A hot issue in current research regarding stem cells for regenerative medicine is the retainment of the stemness and multipotency of stem cell. Endothelial progenitor cells (EPCs) are characterized by an angiogenic switch that induces angiogenesis and further ameliorates the local microenvironment in ischemic organs. This study investigated whether EPCs could modulate the multipotent and differential abilities of mesenchymal stem cells (MSCs) in vitro and in vivo. We established an EPC/MSC indirect Transwell coculture system and then examined the effects of EPCs on the regulation of MSC biological properties in vitro and bone formation in vivo. The in vitro studies showed that cocultured MSCs (coMSCs) display no overt changes in cell morphology but an enhanced MSC phenotype compared with monocultured MSCs (monoMSCs). Our studies regarding the cellular, molecular, and protein characteristics of coMSCs and monoMSCs demonstrated that EPCs greatly promote the proliferation and differentiation potentials of coMSCs under indirect coculture condition. The expression of the pluripotency factors OCT4, SOX2, Nanog, and Klf4 was also upregulated in coMSCs. Furthermore, coMSCs combined with fibrin glue showed improved bone regeneration when used to repair rat alveolar bone defects compared with monoMSC grafts in vivo. This study is the first to demonstrate that EPCs have dynamic roles in maintaining MSC stemness and regulating MSC differentiation potential.
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References
1.
ChamberlainG, FoxJ, AshtonB and MiddletonJ. (2007). Concise review: mesenchymal stem cells: their phenotype, differentiation capacity, immunological features, and potential for homing. Stem Cells, 25:2739–2749.
2.
ChinnaduraiR, WallerEK, GalipeauJ and NookaAK. (2013). From single nucleotide polymorphisms to constant immunosuppression: mesenchymal stem cell therapy for autoimmune diseases. Biomed Res Int, 2013:929842.
3.
RomboutsWJ and PloemacherRE. (2003). Primary murine MSC show highly efficient homing to the bone marrow but lose homing ability following culture. Leukemia, 17:160–170.
4.
BaxterMA, WynnRF, JowittSN, WraithJE, FairbairnLJ and BellantuonoI. (2004). Study of telomere length reveals rapid aging of human marrow stromal cells following in vitro expansion. Stem Cells, 22:675–682.
5.
DuggalS, FronsdalKB, SzokeK, ShahdadfarA, MelvikJE and BrinchmannJE. (2009). Phenotype and gene expression of human mesenchymal stem cells in alginate scaffolds. Tissue Eng Part A, 15:1763–1773.
6.
ScaddenDT. (2006). The stem-cell niche as an entity of action. Nature, 441:1075–1079.
7.
GhaemiSR, HardingFJ, DelalatB, GronthosS and VoelckerNH. (2013). Exploring the mesenchymal stem cell niche using high throughput screening. Biomaterials, 34:7601–7615.
8.
YoshidaS, SukenoM and NabeshimaY. (2007). A vasculature-associated niche for undifferentiated spermatogonia in the mouse testis. Science, 317:1722–1726.
9.
TangW, ZeveD, SuhJM, BosnakovskiD, KybaM, HammerRE, TallquistMD and GraffJM. (2008). White fat progenitor cells reside in the adipose vasculature. Science, 322:583–586.
10.
CrisanM, YapS, CasteillaL, ChenCW, CorselliM, ParkTS, AndrioloG, SunB, ZhengB, et al. (2008). A perivascular origin for mesenchymal stem cells in multiple human organs. Cell Stem Cell, 3:301–313.
11.
AsaharaT, MuroharaT, SullivanA, SilverM, van der ZeeR, LiT, WitzenbichlerB, SchattemanG and IsnerJM. (1997). Isolation of putative progenitor endothelial cells for angiogenesis. Science, 275:964–967.
12.
LarriveeB and KarsanA. (2007). Involvement of marrow-derived endothelial cells in vascularization. Handb Exp Pharmacol. 89–114.
13.
KovacicJC, MooreJ, HerbertA, MaD, BoehmM and GrahamRM. (2008). Endothelial progenitor cells, angioblasts, and angiogenesis—old terms reconsidered from a current perspective. Trends Cardiovasc Med, 18:45–51.
14.
LuttunA, CarmelietG and CarmelietP. (2002). Vascular progenitors: from biology to treatment. Trends Cardiovasc Med, 12:88–96.
15.
KanczlerJM and OreffoRO. (2008). Osteogenesis and angiogenesis: the potential for engineering bone. Eur Cell Mater, 15:100–114.
HanY and HsiehFH. (2014). Osteogenic differentiation of late-outgrowth CD45-negative endothelial progenitor cells. J Vasc Res, 51:369–375.
18.
LiR, NauthA, LiC, QamiraniE, AtesokK and SchemitschEH. (2012). Expression of VEGF gene isoforms in a rat segmental bone defect model treated with EPCs. J Orthop Trauma, 26:689–692.
19.
LiR, NauthA, GandhiR, SyedK and SchemitschEH. (2014). BMP-2 mRNA expression after endothelial progenitor cell therapy for fracture healing. J Orthop Trauma, 28 (Suppl. 1):S24–S27.
20.
Zigdon-GiladiH, BickT, LewinsonD and MachteiEE. (2013). Co-transplantation of endothelial progenitor cells and mesenchymal stem cells promote neovascularization and bone regeneration. Clin Implant Dent Relat Res, 17:353–359.
21.
Zigdon-GiladiH, BickT, LewinsonD and MachteiEE. (2014). Mesenchymal stem cells and endothelial progenitor cells stimulate bone regeneration and mineral density. J Periodontol, 85:984–990.
22.
ItoK, YamadaY, NaikiT and UedaM. (2006). Simultaneous implant placement and bone regeneration around dental implants using tissue-engineered bone with fibrin glue, mesenchymal stem cells and platelet-rich plasma. Clin Oral Implants Res, 17:579–586.
23.
ZhangD and KilianKA. (2013). The effect of mesenchymal stem cell shape on the maintenance of multipotency. Biomaterials, 34:3962–3969.
24.
YangZ, JinF, ZhangX, MaD, HanC, HuoN, WangY, ZhangY, LinZ and JinY. (2009). Tissue engineering of cementum/periodontal-ligament complex using a novel three-dimensional pellet cultivation system for human periodontal ligament stem cells. Tissue Eng Part C Methods, 15:571–581.
25.
HenrichD, SeebachC, KaehlingC, ScherzedA, WilhelmK, TewksburyR, PowerskiM and MarziI. (2009). Simultaneous cultivation of human endothelial-like differentiated precursor cells and human marrow stromal cells on beta-tricalcium phosphate. Tissue Eng Part C Methods, 15:551–560.
26.
SalicA and MitchisonTJ. (2008). A chemical method for fast and sensitive detection of DNA synthesis in vivo. Proc Natl Acad Sci U S A, 105:2415–2420.
27.
LivakKJ and SchmittgenTD. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods, 25:402–408.
28.
HouJ, HanZP, JingYY, YangX, ZhangSS, SunK, HaoC, MengY, YuFH, et al. (2013). Autophagy prevents irradiation injury and maintains stemness through decreasing ROS generation in mesenchymal stem cells. Cell Death Dis, 4:e844.
29.
HungSP, HoJH, ShihYR, LoT and LeeOK. (2012). Hypoxia promotes proliferation and osteogenic differentiation potentials of human mesenchymal stem cells. J Orthop Res, 30:260–266.
30.
LeeKD, KuoTK, Whang-PengJ, ChungYF, LinCT, ChouSH, ChenJR, ChenYP and LeeOK. (2004). In vitro hepatic differentiation of human mesenchymal stem cells. Hepatology, 40:1275–1284.
31.
IngramDA, CapliceNM and YoderMC. (2005). Unresolved questions, changing definitions, and novel paradigms for defining endothelial progenitor cells. Blood, 106:1525–1531.
32.
KahlerCM, WechselbergerJ, HilbeW, GschwendtnerA, ColleselliD, NiedereggerH, BonebergEM, SpizzoG, WendelA, et al. (2007). Peripheral infusion of rat bone marrow derived endothelial progenitor cells leads to homing in acute lung injury. Respir Res, 8:50.
33.
KwonSM, LeeYK, YokoyamaA, JungSY, MasudaH, KawamotoA, LeeYM and AsaharaT. (2011). Differential activity of bone marrow hematopoietic stem cell subpopulations for EPC development and ischemic neovascularization. J Mol Cell Cardiol, 51:308–317.
34.
Kawabe-YakoR, IiM, MasuoO, AsaharaT and ItakuraT. (2011). Cilostazol activates function of bone marrow-derived endothelial progenitor cell for re-endothelialization in a carotid balloon injury model. PLoS One, 6:e24646.
35.
AminiAR, LaurencinCT and NukavarapuSP. (2012). Differential analysis of peripheral blood- and bone marrow-derived endothelial progenitor cells for enhanced vascularization in bone tissue engineering. J Orthop Res, 30:1507–1515.
36.
HackneyJA, CharbordP, BrunkBP, StoeckertCJ, LemischkaIR and MooreKA. (2002). A molecular profile of a hematopoietic stem cell niche. Proc Natl Acad Sci U S A, 99:13061–13066.
37.
AguirreA, PlanellJA and EngelE. (2010). Dynamics of bone marrow-derived endothelial progenitor cell/mesenchymal stem cell interaction in co-culture and its implications in angiogenesis. Biochem Biophys Res Commun, 400:284–291.
38.
LoiblM, BinderA, HerrmannM, DuttenhoeferF, RichardsRG, NerlichM, AliniM and VerrierS. (2014). Direct cell-cell contact between mesenchymal stem cells and endothelial progenitor cells induces a pericyte-like phenotype in vitro. Biomed Res Int, 2014:395781.
39.
NehlsV and DrenckhahnD. (1993). The versatility of microvascular pericytes: from mesenchyme to smooth muscle?. Histochemistry, 99:1–12.
40.
WinklerEA, BellRD and ZlokovicBV. (2010). Pericyte-specific expression of PDGF beta receptor in mouse models with normal and deficient PDGF beta receptor signaling. Mol Neurodegener, 5:32.
41.
HuangFJ, YouWK, BonaldoP, SeyfriedTN, PasqualeEB and StallcupWB. (2010). Pericyte deficiencies lead to aberrant tumor vascularizaton in the brain of the NG2 null mouse. Dev Biol, 344:1035–1046.
42.
QiH and PeiD. (2007). The magic of four: induction of pluripotent stem cells from somatic cells by Oct4, Sox2, Myc and Klf4. Cell Res, 17:578–580.
43.
TakahashiK, TanabeK, OhnukiM, NaritaM, IchisakaT, TomodaK and YamanakaS. (2007). Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell, 131:861–872.
44.
ZhangL, WangP, MeiS, LiC, CaiC and DingY. (2012). In vivo alveolar bone regeneration by bone marrow stem cells/fibrin glue composition. Arch Oral Biol, 57:238–244.
45.
KornP, SchulzMC, RangeU, LauerG and PradelW. (2014). Efficacy of tissue engineered bone grafts containing mesenchymal stromal cells for cleft alveolar osteoplasty in a rat model. J Craniomaxillofac Surg, 42:1277–1285.
46.
YoderMC, MeadLE, PraterD, KrierTR, MrouehKN, LiF, KrasichR, TemmCJ, PrchalJT and IngramDA. (2007). Redefining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals. Blood, 109:1801–1809.
47.
ZhangSJ, ZhangH, HouM, ZhengZ, ZhouJ, SuW, WeiY and HuS. (2007). Is it possible to obtain “true endothelial progenitor cells” by in vitro culture of bone marrow mononuclear cells?. Stem Cells Dev, 16:683–690.
48.
BerryMF, EnglerAJ, WooYJ, PirolliTJ, BishLT, JayasankarV, MorineKJ, GardnerTJ, DischerDE and SweeneyHL. (2006). Mesenchymal stem cell injection after myocardial infarction improves myocardial compliance. Am J Physiol Heart Circ Physiol, 290:H2196–H2203.
49.
XuJ, LiuX, ChenJ, ZacharekA, CuiX, Savant-BhonsaleS, LiuZ and ChoppM. (2009). Simvastatin enhances bone marrow stromal cell differentiation into endothelial cells via notch signaling pathway. Am J Physiol Cell Physiol, 296:C535–C543.
50.
DazziF, RamasamyR, GlennieS, JonesSP and RobertsI. (2006). The role of mesenchymal stem cells in haemopoiesis. Blood Rev, 20:161–171.
51.
XuJ, LiuX, ChenJ, ZacharekA, CuiX, Savant-BhonsaleS, ChoppM and LiuZ. (2010). Cell-cell interaction promotes rat marrow stromal cell differentiation into endothelial cell via activation of TACE/TNF-alpha signaling. Cell Transplant, 19:43–53.
52.
GalasRJJr. and LiuJC. (2014). Vascular endothelial growth factor does not accelerate endothelial differentiation of human mesenchymal stem cells. J Cell Physiol, 229:90–96.
53.
BalajiS, KingA, CrombleholmeTM and KeswaniSG. (2013). The role of endothelial progenitor cells in postnatal vasculogenesis: implications for therapeutic neovascularization and wound healing. Adv Wound Care (New Rochelle), 2:283–295.
54.
HynesB, KumarAH, O'SullivanJ, Klein BunekerC, LeblondAL, WeissS, SchmeckpeperJ, MartinK and CapliceNM. (2013). Potent endothelial progenitor cell-conditioned media-related anti-apoptotic, cardiotrophic, and pro-angiogenic effects post-myocardial infarction are mediated by insulin-like growth factor-1. Eur Heart J, 34:782–789.
55.
DevescoviV, LeonardiE, CiapettiG and CenniE. (2008). Growth factors in bone repair. Chir Organi Mov, 92:161–168.
56.
BallSG, ShuttleworthCA and KieltyCM. (2007). Vascular endothelial growth factor can signal through platelet-derived growth factor receptors. J Cell Biol, 177:489–500.
57.
KempenDH, LuL, HeijinkA, HefferanTE, CreemersLB, MaranA, YaszemskiMJ and DhertWJ. (2009). Effect of local sequential VEGF and BMP-2 delivery on ectopic and orthotopic bone regeneration. Biomaterials, 30:2816–2825.
58.
BaiY, LiP, YinG, HuangZ, LiaoX, ChenX and YaoY. (2013). BMP-2, VEGF and bFGF synergistically promote the osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells. Biotechnol Lett, 35:301–308.
59.
BerendsenAD and OlsenBR. (2014). How vascular endothelial growth factor-A (VEGF) regulates differentiation of mesenchymal stem cells. J Histochem Cytochem, 62:103–108.
60.
MaheraliN, SridharanR, XieW, UtikalJ, EminliS, ArnoldK, StadtfeldM, YachechkoR, TchieuJ, et al. (2007). Directly reprogrammed fibroblasts show global epigenetic remodeling and widespread tissue contribution. Cell Stem Cell, 1:55–70.
61.
GlaucheI, HerbergM and RoederI. (2010). Nanog variability and pluripotency regulation of embryonic stem cells—insights from a mathematical model analysis. PLoS One, 5:e11238.
62.
PapathanasopoulosA and GiannoudisPV. (2008). Biological considerations of mesenchymal stem cells and endothelial progenitor cells. Injury 39, (Suppl. 2)::S21–S32.
63.
GaoD, NolanD, McDonnellK, VahdatL, BenezraR, AltorkiN and MittalV. (2009). Bone marrow-derived endothelial progenitor cells contribute to the angiogenic switch in tumor growth and metastatic progression. Biochim Biophys Acta, 1796:33–40.
64.
Li CalziS, NeuMB, ShawLC, KielczewskiJL, MoldovanNI and GrantMB. (2010). EPCs and pathological angiogenesis: when good cells go bad. Microvasc Res, 79:207–216.
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