Directed pancreatic acinar differentiation of mouse embryonic stem cells via embryonic signalling molecules and exocrine transcription factors.
Pluripotent embryonic stem cells (ESC) are a promising cellular system for generating an unlimited source of tissue for the treatment of chronic diseases and valuable in vitro differentiation models for drug testing. Our aim was to direct differentiation of mouse ESC into pancreatic acinar cells, which play key roles in pancreatitis and pancreatic cancer. To that end, ESC were first differentiated as embryoid bodies and sequentially incubated with activin A, inhibitors of Sonic hedgehog (Shh) and bone morphogenetic protein (BMP) pathways, fibroblast growth factors (FGF) and retinoic acid (RA) in order to achieve a stepwise increase in the expression of mRNA transcripts encoding for endodermal and pancreatic progenitor markers. Subsequent plating in Matrigel® and concomitant modulation of FGF, glucocorticoid, and folllistatin signalling pathways involved in exocrine differentiation resulted in a significant increase of mRNAs encoding secretory enzymes and in the number of cells co-expressing their protein products. Also, pancreatic endocrine marker expression was down-regulated and accompanied by a significant reduction in the number of hormone-expressing cells with a limited presence of hepatic marker expressing-cells. These findings suggest a selective activation of the acinar differentiation program. The newly differentiated cells were able to release α-amylase and this feature was greatly improved by lentiviral-mediated expression of Rbpjl and Ptf1a, two transcription factors involved in the maximal production of digestive enzymes. This study provides a novel method to produce functional pancreatic exocrine cells from ESC.
Cancer Research Program, Institut Hospital del Mar d'Investigacions Mèdiques (IMIM), Biomedical Research Park, Barcelona, Spain.
Fabien
F
Delaspre
Mohammad
M
Massumi
Marta
M
Salido
Bernat
B
Soria
Philippe
P
Ravassard
Pierre
P
Savatier
Anouchka
A
Skoudy
D061354:Acinar Cells / Q000166:cytology* / Q000378:metabolism; D028341:Activins / Q000494:pharmacology; D000818:Animals; D002454:Cell Differentiation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D058732:Embryoid Bodies / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D018613:Microscopy, Confocal; D010179:Pancreas / Q000166:cytology* / Q000378:metabolism; D046790:Pancreas, Exocrine / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic; D014212:Tretinoin / Q000494:pharmacology; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism
D004268:DNA-Binding Proteins; C484553:Rbpsuhl protein, mouse; D014157:Transcription Factors; C422405:activin A; C062602:transcription factor PTF1; D028341:Activins; D014212:Tretinoin; D005346:Fibroblast Growth Factors; D000516:alpha-Amylases