Allogenic demineralized bone matrix (DBM) is widely used for bone repair and regeneration due to its osteoinductivity and osteoconductivity. The present study utilized acellular dermis microfibers to improve the DBM’s clinical handling properties and to enhance bone regeneration. Donated human cadaver skin was de-epidermized and decellularized to be acellular dermal matrix (ADM), which was further processed into microfibers. Donated human bone was micronized and partially demineralized (∼30% calcium removal) for optimal bone regeneration. A flexible ADM/DBM composite foam was fabricated with ADM microfibers and DBM particles. Structural analysis found that the ADM/DBM composite foam had proper porosity with interconnected micropores and rapid wettability, and good stability upon cyclic compressions, whereas cytotoxicity test, in vitro collagenase degradation, and rat subcutaneous implantation showed good biocompatibility and biodegradability. The composite foam, used for in vitro coculture, significantly increased the alkaline phosphatase activity of C2C12 cells and upregulated the expression of osteogenesis-related genes of human umbilical cord mesenchymal stem cells. Using the rat Φ8 mm calvarium defect repair model, the ADM/DBM composite foam demonstrated superior osteogenicity by rapidly inducing new bone formation and achieving complete closure of the bone defects, as compared with the commercially available bone graft for skull repair (SkuHeal). Therefore, the ADM/DBM composite foam holds promise as a superior DBM-based product for repairing critical bone defects.
Impact Statement
The study utilized acellular dermis microfibers to improve the demineralized bone matrix’s (DBM) clinical handling properties and to enhance bone regeneration. The acellular dermal matrix (ADM)/DBM composite foam exhibited proper porosity with interconnected micropores and rapid wettability, excellent stability upon cyclic compressions, good biocompatibility and biodegradability, and osteogenic bioactivity in in vitro cell coculture. In vivo testing using a rat Φ8 mm calvarium defect repair model showed that the ADM/DBM composite foam significantly induced new bone formation and achieved complete closure of critical bone defects. Our results suggest that the ADM/DBM composite foam holds promise as a superior DBM-based product for use as bone graft substitute.
Get full access to this article
View all access options for this article.
References
1.
van der StokJ, HartholtKA, SchoenmakersDAL, et al.The available evidence on demineralised bone matrix in trauma and orthopaedic surgery: A systematic review. Bone Joint Res, 2017; 6(7):423–432; doi: 10.1302/2046-3758.67.BJR-2017-0027.R1
2.
WangW, YeungKWK. Bone grafts and biomaterials substitutes for bone defect repair: A review. Bioact Mater, 2017; 2(4):224–247; doi: 10.1016/j.bioactmat.2017.05.007
3.
GruskinE, DollBA, FutrellFW, et al.Demineralized bone matrix in bone repair: History and use. Adv Drug Deliv Rev, 2012; 64(12):1063–1077; doi: 10.1016/j.addr.2012.06.008
4.
ZhangH, YangL, YangXG, et al.Demineralized bone matrix carriers and their clinical applications: An overview. Orthop Surg, 2019; 11(5):725–737; doi: 10.1111/os.12509
5.
IwataH, SakanoS, ItohT, et al.Demineralized bone matrix and native bone morphogenetic protein in orthopaedic surgery. Clin Orthop Relat Res, 2002; 395(395):99–109; doi: 10.1097/00003086-200202000-00010
6.
ChenS, CleavengerG, PhelpsD, et al.Demineralized bone fibers and preparation thereof. US Patent 11,033,656 B2. LifeNet Health: U.S.; 2021.
7.
PrewettAB, StikeleatherRC. Process for producing flowable osteogenic composition containing demineralized bone particles. US Patent 5,510,396A. LifeNet Health: U.S.; 1996.
8.
WolfinbargerLJr, Process for producing osteoinductive bone, and osteoinductive bone produced thereby. US Patent 6,305,379 B1. LifeNet Health: U.S.; 2001.
9.
ConnorJ, QiuQ-Q. Bone graft composition. US Patent 2007/0248575 A1. LifeCell Corp U.S.; 2007.
10.
OsborneJ, NormanK, MayeT, et al.American Association of Tissue Banks: Standards for tissue banking. 14th ed. American Association of Tissue Banks; 2016.
11.
ZhangM, PowersRMJr, WolfinbargerLJr, Effect(s) of the demineralization process on the osteoinductivity of demineralized bone matrix. J Periodontol, 1997; 68(11):1085–1092; doi: 10.1902/jop.1997.68.11.1085
12.
HeroldRW, PashleyDH, CueninMF, et al.The effects of varying degrees of allograft decalcification on cultured porcine osteoclast cells. J Periodontol, 2002; 73(2):213–219; doi: 10.1902/jop.2002.73.2.213
13.
TuronisJW, McPhersonJC, CueninMF, et al.The effect of residual calcium in decalcified freeze-dried bone allograft in a critical-sized defect in the Rattus norvegicus calvarium. J Oral Implantol, 2006; 32(2):55–62; doi: 10.1563/780.1
14.
WolfinbargerLJr, EisenlohrLM, RuthK. Demineralized bone matrix: maximizing new bone formation for successful bone implantation. In: Musculoskeletal Tissue Regeneration: Biological Materials and Methods. (PietrzakWS. eds.) Humana Press: Totowa, NJ; 2008; pp. 93–117.
15.
HonsawekS, PowersRM, WolfinbargerL. Extractable bone morphogenetic protein and correlation with induced new bone formation in an in vivo assay in the athymic mouse model. Cell Tissue Bank, 2005; 6(1):13–23; doi: 10.1007/s10561-005-1445-4
16.
MaT, RenD, WangJ, et al.Enhanced osteogenicity of the demineralized bone-dermal matrix composite by the optimal partial demineralization for sustained release of bioactive molecules. J Biomed Mater Res B Appl Biomater, 2024; 112(1):e35358; doi: 10.1002/jbm.b.35358
17.
TianM, YangZ, KuwaharaK, et al.Delivery of demineralized bone matrix powder using a thermogelling chitosan carrier. Acta Biomater, 2012; 8(2):753–762; doi: 10.1016/j.actbio.2011.10.030
18.
ShehadiJA, ElzeinSM. Review of commercially available demineralized bone matrix products for spinal fusions: A selection paradigm. Surg Neurol Int, 2017; 8:203; doi: 10.4103/sni.sni_155_17
19.
CohenJD, KanimLE, TronitsAJ, et al.Allografts and spinal fusion. Int J Spine Surg, 2021; 15(s1):68–93; doi: 10.14444/8056
20.
BordenM. Bone grafts based on demineralized bone matrix. In: Bone Graft Substitutes and Bone Regenerative Engineering. (LaurencinCT, JiangT. eds.) ASTM International: West Conshohocken, PA; 2014; pp. 49–72.
21.
AcarturkTO, HollingerJO. Commercially available demineralized bone matrix compositions to regenerate calvarial critical-sized bone defects. Plast Reconstr Surg, 2006; 118(4):862–873; doi: 10.1097/01.prs.0000232385.81219.87
22.
WangJC, AlanayA, MarkD, et al.A comparison of commercially available demineralized bone matrix for spinal fusion. Eur Spine J, 2007; 16(8):1233–1240; doi: 10.1007/s00586-006-0282-x
23.
BostromMP, YangX, KennanM, et al.An unexpected outcome during testing of commercially available demineralized bone graft materials: How safe are the nonallograft components? Spine (Phila Pa 1976), 2001; 26(13):1425–1428; doi: 10.1097/00007632-200107010-00007
24.
WangJC, KanimLE, NagakawaIS, et al.Dose-dependent toxicity of a commercially available demineralized bone matrix material. Spine (Phila Pa 1976), 2001; 26(13):1429–1435; discussion 1435-6; doi: 10.1097/00007632-200107010-00008
25.
ElgaliI, OmarO, DahlinC, et al.Guided bone regeneration: Materials and biological mechanisms revisited. Eur J Oral Sci, 2017; 125(5):315–337; doi: 10.1111/eos.12364
26.
TarnowDP, FletcherP. Histologic evidence of the ability of dermal allograft to function as a barrier during guided bone regeneration: A case report. Clin Adv Periodontics, 2015; 5(3):201–207; doi: 10.1902/cap.2014.130096
27.
LiW, ShengK, RanY, et al.Transformation of acellular dermis matrix with dicalcium phosphate into 3D porous scaffold for bone regeneration. J Biomater Sci Polym Ed, 2021; 32(16):2071–2087; doi: 10.1080/09205063.2021.1955817
28.
QiuQQ, ShihMS, StockK, et al.Evaluation of DBM/AM composite as a graft substitute for posterolateral lumbar fusion. J Biomed Mater Res B Appl Biomater, 2007; 82(1):239–245; doi: 10.1002/jbm.b.30726
29.
QiuQQ, MendenhallHV, GarlickDS, et al.Evaluation of bone regeneration at critical-sized calvarial defect by DBM/AM composite. J Biomed Mater Res B Appl Biomater, 2007; 81(2):516–523; doi: 10.1002/jbm.b.30692
30.
KimJ, LeeK-W, AhnJ-H, et al.Osteoinductivity depends on the ratio of demineralized bone matrix to acellular dermal matrix in defects in rat skulls. Tissue Eng Regen Med, 2013; 10(5):246–251; doi: 10.1007/s13770-012-1083-4
31.
Ignat’evaNY, DanilovNA, AverkievSV, et al.Determination of hydroxyproline in tissues and the evaluation of the collagen content of the tissues. J Anal Chem, 2007; 62(1):51–57; doi: 10.1134/s106193480701011x
32.
AlomN, PetoH, KirkhamGR, et al.Bone extracellular matrix hydrogel enhances osteogenic differentiation of C2C12 myoblasts and mouse primary calvarial cells. J Biomed Mater Res B Appl Biomater, 2018; 106(2):900–908; doi: 10.1002/jbm.b.33894
33.
HanB, TangB, NimniME. Quantitative and sensitive in vitro assay for osteoinductive activity of demineralized bone matrix. J Orthop Res, 2003; 21(4):648–654; doi: 10.1016/s0736-0266(03)00005-6
34.
WangL, DormerNH, BonewaldLF, et al.Osteogenic differentiation of human umbilical cord mesenchymal stromal cells in polyglycolic acid scaffolds. Tissue Eng Part A, 2010; 16(6):1937–1948; doi: 10.1089/ten.TEA.2009.0706
35.
NooriA, AshrafiSJ, Vaez-GhaemiR, et al.A review of fibrin and fibrin composites for bone tissue engineering. Int J Nanomedicine, 2017; 12:4937–4961; doi: 10.2147/IJN.S124671
36.
CrapoPM, GilbertTW, BadylakSF. An overview of tissue and whole organ decellularization processes. Biomaterials, 2011; 32(12):3233–3243; doi: 10.1016/j.biomaterials.2011.01.057
37.
MaT, SunWQ, WangJ. Thermophysical stability and biodegradability of regenerative tissue scaffolds. J Therm Anal Calorim, 2022; 147(16):8757–8764; doi: 10.1007/s10973-021-11184-5
38.
SunWQ, LeungP. Calorimetric study of extracellular tissue matrix degradation and instability after gamma irradiation. Acta Biomater, 2008; 4(4):817–826; doi: 10.1016/j.actbio.2008.02.006
39.
RobertsTT, RosenbaumAJ. Bone grafts, bone substitutes and orthobiologics: The bridge between basic science and clinical advancements in fracture healing. Organogenesis, 2012; 8(4):114–124; doi: 10.4161/org.23306
40.
GreenwaldAS, BodenSD, GoldbergVM, et al.American Academy of Orthopaedic Surgeons. The Committee on Biological Implants. Bone-graft substitutes: Facts, fictions, and applications. J Bone Joint Surg Am, 2001; 83-A Suppl 2 Pt 2:98–103; doi: 10.2106/00004623-200100022-00007
SteinicheT, HaugeEM. Normal structure and function of bone. In: Handbook of Histology Methods for Bone and Cartilage. (AnYH, MartinKL. eds.) Humana Press: Totowa, NJ; 2003; pp. 59–72.
43.
QiuZ-Y, CuiY, WangX-M. Natural bone tissue and its biomimetic. In: Mineralized Collagen Bone Graft Substitutes. (WangX-M, QiuZ-Y, CuiH. eds.) Woodhead Publishing: Duxford; 2019; pp. 1–22.
44.
BrobergA, NissinenL, PotilaM, et al.Three-dimensional collagen regulates collagen gene expression by a mechanism that requires serine/threonine kinases and is independent of mechanical contraction. Biochem Biophys Res Commun, 2001; 280(1):328–333; doi: 10.1006/bbrc.2000.4132
45.
LangholzO, RöckelD, MauchC, et al.Collagen and collagenase gene expression in three-dimensional collagen lattices are differentially regulated by alpha 1 beta 1 and alpha 2 beta 1 integrins. J Cell Biol, 1995; 131(6 Pt 2):1903–1915; doi: 10.1083/jcb.131.6.1903
46.
MagriKA, BenedictMR, EwtonDZ, et al.Negative feedback regulation of insulin-like growth factor-II gene expression in differentiating myoblasts in vitro. Endocrinology, 1994; 135(1):53–62; doi: 10.1210/endo.135.1.8013391
47.
SpicerPP, KretlowJD, YoungS, et al.Evaluation of bone regeneration using the rat critical size calvarial defect. Nat Protoc, 2012; 7(10):1918–1929; doi: 10.1038/nprot.2012.113
48.
VajgelA, MardasN, FariasBC, et al.A systematic review on the critical size defect model. Clin Oral Implants Res, 2014; 25(8):879–893; doi: 10.1111/clr.12194
49.
DialloAM, RotaS, BoissiereM, et al.Osteoformation potential of an allogenic partially demineralized bone matrix in critical-size defects in the rat calvarium. Mater Sci Eng C Mater Biol Appl, 2021; 127:112207; doi: 10.1016/j.msec.2021.112207
50.
Grossi-OliveiraG, FaveraniLP, MendesBC, et al.Comparative evaluation of bone repair with four different bone substitutes in critical size defects. Int J Biomater, 2020; 2020:5182845; doi: 10.1155/2020/5182845
