Restoring β-cell mass by the transplantation of pancreatic islets is an effective diabetes treatment, but it is limited by the shortage of donor organs. CD133-expressing pancreatic ductal epithelial cells (PDECs) have the ability to generate insulin-producing cells. The expansion of these cells is dependent on extrinsic niche factors, but few of those signals have been identified. In this study, CD133-expressing PDECs were purified by sorting from adult wild-type C57BL/6 mice and TGFβRIInull/null mice. Furthermore, using immunofluorescence and transplantation assays, we found that the inhibition of the transforming growth factor-β (TGF-β) pathway promoted the expansion of CD133-expressing PDECs for many generations and maintained the ability of CD133-expressing PDECs to generate insulin-producing cells. Moreover, western blot, qRT-PCR, and dual luciferase assays using TGF-β inhibitors were performed to identify the mechanisms by which TGF-β signaling regulates proliferation and differentiation. The results showed that the inhibition of TGF-β signaling enhanced Id2 binding to the promoter region of the cell proliferation repressor p16 and promoted the expansion of CD133-expressing PDECs, and the increased Id2 binding to NeuroD1 decreased the transcription of Pax6 to maintain CD133-expressing PDECs in the Pdx1-expression stage. Taken together, the effect of TGF-β antagonists on CD133-expressing PDECs reveals a novel paradigm of signaling that explains the balance between the expansion and differentiation of pancreatic duct epithelial progenitors.
MillmanJR, XieC, Van DervortA, GurtlerM, PagliucaFW and MeltonDA. (2016). Generation of stem cell-derived beta-cells from patients with type 1 diabetes. Nat Commun, 7:11463.
6.
ChenS, BorowiakM, FoxJL, MaehrR, OsafuneK, DavidowL, LamK, PengLF, SchreiberSL, RubinLL and MeltonD. (2009). A small molecule that directs differentiation of human ESCs into the pancreatic lineage. Nat Chem Biol, 5:258–265.
7.
XuX, D'HokerJ, StangeG, BonneS, De LeuN, XiaoX, Van de CasteeleM, MellitzerG, LingZ, et al. (2008). Beta cells can be generated from endogenous progenitors in injured adult mouse pancreas. Cell, 132:197–207.
8.
PagliucaFW, MillmanJR, GurtlerM, SegelM, Van DervortA, RyuJH, PetersonQP, GreinerD and MeltonDA. (2014). Generation of functional human pancreatic beta cells in vitro. Cell, 159:428–439.
9.
PictetR, RutterWJ, SterinerDF and FrenkelN. (1972). Development of the embryonic endocrine pancreas. In: Steiner D, Freinkel N (eds.) Handbook of physiology, section 7. Williams & Wilkins, Baltimore,25–66.
10.
SlackJM. (1995). Developmental biology of the pancreas. Development, 121:1569–1580.
11.
OshimaY, SuzukiA, KawashimoK, IshikawaM, OhkohchiN and TaniguchiH. (2007). Isolation of mouse pancreatic ductal progenitor cells expressing CD133 and c-Met by flow cytometric cell sorting. Gastroenterology, 132:720–732.
12.
HoriY, FukumotoM and KurodaY. (2008). Enrichment of putative pancreatic progenitor cells from mice by sorting for prominin1 (CD133) and platelet-derived growth factor receptor beta. Stem Cells, 26:2912–2920.
13.
NikolovaT, WuM, BrumbarovK, AltR, OpitzH, BohelerKR, CrossM and WobusAM. (2007). WNT-conditioned media differentially affect the proliferation and differentiation of cord blood-derived CD133+ cells in vitro. Differentiation, 75:100–111.
14.
JinL, FengT, ShihHP, ZerdaR, LuoA, HsuJ, MahdaviA, SanderM, TirrellDA, RiggsAD and KuHT. (2013). Colony-forming cells in the adult mouse pancreas are expandable in Matrigel and form endocrine/acinar colonies in laminin hydrogel. Proc Natl Acad Sci U S A, 110:3907–3912.
15.
StonerGD, HarrisCC, BostwickDG, JonesRT, TrumpBF, KingsburyEW, FinemanE and NewkirkC. (1978). Isolation and characterization of epithelial cells from bovine pancreatic duct. In Vitro, 14:581–590.
16.
KolarC, CaffreyT, HollingsworthM, ScheetzM, SutherlinM, WeideL and LawsonT. (1997). Duct epithelial cells cultured from human pancreas processed for transplantation retain differentiated ductal characteristics. Pancreas, 15:265–271.
17.
GithensS, PatkeCL and SchexnayderJA. (1994). Isolation and culture of rhesus monkey pancreatic ductules and ductule-like epithelium. Pancreas, 9:20–31.
18.
VermeTB and HootmanSR. (1990). Regulation of pancreatic duct epithelial growth in vitro. Am J Physiol, 258:G833–G840.
19.
YuanS, MetrakosP, DuguidWP and RosenbergL. (1995). Isolation and culture of intralobular ducts from the hamster pancreas. In Vitro Cell Dev Biol Anim, 31:77–80.
20.
RichardsJ, PascoD, YangJ, GuzmanR and NandiS. (1983). Comparison of the growth of normal and neoplastic mouse mammary cells on plastic, on collagen gels and in collagen gels. Exp Cell Res, 146:1–14.
21.
Bonner-WeirS, TanejaM, WeirGC, TatarkiewiczK, SongKH, SharmaA and O'NeilJJ. (2000). In vitro cultivation of human islets from expanded ductal tissue. Proc Natl Acad Sci U S A, 97:7999–8004.
22.
De LisleRC and LogsdonCD. (1990). Pancreatic acinar cells in culture: expression of acinar and ductal antigens in a growth-related manner. Eur J Cell Biol, 51:64–75.
23.
HallPA and LemoineNR. (1992). Rapid acinar to ductal transdifferentiation in cultured human exocrine pancreas. J Pathol, 166:97–103.
24.
VilaMR, LloretaJ and RealFX. (1994). Normal human pancreas cultures display functional ductal characteristics. Lab Invest, 71:423–431.
25.
MaD, TangS, SongJ, WuQ, ZhangF, XingY, PanY, ZhangY, JiangJ, ZhangY and JinL. (2017). Culturing and transcriptome profiling of progenitor-like colonies derived from adult mouse pancreas. Stem Cell Res Ther, 8:172.
26.
GuaschG, SchoberM, PasolliHA, ConnEB, PolakL and FuchsE. (2007). Loss of TGFbeta signaling destabilizes homeostasis and promotes squamous cell carcinomas in stratified epithelia. Cancer Cell, 12:313–327.
27.
HeXC, ZhangJ, TongWG, TawfikO, RossJ, ScovilleDH, TianQ, ZengX, HeX, et al. (2004). BMP signaling inhibits intestinal stem cell self-renewal through suppression of Wnt-beta-catenin signaling. Nat Genet, 36:1117–1121.
28.
QiX, LiTG, HaoJ, HuJ, WangJ, SimmonsH, MiuraS, MishinaY and ZhaoGQ. (2004). BMP4 supports self-renewal of embryonic stem cells by inhibiting mitogen-activated protein kinase pathways. Proc Natl Acad Sci U S A, 101:6027–6032.
29.
OshimoriN and FuchsE. (2012). Paracrine TGF-beta signaling counterbalances BMP-mediated repression in hair follicle stem cell activation. Cell Stem Cell, 10:63–75.
30.
ChapellierM, Bachelard-CascalesE, SchmidtX, ClementF, TreilleuxI, DelayE, JammotA, Menetrier-CauxC, PochonG, et al. (2015). Disequilibrium of BMP2 levels in the breast stem cell niche launches epithelial transformation by overamplifying BMPR1B cell response. Stem Cell Rep, 4:239–254.
31.
ChengL, HuW, QiuB, ZhaoJ, YuY, GuanW, WangM, YangW and PeiG. (2014). Generation of neural progenitor cells by chemical cocktails and hypoxia. Cell Res, 24:665–679.
32.
SunZ and EttensohnCA. (2017). TGF-beta sensu stricto signaling regulates skeletal morphogenesis in the sea urchin embryo. Dev Biol, 421:149–160.
33.
TangY and ChengL. (2017). Cocktail of chemical compounds robustly promoting cell reprogramming protects liver against acute injury. Protein Cell, 8:273–283.
34.
DhawanS, DiriceE, KulkarniRN and BhushanA. (2016). Inhibition of TGF-beta signaling promotes human pancreatic beta-cell replication. Diabetes, 65:1208–1218.
35.
KandasamyM, LehnerB, KrausS, SanderPR, MarschallingerJ, RiveraFJ, TrumbachD, UeberhamU, ReitsamerHA, et al. (2014). TGF-beta signalling in the adult neurogenic niche promotes stem cell quiescence as well as generation of new neurons. J Cell Mol Med, 18:1444–1459.
36.
KawanoM, TanakaK, ItonagaI, IwasakiT and TsumuraH. (2017). MicroRNA-20b promotes cell proliferation via targeting of TGF-beta receptor II and upregulates MYC expression in Ewing's sarcoma cells. Int J Oncol, 51:1842–1850.
ChaeJH, SteinGH and LeeJE. (2004). NeuroD: the predicted and the surprising. Mol Cells, 18:271–288.
39.
GhilSH, JeonYJ and Suh-KimH. (2002). Inhibition of BETA2/NeuroD by Id2. Exp Mol Med, 34:367–373.
40.
Bastidas-PonceA, RoscioniSS, BurtscherI, BaderE, SterrM, BakhtiM and LickertH. (2017). Foxa2 and Pdx1 cooperatively regulate postnatal maturation of pancreatic beta-cells. Mol Metab, 6:524–534.
41.
MfopouJK, ChenB, MateizelI, SermonK and BouwensL. (2010). Noggin, retinoids, and fibroblast growth factor regulate hepatic or pancreatic fate of human embryonic stem cells. Gastroenterology, 138:2233–2245, 2245.e1–e14.
42.
Bonner-WeirS, ToschiE, InadaA, ReitzP, FonsecaSY, AyeT and SharmaA. (2004). The pancreatic ductal epithelium serves as a potential pool of progenitor cells. Pediatr Diabetes 5 Suppl, 2:16–22.
43.
HabenerJF. (2004). A perspective on pancreatic stem/progenitor cells. Pediatr Diabetes 5 Suppl, 2:29–37.
44.
FortunelNO, HatzfeldJA, RosemaryPA, FerrarisC, MonierMN, HaydontV, LonguetJ, BrethonB, LimB, et al. (2003). Long-term expansion of human functional epidermal precursor cells: promotion of extensive amplification by low TGF-beta1 concentrations. J Cell Sci, 116:4043–4052.
45.
MoonH, JuHL, ChungSI, ChoKJ, EunJW, NamSW, HanKH, CalvisiDF and RoSW. (2017). Transforming Growth Factor-beta promotes liver tumorigenesis in mice via up-regulation of snail. Gastroenterology, 153:1378–1391.e6.
46.
de MirandaNF, van DintherM, van den AkkerBE, van WezelT, ten DijkeP and MorreauH. (2015). Transforming growth factor beta signaling in colorectal cancer cells with microsatellite instability despite biallelic mutations in TGFBR2. Gastroenterology, 148:1427–1437.e8.
47.
KrishnamurthyJ, RamseyMR, LigonKL, TorriceC, KohA, Bonner-WeirS and SharplessNE. (2006). p16INK4a induces an age-dependent decline in islet regenerative potential. Nature, 443:453–457.
48.
KongY, SharmaRB, LyS, StamaterisRE, JesdaleWM and AlonsoLC. (2018). CDKN2A/B T2D genome-wide association study risk SNPs impact locus gene expression and proliferation in human islets. Diabete, 67:872–884.
49.
BrackenAP, Kleine-KohlbrecherD, DietrichN, PasiniD, GargiuloG, BeekmanC, Theilgaard-MonchK, MinucciS, PorseBT, et al. (2007). The Polycomb group proteins bind throughout the INK4A-ARF locus and are disassociated in senescent cells. Genes Dev, 21:525–530.
50.
MouH, VinarskyV, TataPR, BrazauskasK, ChoiSH, CrookeAK, ZhangB, SolomonGM, TurnerB, et al. (2016). Dual SMAD signaling inhibition enables long-term expansion of diverse epithelial basal cells. Cell Stem Cell, 19:217–231.
51.
NishimuraEK, SuzukiM, IgrasV, DuJ, LonningS, MiyachiY, RoesJ, BeermannF and FisherDE. (2010). Key roles for transforming growth factor beta in melanocyte stem cell maintenance. Cell Stem Cell, 6:130–140.
52.
Ashery-PadanR, ZhouX, MarquardtT, HerreraP, ToubeL, BerryA and GrussP. (2004). Conditional inactivation of Pax6 in the pancreas causes early onset of diabetes. Dev Biol, 269:479–488.
53.
MarsichE, VetereA, Di PiazzaM, TellG and PaolettiS. (2003). The PAX6 gene is activated by the basic helix-loop-helix transcription factor NeuroD/BETA2. Biochem J, 376:707–715.
54.
KondratyevaLG, SveshnikovaAA, GrankinaEV, ChernovIP, KopantsevaMR, KopantzevEP and SverdlovED. (2016). Downregulation of expression of mater genes SOX9, FOXA2, and GATA4 in pancreatic cancer cells stimulated with TGFbeta1 epithelial-mesenchymal transition. Dokl Biochem Biophys, 469:257–259.
55.
LeeCS, SundNJ, VatamaniukMZ, MatschinskyFM, StoffersDA and KaestnerKH. (2002). Foxa2 controls Pdx1 gene expression in pancreatic beta-cells in vivo. Diabetes, 51:2546–2551.
56.
GaoN, LeLayJ, VatamaniukMZ, RieckS, FriedmanJR and KaestnerKH. (2008). Dynamic regulation of Pdx1 enhancers by Foxa1 and Foxa2 is essential for pancreas development. Genes Dev, 22:3435–3448.
57.
TulachanSS, TeiE, HembreeM, CriseraC, PrasadanK, KoizumiM, ShahS, GuoP, BottingerE and GittesGK. (2007). TGF-beta isoform signaling regulates secondary transition and mesenchymal-induced endocrine development in the embryonic mouse pancreas. Dev Biol, 305:508–521.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
