Limits...
Progenitor potential of nkx6.1-expressing cells throughout zebrafish life and during beta cell regeneration.

Ghaye AP, Bergemann D, Tarifeño-Saldivia E, Flasse LC, Von Berg V, Peers B, Voz ML, Manfroid I - BMC Biol. (2015)

Bottom Line: These two genes are initially co-expressed in the pancreatic primordium and their domains segregate, not as a result of mutual repression, but through the opposite effects of Notch signaling, maintaining nkx6.1 expression while repressing ascl1b in progenitors.In contrast to the mouse, pancreatic progenitor markers nkx6.1 and pdx1 continue to be expressed in adult ductal cells, a subset of which we show are still able to proliferate and undergo ductal and endocrine differentiation, providing robust evidence of the existence of pancreatic progenitor/stem cells in the adult zebrafish.Further characterization of these cells will open up new perspectives for anti-diabetic therapies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Zebrafish Development and Disease Models (ZDDM), GIGA-Research, (Sart-Tilman) University of Liège, Avenue de l'Hôpital 1, B34, 4000, Liège, Belgium. aurelieghaye@outlook.be.

ABSTRACT

Background: In contrast to mammals, the zebrafish has the remarkable capacity to regenerate its pancreatic beta cells very efficiently. Understanding the mechanisms of regeneration in the zebrafish and the differences with mammals will be fundamental to discovering molecules able to stimulate the regeneration process in mammals. To identify the pancreatic cells able to give rise to new beta cells in the zebrafish, we generated new transgenic lines allowing the tracing of multipotent pancreatic progenitors and endocrine precursors.

Results: Using novel bacterial artificial chromosome transgenic nkx6.1 and ascl1b reporter lines, we established that nkx6.1-positive cells give rise to all the pancreatic cell types and ascl1b-positive cells give rise to all the endocrine cell types in the zebrafish embryo. These two genes are initially co-expressed in the pancreatic primordium and their domains segregate, not as a result of mutual repression, but through the opposite effects of Notch signaling, maintaining nkx6.1 expression while repressing ascl1b in progenitors. In the adult zebrafish, nkx6.1 expression persists exclusively in the ductal tree at the tip of which its expression coincides with Notch active signaling in centroacinar/terminal end duct cells. Tracing these cells reveals that they are able to differentiate into other ductal cells and into insulin-expressing cells in normal (non-diabetic) animals. This capacity of ductal cells to generate endocrine cells is supported by the detection of ascl1b in the nkx6.1:GFP ductal cell transcriptome. This transcriptome also reveals, besides actors of the Notch and Wnt pathways, several novel markers such as id2a. Finally, we show that beta cell ablation in the adult zebrafish triggers proliferation of ductal cells and their differentiation into insulin-expressing cells.

Conclusions: We have shown that, in the zebrafish embryo, nkx6.1+ cells are bona fide multipotent pancreatic progenitors, while ascl1b+ cells represent committed endocrine precursors. In contrast to the mouse, pancreatic progenitor markers nkx6.1 and pdx1 continue to be expressed in adult ductal cells, a subset of which we show are still able to proliferate and undergo ductal and endocrine differentiation, providing robust evidence of the existence of pancreatic progenitor/stem cells in the adult zebrafish. Our findings support the hypothesis that nkx6.1+ pancreatic progenitors contribute to beta cell regeneration. Further characterization of these cells will open up new perspectives for anti-diabetic therapies.

No MeSH data available.


Related in: MedlinePlus

nkx6.1 and ascl1b are first co-expressed in the endocrine precursors of the dorsal bud but rapidly their expression domain moves apart. Immunodetection of endogenous Nkx6.1 and GFP in Tg(ascl1b:eGFP-creERT2) embryos at 14 hpf (a–a") and 15 hpf (b–b"). All views are ventral with the anterior part to the left and represent confocal projection images. Scale bars = 40 μm. H hypochord, M medial cells
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4556004&req=5

Fig4: nkx6.1 and ascl1b are first co-expressed in the endocrine precursors of the dorsal bud but rapidly their expression domain moves apart. Immunodetection of endogenous Nkx6.1 and GFP in Tg(ascl1b:eGFP-creERT2) embryos at 14 hpf (a–a") and 15 hpf (b–b"). All views are ventral with the anterior part to the left and represent confocal projection images. Scale bars = 40 μm. H hypochord, M medial cells

Mentions: As presented above, nkx6.1 is expressed in the multipotent pancreatic progenitors and ascl1b in the endocrine precursors; we therefore analyzed the relationship between these two populations by comparing the Nkx6.1 and GFP proteins in Tg(ascl1b:eGFP-2A-creERT2) embryos. At 14 hpf, the ascl1b:eGFP cells delineate two lines adjacent to the midline (Fig. 4a–a"). These cells correspond to the most medial endodermal cells (indicated by M in Fig. 4a'), reported to give rise mostly to pancreatic endocrine cells [29, 30]. At this stage, all of these ascl1b:eGFP cells also express Nkx6.1. In contrast, the Nkx6.1 expression domain is larger and, in addition to its expression in the hypochord (indicated by H in Fig. 4a), it is also expressed in the lateral cells, reported to give rise to exocrine and intestinal cells [29]. Rapidly, these two domains segregate since, as early as 1 hour later, the majority of ascl1b:eGFP+ cells no longer express Nkx6.1 (Fig. 4b–b"). Thus, separation of the two domains is largely completed when hormone-expressing cells start differentiating.Fig. 4


Progenitor potential of nkx6.1-expressing cells throughout zebrafish life and during beta cell regeneration.

Ghaye AP, Bergemann D, Tarifeño-Saldivia E, Flasse LC, Von Berg V, Peers B, Voz ML, Manfroid I - BMC Biol. (2015)

nkx6.1 and ascl1b are first co-expressed in the endocrine precursors of the dorsal bud but rapidly their expression domain moves apart. Immunodetection of endogenous Nkx6.1 and GFP in Tg(ascl1b:eGFP-creERT2) embryos at 14 hpf (a–a") and 15 hpf (b–b"). All views are ventral with the anterior part to the left and represent confocal projection images. Scale bars = 40 μm. H hypochord, M medial cells
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4556004&req=5

Fig4: nkx6.1 and ascl1b are first co-expressed in the endocrine precursors of the dorsal bud but rapidly their expression domain moves apart. Immunodetection of endogenous Nkx6.1 and GFP in Tg(ascl1b:eGFP-creERT2) embryos at 14 hpf (a–a") and 15 hpf (b–b"). All views are ventral with the anterior part to the left and represent confocal projection images. Scale bars = 40 μm. H hypochord, M medial cells
Mentions: As presented above, nkx6.1 is expressed in the multipotent pancreatic progenitors and ascl1b in the endocrine precursors; we therefore analyzed the relationship between these two populations by comparing the Nkx6.1 and GFP proteins in Tg(ascl1b:eGFP-2A-creERT2) embryos. At 14 hpf, the ascl1b:eGFP cells delineate two lines adjacent to the midline (Fig. 4a–a"). These cells correspond to the most medial endodermal cells (indicated by M in Fig. 4a'), reported to give rise mostly to pancreatic endocrine cells [29, 30]. At this stage, all of these ascl1b:eGFP cells also express Nkx6.1. In contrast, the Nkx6.1 expression domain is larger and, in addition to its expression in the hypochord (indicated by H in Fig. 4a), it is also expressed in the lateral cells, reported to give rise to exocrine and intestinal cells [29]. Rapidly, these two domains segregate since, as early as 1 hour later, the majority of ascl1b:eGFP+ cells no longer express Nkx6.1 (Fig. 4b–b"). Thus, separation of the two domains is largely completed when hormone-expressing cells start differentiating.Fig. 4

Bottom Line: These two genes are initially co-expressed in the pancreatic primordium and their domains segregate, not as a result of mutual repression, but through the opposite effects of Notch signaling, maintaining nkx6.1 expression while repressing ascl1b in progenitors.In contrast to the mouse, pancreatic progenitor markers nkx6.1 and pdx1 continue to be expressed in adult ductal cells, a subset of which we show are still able to proliferate and undergo ductal and endocrine differentiation, providing robust evidence of the existence of pancreatic progenitor/stem cells in the adult zebrafish.Further characterization of these cells will open up new perspectives for anti-diabetic therapies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Zebrafish Development and Disease Models (ZDDM), GIGA-Research, (Sart-Tilman) University of Liège, Avenue de l'Hôpital 1, B34, 4000, Liège, Belgium. aurelieghaye@outlook.be.

ABSTRACT

Background: In contrast to mammals, the zebrafish has the remarkable capacity to regenerate its pancreatic beta cells very efficiently. Understanding the mechanisms of regeneration in the zebrafish and the differences with mammals will be fundamental to discovering molecules able to stimulate the regeneration process in mammals. To identify the pancreatic cells able to give rise to new beta cells in the zebrafish, we generated new transgenic lines allowing the tracing of multipotent pancreatic progenitors and endocrine precursors.

Results: Using novel bacterial artificial chromosome transgenic nkx6.1 and ascl1b reporter lines, we established that nkx6.1-positive cells give rise to all the pancreatic cell types and ascl1b-positive cells give rise to all the endocrine cell types in the zebrafish embryo. These two genes are initially co-expressed in the pancreatic primordium and their domains segregate, not as a result of mutual repression, but through the opposite effects of Notch signaling, maintaining nkx6.1 expression while repressing ascl1b in progenitors. In the adult zebrafish, nkx6.1 expression persists exclusively in the ductal tree at the tip of which its expression coincides with Notch active signaling in centroacinar/terminal end duct cells. Tracing these cells reveals that they are able to differentiate into other ductal cells and into insulin-expressing cells in normal (non-diabetic) animals. This capacity of ductal cells to generate endocrine cells is supported by the detection of ascl1b in the nkx6.1:GFP ductal cell transcriptome. This transcriptome also reveals, besides actors of the Notch and Wnt pathways, several novel markers such as id2a. Finally, we show that beta cell ablation in the adult zebrafish triggers proliferation of ductal cells and their differentiation into insulin-expressing cells.

Conclusions: We have shown that, in the zebrafish embryo, nkx6.1+ cells are bona fide multipotent pancreatic progenitors, while ascl1b+ cells represent committed endocrine precursors. In contrast to the mouse, pancreatic progenitor markers nkx6.1 and pdx1 continue to be expressed in adult ductal cells, a subset of which we show are still able to proliferate and undergo ductal and endocrine differentiation, providing robust evidence of the existence of pancreatic progenitor/stem cells in the adult zebrafish. Our findings support the hypothesis that nkx6.1+ pancreatic progenitors contribute to beta cell regeneration. Further characterization of these cells will open up new perspectives for anti-diabetic therapies.

No MeSH data available.


Related in: MedlinePlus