Cytotoxic effect of natural cuttlefish bone xenograft: an in vitro and in vivo study
BACKGROUND Commercialized synthetic bone grafts are commonly used to replace the bone defect. Cuttlefish bone is naturally available and widely studied, but the specific cytotoxicity test has not been conducted. This study aimed to evaluate the cytotoxicity of the xenograft compared to commercial grafts.
METHODS We performed an in vitro test evaluating the viability of human mesenchymal stem cells (hMSCs) when cultured for 48 hours with the tested materials (cuttlefish bone graft and fabricated PerOssal®). The trypsinized mitochondrial activity of the viable hMSC was assayed based on colorimetry of the formazan color change. The tested material was considered nontoxic if >70% of the hMSCs were viable. The in vivo cytotoxic effect was evaluated by implanting the graft material in the femoral muscle of New Zealand (NZ) white rabbits. Nine rabbits were used in each test (cuttlefish bone, PerOssal®, and NaCl 0.9%). The systemic acute pyrogenic effect was evaluated based on 72 hours body weight changes and rectal temperature changes every 30 min in the first 3 hours and 72 hours post-implantation.
RESULTS The mean percentage of hMSC viability when cultured with cuttlefish bone graft and PerOssal® was comparable (93.47% and 105.37%, respectively, p = 0.240). The in vivo cytotoxicity on NZ rabbit was similar between all tested materials, as shown by the minor changes in body weight (<10% body weight, p = 0.391) and rectal temperature (<0.5°C, p = 0.127).
CONCLUSIONS Cuttlefish bone xenograft and fabricated PerOssal® have a similar non-cytotoxic effects on hMSCs and non-pyrogenic systemic effects on rabbits.
Hung NN. Basic knowledge of bone grafting. In: Zorzi A, editor. Bone grafting. London: IntechOpen; 2012. p. 11-38. Available from: https://www.intechopen.com/books/bone-grafting/basicknowledge-of-bone-grafting.
Kinaci A, Neuhaus V, Ring DC. Trends in bone graft use in the United States. Orthopedics. 2014;37(9):e783-8. https://doi.org/10.3928/01477447-20140825-54
Finkemeier CG. Bone-grafting and bone-graft substitutes. J Bone Joint Surg Am. 2002;84(3):454-64. https://doi.org/10.2106/00004623-200203000-00020
Pape HC, Evans A, Kobbe P. Autologous bone graft: properties and techniques. J Orthop Trauma. 2010;24Suppl 1:S36-40. https://doi.org/10.1097/BOT.0b013e3181cec4a1
Sarin P, Lee SJ, Apostolov ZD, Kriven WM. Porous biphasic calcium phosphate scaffolds from cuttlefish bone. J Am Ceram Soc. 2011;94(8):2362-70. https://doi.org/10.1111/j.1551-2916.2011.04404.x
Kloping LP, Purwati, Edward M. The healing effect of cuttlefish bone on fractured bone in rat model. Bali Med J. 2016;5(2):193-6. https://doi.org/10.15562/bmj.v5i2.195
Irianto KA, Pribadi A, Irsyam IA, Kloping YP, Sindrawati O. A comparison of osteoblast cell proliferation and osteocalcin expression in cuttlefish bone and bovine bone xenograft. Mol Cell Biomed Sci. 2019;3(2):75-80. https://doi.org/10.21705/mcbs.v3i2.58
Dogan E, Okumus Z. Cuttlebone used as a bone xenograft in bone healing. Vet Med. 2014;59(5):254-60. https://doi.org/10.17221/7519-VETMED
Cadman J, Zhou S, Chen Y, Li Q. Cuttlebone: characterisation, application and development of biomimetic materials. J Bionic Eng. 2012;9(3):367-76. https://doi.org/10.1016/S1672-6529(11)60132-7
Medical Device and Diagnostic Industry (MDDI) Online. Biocompatibility safety assessment of medical devices: FDA, ISO, and Japanese guidelines [Internet]. 2000 [cited 2018 Sep 18]. Available from: https://www.mddionline.com/biocompatibility-safety-assessment-medical-devices-fdaisoand-japanese-guidelines.
Mapara M, Thomas BS, Bhat KM. Rabbit as an animal model for experimental research. Dent Res J. 2012;9(1):111-8. https://doi.org/10.4103/1735-3327.92960
Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010;8(6):e1000412. https://doi.org/10.1371/journal.pbio.1000412
Rantam FA, P Ferdiansyah, A Purwati. Mesenchymal stem cell, hematopoietic, and application model. Surabaya: Universitas Airlangga; 2014. Indonesian.
Vajrabhaya LO, Korsuwannawong S, Surarit R. Cytotoxic and the proliferative effect of cuttlefish bone on MC3T3-E1 osteoblast cell line. Eur J Dent. 2017;11(4):503-7. https://doi.org/10.4103/ejd.ejd_159_17
BioVision Incorporated. Ready-to-use cell proliferation colorimetric reagent, WST-1 (catalog #K304-2500; assays; store at -20oC). Milpitas: BioVision Incorporated.
Supranto J. Simple random sampling. In: Supranto J. Sampling technique in survey and experimental study. Jakarta: Rineka Cipta; 2000. Indonesian.
Aminatun, Siswanto, Penga YM, Istifarah, Apsari R. The effect of sintering process on the characteristics of hydroxyapatite from cuttlefish bone (Sepia Sp.) Res J Pharm Biol Chem Sci. 2013;4(4):1431-42.
Dimarino AM, Caplan AI, Bonfield TL. Mesenchymal stem cells in tissue repair. Front Immunol. 2013;4:201. https://doi.org/10.3389/fimmu.2013.00201
Park JW, Bae SR, Suh JY, Lee DH, Kim SH, Kim H, et al. Evaluation of bone healing with eggshell-derived bone graft substitutes in rat calvaria: a pilot study. J Biomed Mater Res A. 2008;87(1):203-14. https://doi.org/10.1002/jbm.a.31768
Guda T, Walker JA, Pollot BE, Appleford MR, Oh S, Ong JL, et al. In vivo performance of bilayer hydroxyapatite scaffolds for bone tissue regeneration in the rabbit radius. J Mater Sci Mater Med. 2011;22(3):647-56. https://doi.org/10.1007/s10856-011-4241-7
Blokhuis TJ, Arts JJ. Bioactive and osteoinductive bone graft substitutes: definitions, facts and myths. Injury. 2011;42 Suppl 2:S26-9. https://doi.org/10.1016/j.injury.2011.06.010
Cerrolaza M, Duarte V, Garzón-Alvarado D. Analysis of bone remodeling under piezoelectricity effects using boundary elements. J Bionic Eng. 2017;14(4):659-71. https://doi.org/10.1016/S1672-6529(16)60432-8
Checa AG, Cartwright JH, Sánchez-Almazo I, Andrade JP, Ruiz-Raya F. The cuttlefish Sepia officinalis (Sepiidae, Cephalopoda) constructs cuttlebone from a liquid-crystal precursor. Sci Rep. 2015;5:11513. https://doi.org/10.1038/srep11513
Oryan A, Alidadi S, Moshiri A, Maffulli N. Bone regenerative medicine: classic options, novel strategies, and future directions. J Orthop Surg Res. 2014;9(1):18. https://doi.org/10.1186/1749-799X-9-18
Nazirkar G, Singh S, Dole V, Nikam A. Effortless effort in bone regeneration: a review. J Int Oral Health. 2014;6(3):120-4.
Copyright (c) 2020 komang agung irianto, suyenci limbong
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
Authors who publish with Medical Journal of Indonesia agree to the following terms:
- Authors retain copyright and grant Medical Journal of Indonesia right of first publication with the work simultaneously licensed under a Creative Commons Attribution-NonCommercial License that allows others to remix, adapt, build upon the work non-commercially with an acknowledgment of the work’s authorship and initial publication in Medical Journal of Indonesia.
- Authors are permitted to copy and redistribute the journal's published version of the work non-commercially (e.g., post it to an institutional repository or publish it in a book), with an acknowledgment of its initial publication in Medical Journal of Indonesia.