Preparation and characterization of bovine dental pulp-derived extracellular matrix hydrogel for regenerative endodontic applications: an in vitro study.
Biomimetic scaffolds
Dental pulp-derived extracellular matrix
Dentin-pulp regeneration
Extracellular matrix hydrogels
Hyaluronic acid
Regenerative endodontics
Journal
BMC oral health
ISSN: 1472-6831
Titre abrégé: BMC Oral Health
Pays: England
ID NLM: 101088684
Informations de publication
Date de publication:
24 Oct 2024
24 Oct 2024
Historique:
received:
25
06
2024
accepted:
03
10
2024
medline:
25
10
2024
pubmed:
25
10
2024
entrez:
25
10
2024
Statut:
epublish
Résumé
The use of biological scaffolds in regenerative endodontics has gained much attention in recent years. The search for a new biomimetic scaffold that contains tissue-specific cell homing factors could lead to more predictable tissue regeneration. The aim of this study was to prepare and characterize decellularized bovine dental pulp-derived extracellular matrix (P-ECM) hydrogels for regenerative endodontic applications. Freshly extracted bovine molar teeth were collected. Bovine dental pulp tissues were harvested, and stored at -40º C. For decellularization, a 5-day protocol was implemented incorporating trypsin/EDTA, deionized water and DNase treatment. Decellularization was evaluated by DNA quantification and histological examination to assess collagen and glycosaminoglycans (GAGs) content. This was followed by the preparation of P-ECM hydrogel alone or combined with hyaluronic acid gel (P-ECM + HA). The fabricated scaffolds were then characterized using protein quantification, hydrogel topology and porosity, biodegradability, and growth factor content using Enzyme-linked immunosorbent assay (ELISA): transforming growth factor beta-1(TGF-β1), basic fibroblast growth factor (bFGF), bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF). Decellularization was histologically confirmed, and DNA content was below (50 ng/mg tissue). P-ECM hydrogel was prepared with a final ECM concentration of 3.00 mg/ml while P-ECM + HA hydrogel was prepared with a final ECM concentration of 1.5 mg/ml. Total protein content in P-ECM hydrogel was found to be (439.0 ± 123.4 µg/µl). P-ECM + HA showed sustained protein release while the P-ECM group showed gradual decreasing release. Degradation was higher in P-ECM + HA which had a significantly larger fiber diameter, while P-ECM had a larger pore area percentage. ELISA confirmed the retention and release of growth factors where P-ECM hydrogel had higher BMP-2 release, while P-ECM + HA had higher release of TGF-β1, bFGF, and VEGF. Both P-ECM and P-ECM + HA retained their bioactive properties demonstrating a potential role as functionalized scaffolds for regenerative endodontic procedures.
Sections du résumé
BACKGROUND
BACKGROUND
The use of biological scaffolds in regenerative endodontics has gained much attention in recent years. The search for a new biomimetic scaffold that contains tissue-specific cell homing factors could lead to more predictable tissue regeneration. The aim of this study was to prepare and characterize decellularized bovine dental pulp-derived extracellular matrix (P-ECM) hydrogels for regenerative endodontic applications.
METHODS
METHODS
Freshly extracted bovine molar teeth were collected. Bovine dental pulp tissues were harvested, and stored at -40º C. For decellularization, a 5-day protocol was implemented incorporating trypsin/EDTA, deionized water and DNase treatment. Decellularization was evaluated by DNA quantification and histological examination to assess collagen and glycosaminoglycans (GAGs) content. This was followed by the preparation of P-ECM hydrogel alone or combined with hyaluronic acid gel (P-ECM + HA). The fabricated scaffolds were then characterized using protein quantification, hydrogel topology and porosity, biodegradability, and growth factor content using Enzyme-linked immunosorbent assay (ELISA): transforming growth factor beta-1(TGF-β1), basic fibroblast growth factor (bFGF), bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF).
RESULTS
RESULTS
Decellularization was histologically confirmed, and DNA content was below (50 ng/mg tissue). P-ECM hydrogel was prepared with a final ECM concentration of 3.00 mg/ml while P-ECM + HA hydrogel was prepared with a final ECM concentration of 1.5 mg/ml. Total protein content in P-ECM hydrogel was found to be (439.0 ± 123.4 µg/µl). P-ECM + HA showed sustained protein release while the P-ECM group showed gradual decreasing release. Degradation was higher in P-ECM + HA which had a significantly larger fiber diameter, while P-ECM had a larger pore area percentage. ELISA confirmed the retention and release of growth factors where P-ECM hydrogel had higher BMP-2 release, while P-ECM + HA had higher release of TGF-β1, bFGF, and VEGF.
CONCLUSIONS
CONCLUSIONS
Both P-ECM and P-ECM + HA retained their bioactive properties demonstrating a potential role as functionalized scaffolds for regenerative endodontic procedures.
Identifiants
pubmed: 39448989
doi: 10.1186/s12903-024-05004-z
pii: 10.1186/s12903-024-05004-z
doi:
Substances chimiques
Hydrogels
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
1281Informations de copyright
© 2024. The Author(s).
Références
Kim S, Malek M, Sigurdsson A, Lin L, Kahler B. Regenerative endodontics: a comprehensive review. Int Endod J. 2018;51(12):1367–88.
pubmed: 29777616
doi: 10.1111/iej.12954
Galler KM, Widbiller M. Perspectives for cell-homing approaches to engineer dental pulp. J Endod. 2017;43(9):S40–5.
pubmed: 28778503
doi: 10.1016/j.joen.2017.06.008
Eramo S, Natali A, Pinna R, Milia E. Dental pulp regeneration via cell homing. Int Endod J. 2018;51(4):405–19.
pubmed: 29047120
doi: 10.1111/iej.12868
Bakhtiar H, Mazidi SA, Mohammadi Asl S, Ellini M, Moshiri A, Nekoofar M, et al. The role of stem cell therapy in regeneration of dentine-pulp complex: a systematic review. Prog Biomater. 2018;7:249–68.
pubmed: 30267369
pmcid: 6304177
doi: 10.1007/s40204-018-0100-7
Mobarak A, Genena S, Zaazou A, Mokhless NA, Moussa SM. Regenerative pulpotomy as a novel technique for treatment of permanent mature molars diagnosed with irreversible pulpitis using platelet-rich fibrin: a case series study. Alexandria Dent J. 2021;46(1):129–35.
Chen G, Ushida T, Tateishi T. Scaffold design for tissue engineering. Macromol Biosci. 2002;2(2):67–77.
doi: 10.1002/1616-5195(20020201)2:2<67::AID-MABI67>3.0.CO;2-F
Bakhtiar H, Pezeshki-Modaress M, Kiaipour Z, Shafiee M, Ellini MR, Mazidi A, et al. Pulp ECM-derived macroporous scaffolds for stimulation of dental-pulp regeneration process. Dent Mater. 2020;36(1):76–87.
pubmed: 31735424
doi: 10.1016/j.dental.2019.10.011
Matoug-Elwerfelli M, Duggal M, Nazzal H, Esteves F, Raïf E. A biocompatible decellularized pulp scaffold for regenerative endodontics. Int Endod J. 2018;51(6):663–73.
pubmed: 29197101
doi: 10.1111/iej.12882
Badylak SF. The extracellular matrix as a biologic scaffold material. Biomaterials. 2007;28(25):3587–93.
pubmed: 17524477
doi: 10.1016/j.biomaterials.2007.04.043
Crapo PM, Gilbert TW, Badylak SF. An overview of tissue and whole organ decellularization processes. Biomaterials. 2011;32(12):3233–43.
pubmed: 21296410
pmcid: 3084613
doi: 10.1016/j.biomaterials.2011.01.057
Alqahtani Q, Zaky S, Patil A, Beniash E, Ray H, Sfeir C. Decellularized swine dental pulp tissue for regenerative root canal therapy. J Dent Res. 2018;97(13):1460–7.
pubmed: 30067420
doi: 10.1177/0022034518785124
Taylor PM, Cass AE, Yacoub MH. Extracellular matrix scaffolds for tissue engineering heart valves. Prog Pediatr Cardiol. 2006;21(2):219–25.
doi: 10.1016/j.ppedcard.2005.11.010
Davidenko N, Campbell J, Thian E, Watson C, Cameron R. Collagen–hyaluronic acid scaffolds for adipose tissue engineering. Acta Biomater. 2010;6(10):3957–68.
pubmed: 20466086
doi: 10.1016/j.actbio.2010.05.005
Hu L, Gao Z, Xu J, Zhu Z, Fan Z, Zhang C et al. Decellularized swine dental pulp as a bioscaffold for pulp regeneration. BioMed Research International. 2017;2017.
Ramm R, Goecke T, Theodoridis K, Hoeffler K, Sarikouch S, Findeisen K, et al. Decellularization combined with enzymatic removal of N-linked glycans and residual DNA reduces inflammatory response and improves performance of porcine xenogeneic pulmonary heart valves in an ovine in vivo model. Xenotransplantation. 2020;27(2):e12571.
pubmed: 31769101
doi: 10.1111/xen.12571
Yazdanian M, Arefi AH, Alam M, Abbasi K, Tebyaniyan H, Tahmasebi E, et al. Decellularized and biological scaffolds in dental and craniofacial tissue engineering: a comprehensive overview. J Mater Res Technol. 2021;15:1217–51.
doi: 10.1016/j.jmrt.2021.08.083
Sharma LA, Love RM. 22 - scaffolds for regeneration of the pulp–dentine complex. In: Mozafari M, Sefat F, Atala A, editors. Handbook of tissue Engineering Scaffolds. Volume One: Woodhead Publishing; 2019. pp. 459–78.
doi: 10.1016/B978-0-08-102563-5.00022-8
Chang B, Ahuja N, Ma C, Liu X. Injectable scaffolds: Preparation and application in dental and craniofacial regeneration. Mater Sci Engineering: R: Rep. 2017;111:1–26.
doi: 10.1016/j.mser.2016.11.001
Atila D, Chen C-Y, Lin C-P, Lee Y-L, Hasirci V, Tezcaner A, et al. In vitro evaluation of injectable tideglusib-loaded hyaluronic acid hydrogels incorporated with Rg1-loaded chitosan microspheres for vital pulp regeneration. Carbohydr Polym. 2022;278:118976.
pubmed: 34973790
doi: 10.1016/j.carbpol.2021.118976
Chrepa V, Austah O, Diogenes A. Evaluation of a commercially available hyaluronic acid hydrogel (restylane) as injectable scaffold for dental pulp regeneration: an in vitro evaluation. J Endod. 2017;43(2):257–62.
pubmed: 28041686
doi: 10.1016/j.joen.2016.10.026
Pardue EL, Ibrahim S, Ramamurthi A. Role of hyaluronan in angiogenesis and its utility to angiogenic tissue engineering. Organogenesis. 2008;4(4):203–14.
pubmed: 19337400
pmcid: 2634325
doi: 10.4161/org.4.4.6926
Luo Z, Wang Y, Li J, Wang J, Yu Y, Zhao Y. Tailoring hyaluronic acid hydrogels for biomedical applications. Adv Funct Mater. 2023;33(49):2306554.
doi: 10.1002/adfm.202306554
Rao SS, Prabhu A, Kudkuli J, Surya S, Rekha P. Hyaluronic acid sustains platelet stability with prolonged growth factor release and accelerates wound healing by enhancing proliferation and collagen deposition in diabetic mice. J Drug Deliv Sci Technol. 2022;67:102898.
doi: 10.1016/j.jddst.2021.102898
Ilio K, FurukawaKI TE. Hyaluronic acid induces the release of growth factors from platelet-rich plasma Asia-Pacific. J Sports Med Arthrosc Rehabilitation Technol. 2016;4:27.
Nagendrababu V, Murray PE, Ordinola-Zapata R, Peters OA, Rôças IN, Siqueira JF Jr, et al. PRILE 2021 guidelines for reporting laboratory studies in Endodontology: a consensus‐based development. Int Endod J. 2021;54(9):1482–90.
pubmed: 33938010
doi: 10.1111/iej.13542
Matoug-Elwerfelli M, Nazzal H, Raif EM, Wilshaw S-P, Esteves F, Duggal M. Ex-vivo recellularisation and stem cell differentiation of a decellularised rat dental pulp matrix. Sci Rep. 2020;10(1):21553.
pubmed: 33299073
pmcid: 7725831
doi: 10.1038/s41598-020-78477-x
Bakhtiar H, Rajabi S, Pezeshki-Modaress M, Ellini MR, Panahinia M, Alijani S, et al. Optimizing methods for bovine dental pulp decellularization. J Endod. 2021;47(1):62–8.
pubmed: 33049226
doi: 10.1016/j.joen.2020.08.027
Massensini AR, Ghuman H, Saldin LT, Medberry CJ, Keane TJ, Nicholls FJ, et al. Concentration-dependent rheological properties of ECM hydrogel for intracerebral delivery to a stroke cavity. Acta Biomater. 2015;27:116–30.
pubmed: 26318805
pmcid: 4609617
doi: 10.1016/j.actbio.2015.08.040
Pk S. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985;150:76–85.
doi: 10.1016/0003-2697(85)90442-7
Pouliot RA, Link PA, Mikhaiel NS, Schneck MB, Valentine MS, Kamga Gninzeko FJ, et al. Development and characterization of a naturally derived lung extracellular matrix hydrogel. J Biomedical Mater Res Part A. 2016;104(8):1922–35.
doi: 10.1002/jbm.a.35726
Dong L, Wang S-J, Zhao X-R, Zhu Y-F, Yu J-K. 3D-printed poly (ε-caprolactone) scaffold integrated with cell-laden chitosan hydrogels for bone tissue engineering. Sci Rep. 2017;7(1):13412.
pubmed: 29042614
pmcid: 5645328
doi: 10.1038/s41598-017-13838-7
Medberry CJ, Crapo PM, Siu BF, Carruthers CA, Wolf MT, Nagarkar SP, et al. Hydrogels derived from central nervous system extracellular matrix. Biomaterials. 2013;34(4):1033–40.
pubmed: 23158935
doi: 10.1016/j.biomaterials.2012.10.062
Holiel AA, Sedek EM. Marginal adaptation, physicochemical and rheological properties of treated dentin matrix hydrogel as a novel injectable pulp capping material for dentin regeneration. BMC Oral Health. 2023;23(1):938.
pubmed: 38017480
pmcid: 10683231
doi: 10.1186/s12903-023-03677-6
Alonso JM, Andrade del Olmo J, Perez Gonzalez R, Saez-Martinez V. Injectable hydrogels: from laboratory to industrialization. Polymers. 2021;13(4):650.
pubmed: 33671648
pmcid: 7926321
doi: 10.3390/polym13040650
Xie C, Liu G, Wang L, Yang Q, Liao F, Yang X, et al. Synthesis and Properties of Injectable Hydrogel for tissue filling. Pharmaceutics. 2024;16(3):430.
pubmed: 38543325
pmcid: 10975320
doi: 10.3390/pharmaceutics16030430
Schneider CA, Rasband WS, Eliceiri KW. NIH Image to ImageJ: 25 years of image analysis. Nat Methods. 2012;9(7):671–5.
pubmed: 22930834
pmcid: 5554542
doi: 10.1038/nmeth.2089
Saldin LT, Cramer MC, Velankar SS, White LJ, Badylak SF. Extracellular matrix hydrogels from decellularized tissues: structure and function. Acta Biomater. 2017;49:1–15.
pubmed: 27915024
doi: 10.1016/j.actbio.2016.11.068
Han Y, Xu J, Chopra H, Zhang Z, Dubey N, Dissanayaka W, et al. Injectable tissue-specific Hydrogel System for pulp–dentin regeneration. J Dent Res. 2024;103(4):398–408.
pubmed: 38410924
doi: 10.1177/00220345241226649
Qu T, Jing J, Ren Y, Ma C, Feng JQ, Yu Q, et al. Complete pulpodentin complex regeneration by modulating the stiffness of biomimetic matrix. Acta Biomater. 2015;16:60–70.
pubmed: 25644448
doi: 10.1016/j.actbio.2015.01.029
Lu Q, Pandya M, Rufaihah AJ, Rosa V, Tong HJ, Seliktar D, et al. Modulation of dental pulp stem cell odontogenesis in a tunable PEG-fibrinogen hydrogel system. Stem Cells Int. 2015;2015(1):525367.
pubmed: 26124841
pmcid: 4466494
Orti V, Collart-Dutilleul P-Y, Piglionico S, Pall O, Cuisinier F, Panayotov I. Pulp regeneration concepts for nonvital teeth: from tissue engineering to clinical approaches. Tissue Eng Part B: Reviews. 2018;24(6):419–42.
doi: 10.1089/ten.teb.2018.0073
Yang J, Yuan G, Chen Z. Pulp regeneration: current approaches and future challenges. Front Physiol. 2016;7:164062.
doi: 10.3389/fphys.2016.00058
Elnawam H, Abdelmougod M, Mobarak A, Hussein M, Aboualmakarem H, Girgis M, et al. Regenerative endodontics and minimally invasive dentistry: intertwining paths crossing over into clinical translation. Front Bioeng Biotechnol. 2022;10:837639.
pubmed: 35211465
pmcid: 8860982
doi: 10.3389/fbioe.2022.837639
Schenke-Layland K, Vasilevski O, Opitz F, König K, Riemann I, Halbhuber K, et al. Impact of decellularization of xenogeneic tissue on extracellular matrix integrity for tissue engineering of heart valves. J Struct Biol. 2003;143(3):201–8.
pubmed: 14572475
doi: 10.1016/j.jsb.2003.08.002
Gilpin A, Yang Y. Decellularization strategies for regenerative medicine: from processing techniques to applications. BioMed research international. 2017;2017.
Wang Y, Bao J, Wu Q, Zhou Y, Li Y, Wu X, et al. Method for perfusion decellularization of porcine whole liver and kidney for use as a scaffold for clinical-scale bioengineering engrafts. Xenotransplantation. 2015;22(1):48–61.
pubmed: 25291435
doi: 10.1111/xen.12141
Lee DJ, Miguez P, Kwon J, Daniel R, Padilla R, Min S, et al. Decellularized pulp matrix as scaffold for mesenchymal stem cell mediated bone regeneration. J Tissue Eng. 2020;11:2041731420981672.
pubmed: 33414903
pmcid: 7750895
doi: 10.1177/2041731420981672
Song J, Takimoto K, Jeon M, Vadakekalam J, Ruparel N, Diogenes A. Decellularized human dental pulp as a scaffold for regenerative endodontics. J Dent Res. 2017;96(6):640–6.
pubmed: 28196330
doi: 10.1177/0022034517693606
Uhl FE, Zhang F, Pouliot RA, Uriarte JJ, Enes SR, Han X, et al. Functional role of glycosaminoglycans in decellularized lung extracellular matrix. Acta Biomater. 2020;102:231–46.
pubmed: 31751810
doi: 10.1016/j.actbio.2019.11.029
Wang Z, Wang Z, Lu WW, Zhen W, Yang D, Peng S. Novel biomaterial strategies for controlled growth factor delivery for biomedical applications. NPG Asia Mater. 2017;9(10):e435–e.
doi: 10.1038/am.2017.171
Jung Y-s, Park W, Park H, Lee D-K, Na K. Thermo-sensitive injectable hydrogel based on the physical mixing of hyaluronic acid and pluronic F-127 for sustained NSAID delivery. Carbohydr Polym. 2017;156:403–8.
pubmed: 27842839
doi: 10.1016/j.carbpol.2016.08.068
Nyambat B, Manga YB, Chen C-H, Gankhuyag U, Pratomo WPA, Kumar Satapathy M, et al. New insight into natural extracellular matrix: genipin cross-linked adipose-derived stem cell extracellular matrix gel for tissue engineering. Int J Mol Sci. 2020;21(14):4864.
pubmed: 32660134
pmcid: 7402347
doi: 10.3390/ijms21144864
Bakhtiar H, Mousavi MR, Rajabi S, Pezeshki-Modaress M, Ayati A, Ashoori A, et al. Fabrication and characterization of a novel injectable human amniotic membrane hydrogel for dentin-pulp complex regeneration. Dent Mater. 2023;39(8):718–28.
pubmed: 37393152
doi: 10.1016/j.dental.2023.06.008
Nowwarote N, Petit S, Ferre FC, Dingli F, Laigle V, Loew D, et al. Extracellular matrix derived from dental pulp stem cells promotes mineralization. Front Bioeng Biotechnol. 2022;9:740712.
pubmed: 35155398
pmcid: 8829122
doi: 10.3389/fbioe.2021.740712
Silva PAO, Lima SMF, Freire MS, Murad AM, Franco OL, Rezende TMB. Proteomic analysis of human dental pulp in different clinical diagnosis. Clin Oral Invest. 2021;25:3285–95.
doi: 10.1007/s00784-020-03660-3
Kim JY, Xin X, Moioli EK, Chung J, Lee CH, Chen M, et al. Regeneration of dental-pulp-like tissue by chemotaxis-induced cell homing. Tissue Eng Part A. 2010;16(10):3023–31.
pubmed: 20486799
pmcid: 2947424
doi: 10.1089/ten.tea.2010.0181
Hill RC, Calle EA, Dzieciatkowska M, Niklason LE, Hansen KC. Quantification of extracellular matrix proteins from a rat lung scaffold to provide a molecular readout for tissue engineering*[S]. Mol Cell Proteom. 2015;14(4):961–73.
doi: 10.1074/mcp.M114.045260
de Castro Brás LE, Ramirez TA, DeLeon-Pennell KY, Chiao YA, Ma Y, Dai Q, et al. Texas 3-step decellularization protocol: looking at the cardiac extracellular matrix. J Proteom. 2013;86:43–52.
doi: 10.1016/j.jprot.2013.05.004
Bakhtiar H, Ashoori A, Rajabi S, Pezeshki-Modaress M, Ayati A, Mousavi MR, et al. Human amniotic membrane extracellular matrix scaffold for dental pulp regeneration in vitro and in vivo. Int Endod J. 2022;55(4):374–90.
pubmed: 34923640
doi: 10.1111/iej.13675
Gathani KM, Raghavendra SS. Scaffolds in regenerative endodontics: a review. Dent Res J. 2016;13(5):379–86.
doi: 10.4103/1735-3327.192266
Shi Y, Wang Y, Shan Z, Gao Z. Decellularized rat submandibular gland as an alternative scaffold for dental pulp regeneration. Front Bioeng Biotechnol. 2023;11:1148532.
pubmed: 37152652
pmcid: 10160494
doi: 10.3389/fbioe.2023.1148532
Albuquerque M, Valera M, Nakashima M, Nör J, Bottino M. Tissue-engineering-based strategies for regenerative endodontics. J Dent Res. 2014;93(12):1222–31.
pubmed: 25201917
pmcid: 4237634
doi: 10.1177/0022034514549809
Sadaghiani L, Alshumrani AM, Gleeson HB, Ayre WN, Sloan AJ. Growth factor release and dental pulp stem cell attachment following dentine conditioning: an in vitro study. Int Endod J. 2022;55(8):858–69.
pubmed: 35638345
pmcid: 9541952
doi: 10.1111/iej.13781
Xie Z, Shen Z, Zhan P, Yang J, Huang Q, Huang S, et al. Functional dental pulp regeneration: basic research and clinical translation. Int J Mol Sci. 2021;22(16):8991.
pubmed: 34445703
pmcid: 8396610
doi: 10.3390/ijms22168991
Johnson KE, Wilgus TA. Vascular endothelial growth factor and angiogenesis in the regulation of cutaneous wound repair. Adv Wound care. 2014;3(10):647–61.
doi: 10.1089/wound.2013.0517
Beheshtizadeh N, Gharibshahian M, Bayati M, Maleki R, Strachan H, Doughty S, et al. Vascular endothelial growth factor (VEGF) delivery approaches in regenerative medicine. Biomed Pharmacother. 2023;166:115301.
pubmed: 37562236
doi: 10.1016/j.biopha.2023.115301
Tran-Hung L, Laurent P, Camps J, About I. Quantification of angiogenic growth factors released by human dental cells after injury. Arch Oral Biol. 2008;53(1):9–13.
pubmed: 17764655
doi: 10.1016/j.archoralbio.2007.07.001
Laurent P, Camps J, About I. BiodentineTM induces TGF-β1 release from human pulp cells and early dental pulp mineralization. Int Endod J. 2012;45(5):439–48.
pubmed: 22188368
doi: 10.1111/j.1365-2591.2011.01995.x
Ilić J, Radović K, Roganović J, Brković B, Stojić D. The levels of vascular endothelial growth factor and bone morphogenetic protein 2 in dental pulp tissue of healthy and diabetic patients. J Endod. 2012;38(6):764–8.
pubmed: 22595109
doi: 10.1016/j.joen.2012.03.016
Kim HD, Valentini RF. Retention and activity of BMP-2 in hyaluronic acid‐based scaffolds in vitro. J Biomed Mater Res. 2002;59(3):573–84.