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Otoliths-composed gelatin/sodium alginate scaffolds for bone regeneration

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Abstract

Evidence that otoliths, mineral-rich limestone concrescences present in the inner ear of bone fishes, can accelerate bone formation in vivo has been previously reported. The goal of this work was the development, characterization, and evaluation of the cytocompatibility of otoliths-incorporated sodium alginate and gelatin scaffolds. Cynoscion acoupa–derived otoliths were characterized by X-ray fluorescence spectrometry (FRX), particle size, free lime, and weight loss by calcination. Furthermore, otoliths were incorporated into sodium alginate (ALG/OTL-s) or gelatin (GEL/OTL-s) scaffolds, previously developed by freeze-drying. Then, the scaffolds were characterized by thermogravimetric analysis (TGA/DTG), differential scanning calorimetry (DSC), infrared spectroscopy with Fourier transform (FTIR), swelling tests, and scanning electron microscopy (SEM). Cytotoxicity assays were run against J774.G8 macrophages and MC3T3-E1 osteoblasts. Data obtained from TGA/DTG, DSC, and FTIR analyses confirmed the interaction between otoliths and the polymeric scaffolds. SEM showed the homogeneous porous 3D structure rich in otolith micro-fragments in both scaffolds. Swelling of the GEL/OTL-s (63.54 ± 3.0%) was greater than of ALG/OTL-s (13.36 ± 9.9%) (p < 0.001). The viability of J774.G8 macrophages treated with both scaffolds was statistically similar to the group treated with DMEM only (p > 0.05) and significantly higher than that treated with Triton-X (p < 0.01) at 72 h. Both scaffolds showed approximately 100% growth of MC3T3-E1 osteoblasts by 24 h, similarly to control (p > 0.05). However, by 48 h, only ALG/OTL-s showed growth similar to control (p > 0.05), whereas GEL/OTL showed a significantly lower growth index (p < 0.05). In conclusion, the physicochemical profiles suggest proper interaction between the otoliths and the two developed polymeric 3D scaffolds. Moreover, both materials showed cytocompatibility with J774.G8 macrophages but the growth of MC3T3-E1 osteoblasts was higher when exposed to ALG/OTL-s. These data suggest that sodium alginate/otoliths scaffolds are potential biomaterials to be used in bone regeneration applications.

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Author contribution statement

Conceptualization: Daisy Pereira Valido. Methodology: Wilson Déda Gonçalves Júnior, Maria Eliane de Andrade, Allan Andrade Rezende, Felipe Mendes de Andrade de Carvalho, Renata de Lima, Gabriela das Graças Gomes Trindade, Caio de Alcântara Campos, Ana Maria Santos Oliveira, Eloísa Portugal Barros Silva Soares de Souza, Luiza Abrahão Frank. Formal analysis and investigation: Silvia Stanisçuaski Guterres. Writing - original draft preparation: Eliana Midori Sussuchi, Charlene Regina Santos Matos, André Polloni, Adriano Antunes de Souza Araújo, Patrícia Severino. Writing - review and editing: Eliana Barbosa Souto. Funding acquisition: Francine Ferreira Padilha, Supervision: Ricardo Luiz Cavalcanti de Albuquerque Júnior.

Funding

We would like to thank the National Council for Scientific and Technological Development (CNPq) and the Foundation for Research and Technological Innovation Support of the State of Sergipe for the financial support in this study. EMBS acknowledges the sponsorship of the projects M-ERA-NET-0004/2015-PAIRED and UIDB/04469/2020 (strategic fund), received support from the Portuguese Science and Technology Foundation, Ministry of Science and Education (FCT/MEC) through national funds, and was co-financed by FEDER, under the Partnership Agreement PT2020.

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Authors and Affiliations

  1. Tiradentes University, Av. Murilo Dantas, 300, Aracaju, 49010-390, Brazil

    Daisy Pereira Valido, Wilson Déda Gonçalves Júnior, Maria Eliane de Andrade, Allan Andrade Rezende, Felipe Mendes de Andrade de Carvalho, André Polloni, Francine Ferreira Padilha, Patrícia Severino & Ricardo Luiz Cavalcanti de Albuquerque Júnior

  2. Laboratory of Nanomedicine and Nanotecnology, Instituto de Tecnologia e Pesquisa, Av. Murilo Dantas, 300 - Farolândia, Aracaju, SE, 49032-490, Brazil

    Daisy Pereira Valido, Wilson Déda Gonçalves Júnior, Maria Eliane de Andrade, Allan Andrade Rezende, Felipe Mendes de Andrade de Carvalho, André Polloni, Francine Ferreira Padilha, Patrícia Severino & Ricardo Luiz Cavalcanti de Albuquerque Júnior

  3. Department of Biotechnology, University of Sorocaba, Rodovia Raposo Tavares S/N-km 92,5, Sorocaba, SP, CEP 18023-000, Brazil

    Renata de Lima

  4. Department of Physiology, Federal University of Sergipe, São Cristóvão, Sergipe, 49100-00, Brazil

    Gabriela das Graças Gomes Trindade, Caio de Alcântara Campos, Ana Maria Santos Oliveira, Eloísa Portugal Barros Silva Soares de Souza, Eliana Midori Sussuchi, Charlene Regina Santos Matos & Adriano Antunes de Souza Araújo

  5. Faculty of Pharmacy, Federal University of Rio Grande do Sul, Av. Ipiranga, 2759, Porto Alegre, Rio Grande do Sul, 90610-000, Brazil

    Luiza Abrahão Frank & Silvia Stanisçuaski Guterres

  6. Tiradentes Institute, 150 Mt Vernon St, Dorchester, MA, 02125, USA

    Patrícia Severino

  7. Center for Biomedical Engineering, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, 65 Landsdowne Street, Cambridge, MA, 02139, USA

    Patrícia Severino

  8. Faculty of Pharmacy, University of Coimbra, Pólo das Ciências da Saúde, Azinhaga de Santa Comba, 3000-548, Coimbra, Portugal

    Eliana Barbosa Souto

  9. CEB - Centre of Biological Engineering, University of Minho, Campus de Gualtar, 4710-057, Braga, Portugal

    Eliana Barbosa Souto

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  1. Daisy Pereira Valido
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  2. Wilson Déda Gonçalves Júnior
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  11. Luiza Abrahão Frank
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  15. André Polloni
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  16. Adriano Antunes de Souza Araújo
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  17. Francine Ferreira Padilha
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  19. Eliana Barbosa Souto
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  20. Ricardo Luiz Cavalcanti de Albuquerque Júnior
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Correspondence to Patrícia Severino or Ricardo Luiz Cavalcanti de Albuquerque Júnior.

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Valido, D.P., Júnior, W.D.G., de Andrade, M.E. et al. Otoliths-composed gelatin/sodium alginate scaffolds for bone regeneration. Drug Deliv. and Transl. Res. 10, 1716–1728 (2020). https://doi.org/10.1007/s13346-020-00845-x

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