We developed a novel pH- and thermo-sensitive hydrogel as a scaffold for autologous bone tissue engineering. We synthesized this polymer by adding pH-sensitive sulfamethazine oligomers (SMOs) to both ends of a thermo-sensitive poly(ɛ-caprolactone-co-lactide)–poly(ethylene glycol)–poly(ɛ-caprolactone-co-lactide) (PCLA-PEG-PCLA) block copolymer, yielding a pH/thermo-sensitive SMO-PCLA-PEG-PCLA-SMO block copolymer. The synthesized block copolymer solution rapidly formed a stable gel under physiological conditions (pH 7.4 and 37°C), whereas it formed a sol at pH 8.0 and 37°C, making it injectable. This pH/thermo-sensitive hydrogel exhibited high biocompatibility in a Dulbecco's modified Eagle's medium extract test. Under physiological conditions, the hydrogel easily encapsulated human mesenchymal stem cells (hMSCs) and recombinant human bone morphogenetic protein-2 (rhBMP-2), with encapsulating efficiencies of about 90% and 85%, respectively. To assay for ectopic bone formation in vivo, we subcutaneously injected a polymer solution containing hMSCs and rhBMP-2 into the back of mice, after which we could observe hMSC differentiation for up to 7 weeks. Histological studies revealed mineralized tissue formation and high levels of alkaline phosphatase activity in the mineralized tissue. Therefore, this pH/thermo-sensitive SMO-PCLA-PEG-PCLA-SMO block copolymer demonstrated potential as an injectable scaffold for bone tissue engineering, with in situ formation capabilities.
VacantiJ.P., LangerR., UptonJ., MarlerJ.J.Transplantation of cells in matrices for tissue regeneration. Adv Drug Deliv Rev, 33:165. 1998.
3.
PetiteH., ViateauV., BensaidW., MeunierA., de PollakC., BourguignonM., OudinaK., SedelL., GuilleminG.Tissue-engineered bone regeneration. Nat Biotechnol, 18:959. 2000.
4.
MauneyJ.R., VollochV., KaplanD.L.Role of adult mesenchymal stem cells in bone tissue engineering applications: current status and future prospects. Tissue Eng, 11:787. 2005.
5.
NeubauerM., HackerM., Bauer-KreiselP., WeiserB., FischbachC., SchulzM.B., GoepferichA., BlunkT.Adipose tissue engineering based on mesenchymal stem cells and basic fibroblast growth factor in vitro. Tissue Eng, 11:1840. 2005.
6.
ShinM., YoshimotoH., VacantiJ.P.In vivo bone tissue engineering using mesenchymal stem cells on a novel electrospun nanofibrous scaffold. Tissue Eng, 10:33. 2004.
7.
TakahashiY., TabataY.Homogeneous seeding of mesenchymal stem cells into nonwoven fabric for tissue engineering. Tissue Eng, 9:931. 2003.
8.
ZukP.A., ZhuM., AshjianP., de UgarteD.A., HuangJ.I., MizunoH., AlfonsoZ.C., FraserJ.K., BenhaimP., HedrickM.H.Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell, 13:4279. 2002.
PittengerM.F., MackayA.M., BeckS.C., JaiswalR.K., DouglasR., MoscaJ.D., MoormanM.A., SimonettiD.W., CraigS., MarshakD.R.Multilineage potential of adult human mesenchymal stem cells. Science, 284:143. 1999.
11.
LuuH.H., SongW.X., LuoX., ManningD., LuoJ., DengZ.L., SharffK.A., MontagA.G., HaydonR.C., HeT.C.Distinct roles of bone morphogenetic proteins in osteogenic differentiation of mesenchymal stem cells. J Orthop Res, 25:665. 2007.
12.
HollingerJ.O., UludagH., WinnS.R.Sustained release emphasizing recombinant human bone morphogenetic protein-2. Adv Drug Deliv Rev, 31:303. 1998.
13.
GittensS.A., UludagH.Growth factor delivery for bone tissue engineering. J Drug Target, 9:407. 2001.
14.
WangE.A., RosenV., D'AlessandroJ.S., BauduyM., CordesP., HaradaT., IsraelD.I., HewickR.M., KernsK.M., LaPanP.Recombinant human bone morphogenetic protein induces bone formation. Proc Natl Acad Sci USA, 87:2220. 1990.
15.
MeinelL., HofmannS., BetzO., FajardoR., MerkleH.P., LangerR., EvansC.H., Vunjak-NovakovicG., KaplanD.L.Osteogenesis by human mesenchymal stem cells cultured on silk biomaterials: comparison of adenovirus mediated gene transfer and protein delivery of BMP-2. Biomaterials, 27:4993. 2006.
16.
NaK., KimS.W., SunB.K., WooD.G., YangH.N., ChungH.M., ParkK.H.Osteogenic differentiation of rabbit mesenchymal stem cells in thermo-reversible hydrogel constructs containing hydroxyapatite and bone morphogenic protein-2 (BMP-2)Biomaterials, 28:2631. 2007.
17.
MeinelL., KarageorgiouV., HofmannS., FajardoR., SnyderB., LiC., ZichnerL., LangerR., Vunjak-NovakovicG., KaplanD.L.Engineering bone-like tissue in vitro using human bone marrow stem cells and silk scaffolds. J Biomed Mater Res A, 71:25. 2004.
18.
HaiderM., CappelloJ., GhandehariH., LeongK.W.In vitro chondrogenesis of mesenchymal stem cells in recombinant silk-elastinlike hydrogels. Pharm Res, 25:692. 2008.
19.
IkeuchiM., DohiY., HoriuchiK., OhgushiH., NoshiT., YoshikawaT., YamamotoK., SugimuraM.Recombinant human bone morphogenetic protein-2 promotes osteogenesis within atelopeptide type I collagen solution by combination with rat cultured marrow cells. J Biomed Mater Res, 60:61. 2002.
BehraveshE., JoS., ZygourakisK., MikosA.G.Synthesis of in situ cross-linkable macroporous biodegradable poly(propylene fumarate-co-ethylene glycol) hydrogels. Biomacromolecules, 3:374. 2002.
22.
TrojaniC., BoukhechbaF., ScimecaJ.C., VandenbosF., MichielsJ.F., DaculsiG., BoileauP., WeissP., CarleG.F., RochetN.Ectopic bone formation using an injectable biphasic calcium phosphate/Si-HPMC hydrogel composite loaded with undifferentiated bone marrow stromal cells. Biomaterials, 27:3256. 2006.
23.
ChoiY.S., ParkS.N., SuhH.Adipose tissue engineering using mesenchymal stem cells attached to injectable PLGA spheres. Biomaterials, 26:5855. 2005.
24.
JeonO., SongS.J., KangS.W., PutnamA.J., KimB.S.Enhancement of ectopic bone formation by bone morphogenetic protein-2 released from a heparin-conjugated poly(L-lactic-co-glycolic acid) scaffold. Biomaterials, 28:2763. 2007.
25.
FuK., PackD.W., KlibanovA.M., LangerR.Visual evidence of acidic environment within degrading poly(lactic-co-glycolic acid) (PLGA) microspheres. Pharm Res, 17:100. 2000.
26.
Qingpu HouP.A.D.B.K.M.S.Injectable scaffolds for tissue regeneration. J Mater Chem, 14:1915. 2004.
27.
KretlowJ.D., KloudaL., MikosA.G.Injectable matrices and scaffolds for drug delivery in tissue engineering. Adv Drug Deliv Rev, 59:263. 2007.
28.
YamadaY., UedaM., NaikiT., TakahashiM., HataK., NagasakaT.Autogenous injectable bone for regeneration with mesenchymal stem cells and platelet-rich plasma: tissue-engineered bone regeneration. Tissue Eng, 10:955. 2004.
29.
ShimW.S., KimJ.H., ParkH., KimK., Chan KwonI., LeeD.S.Biodegradability and biocompatibility of a pH- and thermo-sensitive hydrogel formed from a sulfonamide-modified poly(epsilon-caprolactone-co-lactide)-poly(ethylene glycol)-poly(epsilon-caprolactone-co-lactide) block copolymer. Biomaterials, 27:5178. 2006.
ShimW.S., YooJ.S., BaeY.H., LeeD.S.Novel injectable pH and temperature sensitive block copolymer hydrogel. Biomacromolecules, 6:2930. 2005.
32.
YanH., SaianiA., GoughJ.E., MillerA.F.Thermoreversible protein hydrogel as cell scaffold. Biomacromolecules, 7:2776. 2006.
33.
ISO document 10993. Biological compatibility of medical devices. 5. Test for cytotoxicity: in vitro methods, 1992. www.iso.org/iso.
34.
VertM., LiS., GarreauH.More about the degradation of LA/GA-derived matrices in aqueous media. J Controlled Release, 16:15. 1991.
35.
YoshidaK., BesshoK., FujimuraK., KusumotoK., OgawaY., TaniY., IizukaT.Osteoinduction capability of recombinant human bone morphogenetic protein-2 in intramuscular and subcutaneous sites: an experimental study. J Craniomaxillofac Surg, 26:112. 1998.
36.
Kroese-DeutmanH.C., RuheP.Q., SpauwenP.H., JansenJ.A.Bone inductive properties of rhBMP-2 loaded porous calcium phosphate cement implants inserted at an ectopic site in rabbits. Biomaterials, 26:1131. 2005.
