gamma-secretase inhibitor induces adipogenesis of adipose-derived stem cells by regulation of Notch and PPAR-gamma.
OBJECTIVE
To determine the inhibitory effect and mechanism of Notch signalling on adipogenesis of mouse adipose-derived stem cells (mASCs).


METHODS
Varied concentrations of N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butylester (DAPT) were added to mASCs 3 days before adipogenic induction with insulin-containing differentiation medium. The process of adipogenesis and ability of lipid droplet accumulation were analysed using oil red-O staining. The Notch signalling pathway (Notch-1, -2, -3, -4, Hes-1 and Hey-1) and adipogenesis-related factors (PPAR-gamma, DLK-1/Pref-1 and Acrp) were tested using real-time PCR, Western blot analysis and immunofluorescence staining assays.


RESULTS
We demonstrated that Notch-2-Hes-1 signalling pathway was inhibited dose-dependently by DAPT in mASCs. In addition, transcription of PPAR-gamma was promoted by DAPT before adipogenic induction, while inhibitor of adipogenesis DLK-1/Pref-1 was further depressed. At early stages of differentiation (2-4 days), adipogenesis in mASCs was advanced and significantly enhanced in 5 and 10 mum DAPT pre-treated cases. On day 4, in differentiated mASCs cases with DAPT pre-treatment, we also found promotion of activation of de-PPAR-gamma and depression of HES-1, DLK-1/Pref-1 mRNA and protein expression.


CONCLUSIONS
We conclude that blocking Notch signalling with DAPT enhances adipogenesis of differentiated mASCs at an early stage. It may be due to depression of DLK-1/Pref-1 and promotion of de-PPAR-gamma activation, which work through inhibition of Notch-2-Hes-1 pathway by DAPT.
Huang
Y
Y
State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, Chengdu, China.
Yang
X
X
Wu
Y
Y
Jing
W
W
Cai
X
X
Tang
W
W
Liu
L
L
Liu
Y
Y
Grottkau
B E
BE
Lin
Y
Y
D017667:Adipocytes / Q000166:cytology / Q000187:drug effects; D050156:Adipogenesis / Q000187:drug effects; D053829:Amyloid Precursor Protein Secretases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D004151:Dipeptides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D051379:Mice; D008815:Mice, Inbred Strains; D047495:PPAR gamma / Q000378:metabolism / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051880:Receptors, Notch / Q000378:metabolism / Q000502:physiology*; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism
D004151:Dipeptides; D004791:Enzyme Inhibitors; C419410:N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester; D047495:PPAR gamma; D012333:RNA, Messenger; D051880:Receptors, Notch; D053829:Amyloid Precursor Protein Secretases
Yi Chuan. 2004 Sep;26(5):653-7
15640080
Cell Prolif. 2007 Apr;40(2):185-95
17472726
Annu Rev Physiol. 2006;68:123-58
16460269
J Vet Med Sci. 2008 Aug;70(8):761-7
18772549
J Neurochem. 2008 Dec;107(6):1471-81
19094054
Differentiation. 2008 Jul;76(6):699-716
18565101
Br J Pharmacol. 2007 Jul;151(6):807-15
17549051
Mol Cell Biol. 2007 Mar;27(6):2294-308
17210639
J Biol Chem. 1997 Feb 21;272(8):5128-32
9030579
Aging Cell. 2004 Dec;3(6):379-89
15569355
Physiol Genomics. 2005 Oct 17;23(2):206-16
16106032
Semin Cancer Biol. 2004 Oct;14(5):348-56
15288260
Curr Mol Med. 2006 Dec;6(8):905-18
17168741
Cell Prolif. 2008 Oct;41(5):803-12
18616697
J Cell Mol Med. 2010 Apr;14(4):922-32
19243475
Cell Prolif. 2006 Apr;39(2):79-92
16542344
Semin Cell Dev Biol. 2003 Apr;14(2):113-9
12651094
Eur J Hum Genet. 2008 Sep;16(9):1126-34
18398438
Nature. 2002 May 30;417(6888):563-7
12037571
J Exp Med. 2005 Jul 4;202(1):157-68
15998794
J Cell Physiol. 2003 Mar;194(3):237-55
12548545
Cell Tissue Res. 2007 Dec;330(3):567-72
17922143
Cytotherapy. 2005;7(1):36-45
16040382
Ai Zheng. 2008 May;27(5):471-5
18479595
Am J Physiol Endocrinol Metab. 2005 Jun;288(6):E1195-205
15644454
Clin Otolaryngol Allied Sci. 2002 Aug;27(4):227-32
12169121
Dev Cell. 2004 Apr;6(4):454-6
15068783
J Biol Chem. 1997 Nov 21;272(47):29729-34
9368042
J Mol Biol. 2007 Apr 13;367(5):1281-93
17320900
Stem Cells. 2004;22(7):1346-55
15579652
Eur J Oral Sci. 2005 Aug;113(4):312-7
16048523
Dev Cell. 2004 Feb;6(2):219-27
14960276
Mol Endocrinol. 2006 Mar;20(3):698-705
16282371
Genesis. 2004 Sep;40(1):40-4
15354292
Kidney Int. 2005 Nov;68(5):1951-2
16221173
Nihon Rinsho. 2002 Jul;60 Suppl 7:593-600
12238104
Biochimie. 1997 Feb-Mar;79(2-3):111-2
9209705
Mol Endocrinol. 2006 Apr;20(4):809-30
16373399
Proc Natl Acad Sci U S A. 2003 Sep 30;100 Suppl 1:11889-95
12909711
Cell Death Differ. 2002 Aug;9(8):842-55
12107827
Curr Opin Genet Dev. 2004 Oct;14(5):506-12
15380241
J Endocrinol. 1998 Dec;159(3):429-39
9834460
Int J Oral Sci. 2009 Jun;1(2):81-9
20687300
Clin Cancer Res. 2006 Feb 15;12(4):1074-9
16489059
Mol Cell Biol. 2004 Apr;24(8):3505-13
15060169
J Cell Biochem. 2008 Mar 1;103(4):1054-65
17973271
J Biol Chem. 2002 Feb 22;277(8):6598-607
11741889
FEBS Lett. 2002 Jun 5;520(1-3):58-62
12044870
J Nutr. 2006 Dec;136(12):2953-6
17116701