Skip to main content
Log in

Evaluation of the In Vivo Biological Effects of Marine Collagen and Hydroxyapatite Composite in a Tibial Bone Defect Model in Rats

  • Original Article
  • Published:
Marine Biotechnology Aims and scope Submit manuscript

Abstract

One of the most promising strategies to improve the biological performance of bone grafts is the combination of different biomaterials. In this context, the aim of this study was to evaluate the effects of the incorporation of marine spongin (SPG) into Hydroxyapatite (HA) for bone tissue engineering proposals. The hypothesis of the current study is that SPG into HA would improve the biocompatibility of material and would have a positive stimulus into bone formation. Thus, HA and HA/SPG materials were produced and scanning electron microscopy (SEM) analysis was performed to characterize the samples. Also, in order to evaluate the in vivo tissue response, samples were implanted into a tibial bone defect in rats. Histopathological, immunohistochemistry, and biomechanical analyses were performed after 2 and 6 weeks of implantation to investigate the effects of the material on bone repair. The histological analysis demonstrated that composite presented an accelerated material degradation and enhanced newly bone formation. Additionally, histomorphometry analysis showed higher values of %BV/TV and N.Ob/T.Ar for HA/SPG. Runx-2 immunolabeling was higher for the composite group and no difference was found for VEGF. Moreover, the biomechanical analysis demonstrated similar values for all groups. These results indicated the potential of SPG to be used as an additive to HA to improve the biological performance for bone regeneration applications. However, further long-term studies should be carried out to provide additional information regarding the material degradation and bone regeneration.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  • Alt V, Cheung WH, Chow SK, Thormann U, Cheung EN, Lips KS, Schnettler R, Leung KS (2016) Bone formation and degradation behavior of nanocrystalline hydroxyapatite with or without collagen-type 1 in osteoporotic bone defects - an experimental study in osteoporotic goats. Injury 47:58–65

    Google Scholar 

  • Bhatt RA, Rozental TD (2012) Bone graft substitutes. Hand Clin 28:457–468

    PubMed  Google Scholar 

  • Brandt J, Henning S, Michler G, Hein W, Bernstein A, Schulz M (2010) Nanocrystalline hydroxyapatite for bone repair: an animal study. J Mater Sci Mater Med 21:283–294

    PubMed  CAS  Google Scholar 

  • Campana V, Milano G, Pagano E, Barba M, Cicione C, Salonna G, Lattanzi W, Logroscino G (2014) Bone substitutes in orthopaedic surgery: from basic science to clinical practice. J Mater Sci Mater Med 25:2445–2461

    PubMed  PubMed Central  CAS  Google Scholar 

  • Cassino PC, Rosseti LS, Ayala OI, Martines MA, Portugual LC, Oliveira CG, Silva IS, de Araujo R (2018) Potencial of different hydroxyapatites as biomaterials in the bone remodeling. Acta Cir Bras 33:816–823

    PubMed  Google Scholar 

  • Denry I, Kuhn LT (2015) Design and characterization of calcium phosphate ceramic scaffolds for bone tissue engineering. Dent Mater 32:43–53

    PubMed  PubMed Central  Google Scholar 

  • Dimitriou R, Mataliotakis GI, Angoules AG, Kanakaris NK, Giannoudis PV (2011) Complications following autologous bone graft harvesting from the iliac crest and using the RIA: a systematic review. Injury 42:3–15

    Google Scholar 

  • Dorozhkin SV (2010) Bioceramics of calcium thophosphates. Biomaterials 31:1465–1485

    PubMed  CAS  Google Scholar 

  • Fernandes KR, Magri AMP, Kido HW, Ueno F, Assis L, Fernandes KPS, Mesquita-Ferrari RA, Martins VC, Plepis AM, Renno ACM (2017) Characterization and biological evaluation of the introduction of PLGA into biosilicate®. J Biomed Mater Res B Appl Biomater 05:1063–1074

    Google Scholar 

  • Fernandes KR, Parisi JR, Magri AMP, Kido HW, Gabbai-Armelin PR, Fortulan CA, Zanotto ED, Peitl O, Granito RN, Renno ACM (2019) Influence of the incorporation of marine spongin into a Biosilicate®: an in vitro study. J Mater Sci Mater Med 30:64

    PubMed  CAS  Google Scholar 

  • Gleeson JP, Plunkett NA, O’Brien FJ (2010) Addition of hydroxyapatite improves stiffness, interconnectivity and osteogenic potential of a highly porous collagen-based scaffold for bone tissue regeneration. Eur Cell Mater 20:218–230

    PubMed  CAS  Google Scholar 

  • Granito RN, Custódio MR, Rennó AC (2017) Natural marine sponges for bone tissue engineering: the state of art and future perspectives. J Biomed Mater Res B Appl Biomater 105:1717–1727

    PubMed  CAS  Google Scholar 

  • Green D, Howard D, Yang X, Kelly M, Oreffo RO (2003) Natural marine sponge fiber skeleton: a biomimetic scaffold for human osteoprogenitor cell attachment, growth, and differentiation. Tissue Eng 9:1159–1166

    PubMed  CAS  Google Scholar 

  • Guerado E, Caso E (2017) Challenges of bone tissue engineering in orthopaedic patients. World J Orthop 8:87–98

    PubMed  PubMed Central  Google Scholar 

  • Haach LCA, Purquerio BM, Silva NF Jr, Gaspar AM, Fortulan CA (2014) Comparison of two composites developed to be used as bone replacement–PMMA/Bioglass 45S5® Microfiber and PMMA/Hydroxyapatite. Bioceram Dev Appl 4:071

    Google Scholar 

  • Iwatsubo T, Kishi R, Miura T, Ohzono T, Yamaguchi T (2015) Formation of hydroxyapatite skeletal materials from hydrogel matrices via artificial biomineralization. J Phys Chem B 119:8793–8799

    PubMed  CAS  Google Scholar 

  • Junqua S, Robert L, Garrone R, Pavans de Ceccatty M, Vacelet J (1974) Biochemical and morphological studies on collagens of horny sponges Ircinia filaments compared to spongines. Connect Tissue Res 2:193–203

    PubMed  CAS  Google Scholar 

  • Komori T (2017) Roles of Runx2 in skeletal development. Adv Exp Med Biol 962:83–93

    PubMed  CAS  Google Scholar 

  • Lin K, Zhang D, Macedo MH, Cui W, Sarmento B, Shen G (2018) Advanced collagen-based biomaterials for regenerative biomedicine. Adv Funct Mater. https://doi.org/10.1002/adfm.201804943

  • Liu WC, Chen S, Zheng L, Qin L (2017) Angiogenesis assays for the evaluation of angiogenic properties of orthopaedic biomaterials - a general review. Adv Healthc Mater 6

  • Lopez-Heredia MA, Sa Y, Salmon P, de Wijn JR, Wolke JG, Jansen J (2012) A. Bulk properties and bioactivity assessment of porous polymethylmethacrylate cement loaded with calcium phosphates under simulated physiological conditions. Acta Biomater 8:3120–3127

    PubMed  CAS  Google Scholar 

  • Magri AM, Fernandes KR, Assis L, Mendes NA, da Silva Santos AL, de Oliveira DE, Rennó AC (2015) Photobiomodulation and bone healing in diabetic rats: evaluation of bone response using a tibial defect experimental model. Lasers Med Sci 30:1949–1957

    PubMed  Google Scholar 

  • Matassi F, Nistri L, Chicon Paez D, Innocenti M (2011) New biomaterials for bone regeneration. Clin Cases Miner Bone Metab 8:21–24

    PubMed  PubMed Central  Google Scholar 

  • Mishra R, Bishop T, Valerio IL, Fisher JP, Dean D (2016) The potential impact of bone tissue engineering in the clinic. Regen Med 11:571–587

    PubMed  PubMed Central  CAS  Google Scholar 

  • Oryan A, Alidadi S, Moshiri A, Maffulli N (2014) Bone regenerative medicine: classic options, novel strategies, and future directions. J Orthop Surg Res 9:18

    PubMed  PubMed Central  Google Scholar 

  • Pang KM, Lee JK, Seo YK, Kim SM, Kim MJ, Lee JH (2015) Biologic properties of nano-hydroxyapatite: an in vivo study of calvarial defects, ectopic bone formation and bone implantation. Biomed Mater Eng 25:25–38

    PubMed  CAS  Google Scholar 

  • Parfitt AM (1988) Bone histomorphometry: standardization of nomenclature, symbols and units. Summary of proposed system. Bone Miner 4:1–5

    PubMed  CAS  Google Scholar 

  • Parisi JR, Fernandes KR, Avanzi IR, Dorileo BP, Santana AF, Andrade AL, Gabbai-Armelin PR, Fortulan CA, Trichês ES, Granito RN, Renno ACM (2019) Incorporation of collagen from marine sponges (spongin) into hydroxyapatite samples: characterization and in vitro biological evaluation. Mar Biotechnol 21:30–37

    PubMed  CAS  Google Scholar 

  • Parizi AM, Oryan A, Shafiei-Sarvestani Z, Bigham-Sadegh A (2013) Effectiveness of synthetic hydroxyapatite versus Persian Gulf coral in an animal model of long bone defect reconstruction. J Orthop Traumatol 14:259–268

    PubMed  PubMed Central  Google Scholar 

  • Pek YS, Gao S, Arshad MS, Leck KJ, Ying JY (2008) Porous collagen-apatite nanocomposite foams as bone regeneration scaffolds. Biomaterials 29:4300–4305

    PubMed  CAS  Google Scholar 

  • Pozzolini M, Scarfì S, Gallus L, Castellano M, Vicini S, Cortese K, Gagliani MC, Bertolino M, Costa G, Giovine M (2018) Production, characterization and biocompatibility evaluation of collagen membranes derived from marine sponge Chondrosia reniformis Nardo, 1847. Mar Drugs 16:111

    PubMed Central  Google Scholar 

  • Raucci MG, Giugliano D, Alvarez-Perez MA, Ambrosio L (2015) Effects on growth and osteogenic differentiation of mesenchymal stem cells by the strontium-added sol–gel hydroxyapatite gel materials. J Mater Sci Mater Med 26:90

    PubMed  Google Scholar 

  • Sarkar SK, Lee BT (2015) Hard tissue regeneration using bone substitutes: an update on innovations in materials. Korean J Intern Med 30:279–293

    PubMed  PubMed Central  Google Scholar 

  • Scarano A, Lorusso F, Staiti G, Sinjari B, Tampieri A, Mortellaro C (2017) Sinus augmentation with biomimetic nanostructured matrix: tomographic, radiological, histological and histomorphometrical results after 6 months in humans. Front Physiol 8:565

    PubMed  PubMed Central  Google Scholar 

  • Siddiqui HA, Pickering KL, Mucalo MR (2018) A review on the use of hydroxyapatite-carbonaceous structure composites in bone replacement materials for strengthening purposes. Materials (Basel) 11:1813

    Google Scholar 

  • Silva TH, Moreira-Silva J, Marques AL, Domingues A, Bayon Y, Reis RL (2014) Marine origin collagens and its potential applications. Mar Drugs 12:5881–5901

    PubMed  PubMed Central  CAS  Google Scholar 

  • Smith JO, Aarvold A, Tayton ER, Dunlop DG, Oreffo RO (2011) Skeletal tissue regeneration: current approaches, challenges, and novel reconstructive strategies for an aging population. Tissue Eng B Rev 17:307–320

    Google Scholar 

  • Sousa THS, Fortulan CA, Antunes ES, Purquerio BM (2019) Concept of a bioactive implant with functional gradient structure. Key Eng Mater 396:221–224

    Google Scholar 

  • Swatschek D, Schatton W, Kellermann J, Müller WE, Kreuter J (2002) Marine sponge collagen: isolation, characterization and effects on the skin parameters surface-pH, moisture and sebum. Eur J Pharm Biopharm 53:107–113

    PubMed  CAS  Google Scholar 

  • Thorwarth M, Schultze-Mosgau S, Kessler P, Wiltfang J, Schlegel KA (2005) Bone regeneration in osseous defects using a resorbable nanoparticular hydroxyapatite. J Oral Maxillofac Surg 63:1626–1633

    PubMed  Google Scholar 

  • Walsh DP, Raftery RM, Chen G, Heise A, O’Brien FJ, Cryan SA (2019) Rapid healing of a critical-sized bone defect using a collagen-hydroxyapatite scaffold to facilitate low dose, combinatorial growth factor delivery. J Tissue Eng Regen Med 13:1843–1853

  • Wang W, Yeung K (2017) Bone grafts and biomaterials substitutes for bone defect repair: a review. Bioact Mater 2:224–247

    PubMed  PubMed Central  Google Scholar 

  • Wang H, Li Y, Zuo Y, Li J, Ma S, Cheng L (2007) Biocompatibility and osteogenesis of biomimetic nano-hydroxyapatite/polyamide composite scaffolds for bone tissue engineering. Biomaterials 28:3338–3348

    PubMed  CAS  Google Scholar 

  • Wang B, Dong J, Zhou X, Lee KJ, Huang R, Zhang S, Liu Y (2009) Nucleosides from the marine sponge Haliclona sp. Z Naturforsch 64:143–148

    CAS  Google Scholar 

  • Wei X, Egawa S, Matsumoto R, Yasuda H, Hirai K, Yoshii T, Okawa A, Nakajima T, Sotome S (2018) Augmentation of fracture healing by hydroxyapatite/collagen paste and bone morphogenetic protein-2 evaluated using a rat femur osteotomy model. J Orthop Res 36:129–137

    PubMed  CAS  Google Scholar 

  • Zhang Z, Li Z, Zhang C, Liu J, Bai Y, Li S, Zhang C (2018) Biomimetic intrafibrillar mineralized collagen promotes bone regeneration via activation of the Wnt signaling pathway. Int J Nanomedicine 13:7503–7516

    PubMed  PubMed Central  CAS  Google Scholar 

Download references

Acknowledgments

The authors would like to acknowledge CAPES Foundation, Ministry of Education of Brazil, Brasilia-DF, Brazil. Prof. Dr. Márcio Reis Custódio from Department of General Physiology of the Institute of Biosciences (IB-USP) for the assistance with this experiment.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Julia Risso Parisi.

Ethics declarations

Conflict of Interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Parisi, J.R., Fernandes, K.R., de Almeida Cruz, M. et al. Evaluation of the In Vivo Biological Effects of Marine Collagen and Hydroxyapatite Composite in a Tibial Bone Defect Model in Rats. Mar Biotechnol 22, 357–366 (2020). https://doi.org/10.1007/s10126-020-09955-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10126-020-09955-6

Keywords

Navigation