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      In Vitro Osteoinductivity Assay of Hydroxylapatite Scaffolds, Obtained with Biomorphic Transformation Processes, Assessed Using Human Adipose Stem Cell Cultures

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          Abstract

          In this study, the in vitro biocompatibility and osteoinductive ability of a recently developed biomorphic hydroxylapatite ceramic scaffold (B-HA) derived from transformation of wood structures were analyzed using human adipose stem cells (hASCs). Cell viability and metabolic activity were evaluated in hASCs, parental cells and in recombinant genetically engineered hASC-eGFP cells expressing the green fluorescence protein. B-HA osteoinductivity properties, such as differentially expressed genes (DEG) involved in the skeletal development pathway, osteocalcin (OCN) protein expression and mineral matrix deposition in hASCs, were evaluated. In vitro induction of osteoblastic genes, such as Alkaline phosphatase ( ALPL), Bone gamma-carboxyglutamate (gla) protein ( BGLAP), SMAD family member 3 ( SMAD3), Sp7 transcription factor ( SP7) and Transforming growth factor, beta 3 ( TGFB3) and Tumor necrosis factor (ligand) superfamily, member 11 ( TNFSF11)/Receptor activator of NF-κB (RANK) ligand ( RANKL), involved in osteoclast differentiation, was undertaken in cells grown on B-HA. Chondrogenic transcription factor SRY (sex determining region Y)-box 9 ( SOX9), tested up-regulated in hASCs grown on the B-HA scaffold. Gene expression enhancement in the skeletal development pathway was detected in hASCs using B-HA compared to sintered hydroxylapatite (S-HA). OCN protein expression and calcium deposition were increased in hASCs grown on B-HA in comparison with the control. This study demonstrates the biocompatibility of the novel biomorphic B-HA scaffold and its potential use in osteogenic differentiation for hASCs. Our data highlight the relevance of B-HA for bone regeneration purposes.

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          Calcium phosphate cements for bone substitution: chemistry, handling and mechanical properties.

          Since their initial formulation in the 1980s, calcium phosphate cements (CPCs) have been increasingly used as bone substitutes. This article provides an overview on the chemistry, kinetics of setting and handling properties (setting time, cohesion and injectability) of CPCs for bone substitution, with a focus on their mechanical properties. Many processing parameters, such as particle size, composition of cement reactants and additives, can be adjusted to control the setting process of CPCs, concomitantly influencing their handling and mechanical performance. Moreover, this review shows that, although the mechanical strength of CPCs is generally low, it is not a critical issue for their application for bone repair--an observation not often realized by researchers and clinicians. CPCs with compressive strengths comparable to those of cortical bones can be produced through densification and/or homogenization of the cement matrix. The real limitation for CPCs appears to be their low fracture toughness and poor mechanical reliability (Weibull modulus), which have so far been only rarely studied.
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            TGF-β Family Signaling in Mesenchymal Differentiation.

            Mesenchymal stem cells (MSCs) can differentiate into several lineages during development and also contribute to tissue homeostasis and regeneration, although the requirements for both may be distinct. MSC lineage commitment and progression in differentiation are regulated by members of the transforming growth factor-β (TGF-β) family. This review focuses on the roles of TGF-β family signaling in mesenchymal lineage commitment and differentiation into osteoblasts, chondrocytes, myoblasts, adipocytes, and tenocytes. We summarize the reported findings of cell culture studies, animal models, and interactions with other signaling pathways and highlight how aberrations in TGF-β family signaling can drive human disease by affecting mesenchymal differentiation.
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              Collagen-Hydroxyapatite Scaffolds Induce Human Adipose Derived Stem Cells Osteogenic Differentiation In Vitro

              Mesenchymal stem cells (MSCs) play a crucial role in regulating normal skeletal homeostasis and, in case of injury, in bone healing and reestablishment of skeletal integrity. Recent scientific literature is focused on the development of bone regeneration models where MSCs are combined with biomimetic three-dimensional scaffolds able to direct MSC osteogenesis. In this work the osteogenic potential of human MSCs isolated from adipose tissue (hADSCs) has been evaluated in vitro in combination with collagen/Mg doped hydroxyapatite scaffolds. Results demonstrate the high osteogenic potential of hADSCs when cultured in specific differentiation induction medium, as revealed by the Alizarin Red S staining and gene expression profile analysis. In combination with collagen/hydroxyapatite scaffold, hADSCs differentiate into mature osteoblasts even in the absence of specific inducing factors; nevertheless, the supplement of the factors markedly accelerates the osteogenic process, as confirmed by the expression of specific markers of pre-osteoblast and mature osteoblast stages, such as osterix, osteopontin (also known as bone sialoprotein I), osteocalcin and specific markers of extracellular matrix maturation and mineralization stages, such as ALPL and osteonectin. Hence, the present work demonstrates that the scaffold per se is able to induce hADSCs differentiation, while the addition of osteo-inductive factors produces a significant acceleration of the osteogenic process. This observation makes the use of our model potentially interesting in the field of regenerative medicine for the treatment of bone defects.
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                Author and article information

                Contributors
                Role: Academic Editor
                Role: Academic Editor
                Role: Academic Editor
                Role: Academic Editor
                Journal
                Int J Mol Sci
                Int J Mol Sci
                ijms
                International Journal of Molecular Sciences
                MDPI
                1422-0067
                30 June 2021
                July 2021
                : 22
                : 13
                : 7092
                Affiliations
                [1 ]Department of Medical Sciences, Section of Experimental Medicine, School of Medicine, University of Ferrara, 64b Fossato di Mortara Street, 44121 Ferrara, Italy; mariarosa.iaquinta@ 123456unife.it (M.R.I.); elena.torreggiani@ 123456unife.it (E.T.); chiara.mazziotta@ 123456unife.it (C.M.); elisa.mazzoni@ 123456unife.it (E.M.)
                [2 ]Institute of Science and Technology for Ceramics, National Research Council, 48018 Faenza, Italy; andrea.ruffini@ 123456istec.cnr.it (A.R.); simone.sprio@ 123456istec.cnr.it (S.S.); anna.tampieri@ 123456istec.cnr.it (A.T.)
                [3 ]Laboratory for Technologies of Advanced Therapies (LTTA), University of Ferrara, 44121 Ferrara, Italy
                Author notes
                [* ]Correspondence: tgm@ 123456unife.it (M.T.); mrf@ 123456unife.it (F.M.)
                Author information
                https://orcid.org/0000-0003-3242-859X
                https://orcid.org/0000-0001-7874-2002
                https://orcid.org/0000-0002-7170-9821
                https://orcid.org/0000-0003-1526-7915
                Article
                ijms-22-07092
                10.3390/ijms22137092
                8267654
                34209351
                cda6fe4c-d639-4fa5-894b-25a2a171461a
                © 2021 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( https://creativecommons.org/licenses/by/4.0/).

                History
                : 17 March 2021
                : 26 June 2021
                Categories
                Article

                Molecular biology
                biomorphic scaffolds,hydroxylapatite,bone regeneration,nanostructure,in vitro osteoinductivity

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