Pro osteon/collagen hydroxylapatite hybrid scaffold is able to induce human mesechymal stem cells (hMSCs) to osteogenic differentiation

Abstract

Event Abstract Back to Event Pro osteon/collagen hydroxylapatite hybrid scaffold is able to induce human mesechymal stem cells (hMSCs) to osteogenic differentiation Elisa Mazzoni1*, Antonio D'Agostino2, Lorenzo Trevisiol2, Maria Rosa Iaquinta1, Chiara Mazziotta1 and Mauro G. Tognon1 1 University of Ferrara, Department of Morphology, Surgery and Experimental Medicine, Italy 2 University of Verona, Department of Surgery, Dentistry, Paediatrics and Gynecology, Italy Aim. Innovative scaffolds are of paramount importance for bone grafting and re-growth. Distinct enhancements of scaffold properties may optimize the product performance for different applications in the fields of maxillofacial and dental sciences/clinics. The most common scaffold improvements were obtained for their biocompatibility, mechanical properties in osteo-conductive and osteo-inductive properties and healing rate. In this study an innovative hydroxylapatite hybrid scaffold composed of granular hydroxylapatite (Pro Osteon®200, Interpore Cross Irvine, CA, USA) and collagen Hemostat (Bard Warwick, Rhode Island, USA) (Coll/HA) was investigated using human bone marrow-derived mesenchymal stem cells (hMSCs) of adult donors. Materials and Methods. In order to evaluate (i) biocompatibility, (ii) osteoconductivity and (iii) osteoinductivity hMSC cultures were grown on the innovative scaffold. The cellular morphology, cytoskeleton organization, and cell viability were studied by immunohistochemistry (IHC), and AlamarBlue metabolic assay respectively. Osteocalcin and osteopontin expression proteins were detected by IHC. The temporal osteocalcin expression protein in hMSCs grown on the biomaterial and in osteogenic condition (OC), and the control (TCPS), were quantified by Human Osteocalcin Instant E.L.I.S.A assays. Expression of osteogenic genes were evaluated by quantitative PCR (Q-PCR) array technologies; the Human Osteogenesis RT² Profiler PCR Array (Qiagen) was used to analyze the expression of 84 genes related to osteogenic differentiation at day 21. Results. Cell morphology of hMSC–eGFP cells was indistinguishable from that of parental hMSC. Indeed, hMSC-eGFP grown on the scaffold showed a normal morphology. Metabolic activity was increased during the 21 days of experiments (P<0.05). The cytoskeleton architecture seemed to be well organized, whereas its integrity remains uninfluenced by the scaffold during the time course. The biomaterial induced the matrix mineralization in hMSCs at day 14. Osteogenic proteins, such as osteocalcin and osteopontin were detected at day 21.The biomaterial induced the up-regulation of osteocalcin protein expression levels, quantified in E.L.I.S.A assay at day 21, compared to control (TCPS) and at day 14. Gene expression analyzed in hMSCs allowed us to detect the upregulation of mRNAs of 16 genes, belonging to the osteogenic differentiation pathway. Specific genes were for (i) the ossification process: BMP2/3, COL2A1, MMP9, NOG, SPP1, TNFSF11, TGFB3, (ii) osteoblast differentiation were: GLI1, SMAD3, SP7, (iii) whereas for the extracellular matrix (ECM) and cell adhesion molecules were: MMP10, ICAM1, ITGAM, CD36. In addition, the growth factor CSF3 was also up-regulated compared to the control, ad day 21. The transcription factor SP7 was the highest gene modulated by the biomaterial with a 3 Log2 fold increase. It has been reported that during the development of the skeletal bone and tooth, SP7 is a key mesenchymal factor necessary for cell fate decisions in the differentiation of specialized cells. Down-regulated genes were that encoding ECM and cell-to cell adhesion molecules such as BGN, CDH11, COL1A1, COL5A1, COMP, CSF2, CTSK, IGF1/2, IGF1R, ALPL. Early transcription factors, such as RUNX2, SMAD1, TWIST1 were down-regulated, at day 21. In addition, FGFR2 and BMPR2 genes were also down-regulated compared to the control. Discussion. Our data demonstrate that the innovative scaffold provides a good microenvironment for the hMSCs adhesion and proliferation. The scaffold demonstrated biocompatibility in term of morphology, viability and cytoskeleton architecture of hMSC grown on the biomaterial. Gene expression profile analyses by array technology demonstrated that, in hMSCs, the scaffold induces up-regulation in specific genes that are involved in ossification process, such as BMP2/3, SPP1 and SP7, at d 21 post-cell seeding. The scaffold induces a up-regulation of the osteocalcin protein with improvement in matrix mineralization, indicating a good osteoinductivity performance. In conclusion, our experimental cell biology and epigenetic analyses suggest that the Coll/HA hybrid scaffold is an excellent biomaterial for the bone repair and bone tissue engineering. References 1. Mazzoni E. et al. FASEB J. 31:4555-65, 2017. 2. D'Agostino A. et al. J. Oral Maxillofac. Surg 74 1238.e1-e15, 2016. 3. Manfrini M. et al. J. Cell. Physiol. 228:1229-37, 2013 Keywords: hMSC = human mesenchymal stem cell, Pro Osteon, Collagen, Osteogenesis, Biocompatibility Conference: 5th National and 1st International Symposium of Italian Society of Oral Pathology and Medicine., Ancona, Italy, 19 Oct - 20 Oct, 2018. Presentation Type: Poster Presentation Topic: Oral Diseases Citation: Mazzoni E, D'Agostino A, Trevisiol L, Iaquinta M, Mazziotta C and Tognon MG (2019). Pro osteon/collagen hydroxylapatite hybrid scaffold is able to induce human mesechymal stem cells (hMSCs) to osteogenic differentiation. Front. Physiol. Conference Abstract: 5th National and 1st International Symposium of Italian Society of Oral Pathology and Medicine.. doi: 10.3389/conf.fphys.2019.27.00018 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 11 Oct 2018; Published Online: 09 Dec 2019. * Correspondence: Dr. Elisa Mazzoni, University of Ferrara, Department of Morphology, Surgery and Experimental Medicine, Ferrara, Emilia-Romagna, 44121, Italy, [email protected] Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Elisa Mazzoni Antonio D'Agostino Lorenzo Trevisiol Maria Rosa Iaquinta Chiara Mazziotta Mauro G Tognon Google Elisa Mazzoni Antonio D'Agostino Lorenzo Trevisiol Maria Rosa Iaquinta Chiara Mazziotta Mauro G Tognon Google Scholar Elisa Mazzoni Antonio D'Agostino Lorenzo Trevisiol Maria Rosa Iaquinta Chiara Mazziotta Mauro G Tognon PubMed Elisa Mazzoni Antonio D'Agostino Lorenzo Trevisiol Maria Rosa Iaquinta Chiara Mazziotta Mauro G Tognon Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.