DLK1 regulates branching morphogenesis and parasympathetic innervation of salivary glands through inhibition of NOTCH signalling.
BACKGROUND
Delta-like proteins 1 and 2 (DLK1, 2) are NOTCH receptor ligands containing epidermal growth factor-like repeats, which regulate NOTCH signalling. We investigated the role of DLK and the NOTCH pathway in the morphogenesis of the submandibular salivary glands (SMGs), using in vitro organotypic cultures.


RESULTS
DLK1 and 2 were present in all stages of SMG morphogenesis, where DLK1 inhibited both NOTCH activity and SMG branching. The addition of NOTCH inhibitory agents, either soluble DLK1 (sDLK1) or N-[N-(3, 5-difluorophenacetyl-L-alanyl]-S-phenylglycine t-buthyl ester (DAPT), to the SMG culture medium did not affect the rate of cell proliferation, but induced a strong reduction in SMG branching, increased epithelial apoptosis, and impaired innervation of the epithelial end buds by local parasympathetic ganglion neurons. SMG innervation could be restored by the acetylcholine analog carbachol (CCh), which also rescued cytokeratin 5 (CK5(+))-expressing epithelial progenitor cells. Despite this, CCh failed to restore normal branching morphogenesis in the presence of either sDLK1 or DAPT. However, it improved recovery of branching morphogenesis in SMGs, once DLK1 or DAPT were removed from the medium.


CONCLUSIONS
Our data suggest that DLK1 regulates SMGs morphogenesis and parasympathetic nerve fibre outgrowth through inhibition of NOTCH signalling.
Department of Cell Biology and Histology, Faculty of Medicine and Dentistry, University of the Basque Country, UPV/EHU, Leioa, 48940, Bizkaia, Spain.
Patricia
P
García-Gallastegui
Gaskon
G
Ibarretxe
José-Javier
JJ
Garcia-Ramírez
Victoriano
V
Baladrón
Maitane
M
Aurrekoetxea
María-Luisa
ML
Nueda
Ana-Isabel
AI
Naranjo
Francisco
F
Santaolalla
Ana
A
Sánchez-del Rey
Jorge
J
Laborda
Fernando
F
Unda
D000818:Animals; D002135:Calcium-Binding Proteins; D004151:Dipeptides / Q000494:pharmacology; D005726:Ganglia, Parasympathetic / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology / Q000502:physiology*; D051379:Mice; D009024:Morphogenesis / Q000502:physiology; D009924:Organ Culture Techniques; D051880:Receptors, Notch / Q000037:antagonists & inhibitors / Q000502:physiology*; D015398:Signal Transduction; D013234:Stem Cells / Q000502:physiology; D013363:Submandibular Gland / Q000196:embryology / Q000294:innervation
D002135:Calcium-Binding Proteins; D004151:Dipeptides; C512925:Dlk1 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; C419410:N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester; D051880:Receptors, Notch
Cell differentiation; Development; Organogenesis