP-negative beta cells within single islets.

Conclusion: The detection of both labeled and unlabeled beta cells in single islets demonstrates that beta cells present in a human islet are derived from multiple progenitors thus providing the first dynamic analysis of human islet formation during development. This human transgenic-like tool can be widely used to elucidate dynamic genetic processes in human tissue formation.

Show MeSH
Related in: MedlinePlus