نوع مقاله : مقاله پژوهشی

نویسندگان

واحد علوم و تحفیفات دانشگاه آزاد اسلامی

چکیده

حفظ ساختار بیولوژیکی اولیه استخوان اسفنجی می تواند آن را برای استفاده به عنوان یک داربست مناسب برای مهندسی موفقیت آمیز بافت استخوان آماده کند. علاوه بر این ، از بین بردن سلولهای متعلق به بستر آن برای افزایش زیست سازگاری آن و کاهش پاسخهای ایمونولوژیکی آنها بسیار حیاتی است. در این مطالعه ، از روشهای شیمیایی برای سلول زدایی داربستهای سه بعدی ساخته شده از استخوان لگن گوساله اسفنجی استفاده شد. برای این منظور ، نمونه های استخوانی که از استخوان لگن گوساله بریده شده بودند با استفاده از روش شیمیایی (سدیم دودسیل سولفات (SDS) و TritonX-100 با غلظت های مختلف) سلول زدایی شدند. نمونه ها با رنگ آمیزی هماتوکسیلین و ائوزین ، رنگ آمیزی تریکروم، میکروسکوپ الکترونی نوری و روبشی مشخصه یابی شدند. در پایان، برای اطمینان از عدم وجود مواد سمی در داربست ، آزمایش سمیت سلولی انجام شد. نتایج نشان داد که نمونه های سلولز شده با TritonX-100 2٪ و به ترتیب در ترکیب محلول 3٪ TritonX-100 و 4٪ SDS می توانند جایگزین بافت استخوانی اسفنجی آسیب دیده شوند.

کلیدواژه‌ها

عنوان مقاله [English]

Comparison of SDS and TritonX-100 effects on cell removing of bovine spongy bone for using in bone replacements

نویسندگان [English]

  • Nahid Hassanzadeh Nemati
  • setareh nikzamir
  • zohreh ansarinezhad

چکیده [English]

Background: Preserving the biological structure of the initial nature of cancellous bone could prepare it for a proper scaffold for successful bone tissue engineering. Moreover, it is vital to eliminate the cells belonging to its bed to increase its biocompatibility and reduce their immunological responses.
Methods: In this study, Chemical methods were used for decellularization of three-dimensional scaffolds made from spongy calfchr('39')s pelvic bone. For this purpose, the bone samples which were cut from calf pelvis bone were degreased, and then their cells were removed through chemical (sodium dodecyl sulfate (SDS) and TritonX-100 with different concentrations) method. The samples were characterized by hematoxylin and eosin staining, trichrome staining, and optical and scanning electron microscope. In the end, to ensure the absence of toxic substances in the scaffold, a cell toxicity test was conducted.
Results: The results show that the decellularized samples with TritonX-100 of 2% and combining solution of 3% TritonX-100 and 4% SDS respectively (T3S4) can substitute for damaged cancellous bone tissue. The results indicated that calf pelvic spongy bone tissue, as a xenograft that has undergone decellularization with SDS and Triton x-100 chemical solutions, can produce an appropriate scaffold for bone tissue engineering. The natural bone tissue with preservation of collagen fibers and the presence of porosity in its structure can provide a suitable environment for tissue regeneration..
Conclusion: The results suggested that T3S4-acellular bone tissue can be further evaluated as a natural scaffold suitable for using in bone tissue engineering and restorative medicine.

کلیدواژه‌ها [English]

  • Bone tissue engineering
  • Xenograft Scaffold
  • Chemical Decellularization
  • Bone Replacement
  1. Ashley B. Which scaffold for which application?. Current Orthopaedics. 2007;21(4):280-7.
    doi: 10.1016/j.cuor. 2007.06.005.
  2. Yuan J, Zhang WJ, Liu G, Wei M, Qi ZL, Liu W, Cui L, Cao YL. Repair of canine mandibular bone defects with bone marrow stromal cells and coral. Tissue Eng Part A. 2010 Apr;16(4):1385-94. doi: 10.1089/ten.TEA.2009.0472.
  3. Jwo-Lin W, Ying-Tai Z, Ching-Cherng T, Chin-I Lin, Shi-Wei L, Guan-Liang C, The assay of Bone Reaction after implantation of Calcium Sulfat and A Composite of Calcium Sulfat and Calcium Phosphate. Journal of Medical and biological Engineering 2003;23(4):205-212.
  4. Fernandez de Grado G, Keller L, Idoux-Gillet Y, Wagner Q, Musset AM, Benkirane-Jessel N, Bornert F, Offner D. Bone substitutes: a review of their characteristics, clinical use, and perspectives for large bone defects management. J Tissue Eng. 2018 Jun 4;9:2041731418776819. doi: 10.1177/2041731418776819.
  5. Fröhlich M, Grayson WL, Wan LQ, Marolt D, Drobnic M, Vunjak-Novakovic G. Tissue engineered bone grafts: biological requirements, tissue culture and clinical relevance. Curr Stem Cell Res Ther. 2008 Dec;3(4):254-64. doi: 10.2174/157488808786733962.
  6. Weitao Y, Kangmei K, Xinjia W, Weili Q. Bone regeneration using an injectable calcium phosphate/autologous iliac crest bone composites for segmental ulnar defects in rabbits. J Mater Sci Mater Med. 2008 Jun;19(6):2485-92. doi: 10.1007/s10856-008-3383-8.
  7. Goldstein SA. Tissue engineering: functional assessment and clinical outcome. Ann N Y Acad Sci. 2002 Jun;961:183-92. doi: 10.1111/j.1749-6632.2002.tb03079.x.
  8. Vacanti CA, Bonassar L J. An overview of tissue engineered bone. Cline.Orthop. 1999;17(4)343-352.
    doi: 10.1097/00003086-199910001-00036.
  9. Jerome CP, Peterson PE. Nonhuman primate models in skeletal research. Bone. 2001 Jul;29(1):1-6.
    doi: 10.1016/s8756-3282(01)00477-x.
  10. Andrew W, Michael M, Geoffrey H, Warrick Be, Hilary B. The evaluation of processed cancellous bovine bone as a bone graft substitute. Clin.OraImpl. 2005;16:379-386. doi: 10.1111/j.1600-0501.2005.01113.x.
  11. Cukierman E, Pankov R, Yamada KM. Cell interactions with three-dimensional matrices. Curr Opin Cell Biol. 2002 Oct;14(5):633-9. doi: 10.1016/s0955-0674(02)00364-2.
  12. Badylak SF. Xenogeneic extracellular matrix as a scaffold for tissue reconstruction. Transpl Immunol. 2004; 12(3-4): 367-77. doi: 10.1016/j.trim.2003.12.016.
  13. Hussey, G.S., Dziki, J.L., Badylak S.F. Extracellular matrix-based materials for regenerative medicine. Nat. Rev. Mater. 2018; 3: 159–173. doi: 10.1038/s41578-018-0023-x.
  14. Mendibil u. et al. Tissue-Specific Decellularization Methods: Rationale and Strategies to Achieve Regenerative Compounds. International Journal of Molecular Sciences. 30 July 2020;21(15):5447. doi:10.3390/ijms21155447.
  15. Wu X et al. Mineralization of Biomaterials for Bone Tissue Engineering. International Journal of bioengineering. 2020;7(4):132. doi: 10.3390/bioengineering7040132.
  16. Wang W., Yeung Bone grafts and biomaterials substitutes for bone defect repair. Bioactive Materials. 2017;2:224-247. doi: 10.1016/j.bioactmat.2017.05.007.
  17. Keane TJ, Swinehart IT, Badylak SF. Methods of tissue decellularization used for preparation of biologic scaffolds and in vivo relevance. Methods. 2015 Aug;84:25-34. doi: 10.1016/j.ymeth.2015.03.005.
  18. Crapo P.M, Gilbert T.W, Badylak S.F. An overview of tissue and whole organ decellularization processes. Biomaterials. 2011;32:3233–3243.
    doi: 10.1016/j.biomaterials.2011.01.057.
  19. White LJ, Taylor AJ, Faulk DM, Keane TJ, Saldin LT, Reing JE, Swinehart IT, Turner NJ, Ratner BD, Badylak SF. The impact of detergents on the tissue decellularization process: A ToF-SIMS study. Acta Biomater. 2017 Mar 1;50:207-219. doi: 10.1016/j.actbio.2016.12.033.
  20. Woods T, Gratzer PF. Effectiveness of three extraction techniques in the development of a decellularized bone-anterior cruciate ligament-bone graft. Biomaterials. 2005 Dec;26(35):7339-49.
    doi: 10.1016/j.biomaterials.2005.05.066.
  21. Cartmell JS, Dunn MG. Effect of chemical treatments on tendon cellularity and mechanical properties. J Biomed Mater Res. 2000 Jan;49(1):134-40. doi: 10.1002/(sici)1097-4636(200001)49:1<134::aid-jbm17>3.0.co;2-d.
  22. Elder BD, Kim DH, Athanasiou KA. Developing an articular cartilage decellularization process toward facet joint cartilage replacement. Neurosurgery. 2010 Apr;66(4):722-7; discussion 727. doi: 10.1227/01.NEU.0000367616.49291.9F.
  23. Chen RN, Ho HO, Tsai YT, Sheu MT. Process development of an acellular dermal matrix (ADM) for biomedical applications. Biomaterials. 2004 Jun;25(13):2679-86. doi: 10.1016/j.biomaterials.2003.09.070.
  24. Tavassoli A, Matin MM, Niaki MA, Mahdavi-Shahri N, Shahabipour F. Mesenchymal stem cells can survive on the extracellular matrix-derived decellularized bovine articular cartilage scaffold. Iran J Basic Med Sci. 2015 Dec;18(12):1221-7.
  25. Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials. 2006 Jul;27(19):3675-83. doi: 10.1016/j.biomaterials.2006.02.014.

 

 

  1. Butcher JT, Mahler GJ, Hockaday LA. Aortic valve disease and treatment: the need for naturally engineered solutions. Adv Drug Deliv Rev. 2011 Apr 30;63(4-5):242-68. doi: 10.1016/j.addr.2011.01.008.
  2. Naso F, Gandaglia A, Formato M, Cigliano A, Lepedda AJ, Gerosa G, Spina M. Differential distribution of structural components and hydration in aortic and pulmonary heart valve conduits: Impact of detergent-based cell removal. Acta Biomater. 2010 Dec;6(12):4675-88.
    doi: 10.1016/j.actbio.2010.06.037.
  3. Seddon AM, Curnow P, Booth PJ. Membrane proteins, lipids and detergents: not just a soap opera. Biochim Biophys Acta. 2004 Nov 3;1666(1-2):105-17.
    doi: 10.1016/j.bbamem.2004.04.011.
  4. Schaner PJ, Martin ND, Tulenko TN, Shapiro IM, Tarola NA, Leichter RF, Carabasi RA, Dimuzio PJ. Decellularized vein as a potential scaffold for vascular tissue engineering. J Vasc Surg. 2004 Jul;40(1):146-53. doi: 10.1016/j.jvs.2004.03.033
  5. Lumpkins SB, Pierre N, McFetridge PS. A mechanical evaluation of three decellularization methods in the design of a xenogeneic scaffold for tissue engineering the temporomandibular joint disc. Acta Biomater. 2008 Jul;4(4):808-16. doi: 10.1016/j.actbio.2008.01.016. -16.
  6. Staubli AE, De Simoni C, Babst R, Lobenhoffer P. TomoFix: a new LCP-concept for open wedge osteotomy of the medial proximal tibia--early results in 92 cases. Injury. 2003 Nov;34 Suppl 2:B55-62. doi: 10.1016/j.injury.2003.09.025.
  7. Meijer GJ, de Bruijn JD, Koole R, van Blitterswijk CA. Cell-based bone tissue engineering. PLoS Med. 2007 Feb;4(2):e9. doi: 10.1371/journal.pmed.0040009.
  8. Hutmacher DW. Scaffold design and fabrication technologies for engineering tissues--state of the art and future perspectives. J Biomater Sci Polym Ed. 2001;12(1):107-24. doi: 10.1163/156856201744489
  9. Greenwald AS, Boden SD, Goldberg VM, Khan Y, Laurencin CT, Rosier RN; 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.
  10. Kim SH, Shin JW, Park SA, Kim YK, Park MS, Mok JM, Yang WI, Lee JW. Chemical, structural properties, and osteoconductive effectiveness of bone block derived from porcine cancellous bone. J Biomed Mater Res B Appl Biomater. 2004 Jan 15;68(1):69-74.
    doi: 10.1002/jbm.b.10084.
  11. McFetridge PS, Daniel JW, Bodamyali T, Horrocks M, Chaudhuri JB. Preparation of porcine carotid arteries for vascular tissue engineering applications. J Biomed Mater Res A. 2004 Aug 1;70(2):224-34. doi: 10.1002/jbm.a.30060.
  12. Ketchedjian A, Jones AL, Krueger P, Robinson E, Crouch K, Wolfinbarger L Jr, Hopkins R. Recellularization of decellularized allograft scaffolds in ovine great vessel reconstructions. Ann Thorac Surg. 2005 Mar;79(3):888-96; discussion 896. doi: 10.1016/j.athoracsur.2004.09.033.
  13. David V, Guignandon A, Martin A, Malaval L, Lafage-Proust MH, Rattner A, Mann V, Noble B, Jones DB, Vico L. Ex Vivo bone formation in bovine trabecular bone cultured in a dynamic 3D bioreactor is enhanced by compressive mechanical strain. Tissue Eng Part A. 2008 Jan;14(1):117-26. doi: 10.1089/ten.a.2007.0051.
  14. Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials. 2006 Jul;27(19):3675-83. doi: 10.1016/j.biomaterials.2006.02.014.
  15. Jank B J, Xiong L, Moser P T et al. Engineered composite tissue as a bioartificial limb graft. Biomaterials. 2015; 61: 246– 256. doi:10.1016/j.biomaterials.2015.04.051
  16. Pang K, Du L, Wu X. A rabbit anterior cornea replacement derived from acellular porcine cornea matrix, epithelial cells and keratocytes. Biomaterials. 2010 Oct;31(28):7257-65. doi: 10.1016/j.biomaterials.2010.05.066.
  17. Wang B, Borazjani A, Tahai M, Curry AL, Simionescu DT, Guan J, To F, Elder SH, Liao J. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrow mononuclear cells. J Biomed Mater Res A. 2010 Sep 15;94(4):1100-10. doi: 10.1002/jbm.a.32781.
  18. Zhou J, Fritze O, Schleicher M, Wendel HP, Schenke-Layland K, Harasztosi C, Hu S, Stock UA. Impact of heart valve decellularization on 3-D ultrastructure, immunogenicity and thrombogenicity. Biomaterials. 2010 Mar;31(9):2549-54. doi: 10.1016/j.biomaterials.2009.11.088.
  19. Syed O, Walters NJ, Day RM, Kim HW, Knowles JC. Evaluation of decellularization protocols for production of tubular small intestine submucosa scaffolds for use in oesophageal tissue engineering. Acta Biomater. 2014 Dec;10(12):5043-5054. doi: 10.1016/j.actbio.2014.08.024.
  20. Sullivan DC, Mirmalek-Sani SH, Deegan DB, Baptista PM, Aboushwareb T, Atala A, Yoo JJ. Decellularization methods of porcine kidneys for whole organ engineering using a high-throughput system. Biomaterials. 2012 Nov;33(31):7756-64. doi: 10.1016/j.biomaterials.2012.07.023.
  21. Schaner PJ, Martin ND, Tulenko TN, Shapiro IM, Tarola NA, Leichter RF, Carabasi RA, Dimuzio PJ. Decellularized vein as a potential scaffold for vascular tissue engineering. J Vasc Surg. 2004 Jul;40(1):146-53. doi: 10.1016/j.jvs.2004.03.033. 2004.
  22. Gilpin SE, Guyette JP, Gonzalez G, Ren X, Asara JM, Mathisen DJ, Vacanti JP, Ott HC. Perfusion decellularization of human and porcine lungs: bringing the matrix to clinical scale. J Heart Lung Transplant. 2014 Mar;33(3):298-308. doi: 10.1016/j.healun.2013.10.030.
  23. O'Neill JD, Anfang R, Anandappa A, Costa J, Javidfar J, Wobma HM, Singh G, Freytes DO, Bacchetta MD, Sonett JR, Vunjak-Novakovic G. Decellularization of human and porcine lung tissues for pulmonary tissue engineering. Ann Thorac Surg. 2013 Sep;96(3):1046-55; discussion 1055-6. doi: 10.1016/j.athoracsur.2013.04.022.
  24. Guyette JP, Charest JM, Mills RW, Jank BJ, Moser PT, Gilpin SE, Gershlak JR, Okamoto T, Gonzalez G, Milan DJ, Gaudette GR, Ott HC. Bioengineering Human Myocardium on Native Extracellular Matrix. Circ Res. 2016 Jan 8;118(1):56-72. doi: 10.1161/CIRCRESAHA.115.306874.
  25. Uygun B, Soto-Gutierrez A, Yagi H. et al. Organ reengineering through development of a transplantable recellularized liver graft using decellularized liver matrix. Nat Med. 2010;16: 814–820. doi; 10.1038/nm.2170.
  26. Ott H, Matthiesen, T, Goh SK. et al. Perfusion-decellularized matrix: using nature's platform to engineer a bioartificial heart. Nat Med.2008;14: 213–221. doi:10.1038/nm16.
  27. Gilpin A, Yang Y. Decellularization Strategies for Regenerative Medicine: From Processing Techniques to Applications. Biomed Res Int. 2017;2017:9831534. doi: 10.1155/2017/9831534.