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This can be additionally supported by employing numerical simulation techniques to assess local physical factors, such as oxygen concentration, and mechanical stimuli, such as shear stress, that can mediate cellular communication. In this study, we developed a Mesenchymal Stem Cell line (MSC) and a Human Umbilical Vein Endothelial Cell line (HUVEC), which were co-cultured under flow conditions in a three-dimensional, porous, natural pullulan\/dextran scaffold that was supplemented with hydroxyapatite crystals that allowed for the spontaneous formation of spheroids. After 2 weeks, their viability was higher under the dynamic conditions (&gt;94%) than the static conditions (&lt;75%), with dead cells central in the spheroids. Mineralization and collagen IV production increased under the dynamic conditions, correlating with osteogenesis and vasculogenesis. The endothelial cells clustered at the spheroidal core by day 7. Proliferation doubled in the dynamic conditions, especially at the scaffold peripheries. Lattice Boltzmann simulations showed negligible wall shear stress in the hydrogel pores but highlighted highly oxygenated zones coinciding with cell proliferation. A strong oxygen gradient likely influenced endothelial migration and cell distribution. Hypoxia was minimal, explaining high viability and spheroid maturation in the dynamic conditions.<\/jats:p>","DOI":"10.3390\/gels10100666","type":"journal-article","created":{"date-parts":[[2024,10,18]],"date-time":"2024-10-18T06:46:52Z","timestamp":1729234012000},"page":"666","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":13,"title":["Bone Spheroid Development Under Flow Conditions with Mesenchymal Stem Cells and Human Umbilical Vein Endothelial Cells in a 3D Porous Hydrogel Supplemented with Hydroxyapatite"],"prefix":"10.3390","volume":"10","author":[{"ORCID":"https:\/\/orcid.org\/0009-0007-6611-4472","authenticated-orcid":false,"given":"Soukaina","family":"El Hajj","sequence":"first","affiliation":[{"name":"Laboratoire de M\u00e9canique Paris-Saclay, CNRS, CentraleSup\u00e9lec, ENS Paris-Saclay, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-2342-2973","authenticated-orcid":false,"given":"Martial Bankou\u00e9","family":"Ntat\u00e9","sequence":"additional","affiliation":[{"name":"Laboratoire de G\u00e9nie des Proc\u00e9d\u00e9s et Mat\u00e9riaux, CentraleSup\u00e9lec, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cyril","family":"Breton","sequence":"additional","affiliation":[{"name":"Laboratoire de G\u00e9nie des Proc\u00e9d\u00e9s et Mat\u00e9riaux, CentraleSup\u00e9lec, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robin","family":"Siadous","sequence":"additional","affiliation":[{"name":"Laboratoire de Bioing\u00e9nierie Tissulaire (BioTis), INSERM U1026, Universit\u00e9 de Bordeaux, 33076 Bordeaux, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rachida","family":"Aid","sequence":"additional","affiliation":[{"name":"Laboratoire de Recherche Vasculaire Translationnelle (LVTS), INSERM U1148, Universit\u00e9 Paris Cit\u00e9, 75018 Paris, France"},{"name":"Laboratoire de Recherche Vasculaire Translationnelle (LVTS), INSERM U1148, Universit\u00e9 Sorbonne Paris Nord, 93430 Villetaneuse, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Magali","family":"Dupuy","sequence":"additional","affiliation":[{"name":"Laboratoire de G\u00e9nie des Proc\u00e9d\u00e9s et Mat\u00e9riaux, CentraleSup\u00e9lec, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2197-9468","authenticated-orcid":false,"given":"Didier","family":"Letourneur","sequence":"additional","affiliation":[{"name":"Laboratoire de Recherche Vasculaire Translationnelle (LVTS), INSERM U1148, Universit\u00e9 Paris Cit\u00e9, 75018 Paris, France"},{"name":"Laboratoire de Recherche Vasculaire Translationnelle (LVTS), INSERM U1148, Universit\u00e9 Sorbonne Paris Nord, 93430 Villetaneuse, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jo\u00eblle","family":"Am\u00e9d\u00e9e","sequence":"additional","affiliation":[{"name":"Laboratoire de Bioing\u00e9nierie Tissulaire (BioTis), INSERM U1026, Universit\u00e9 de Bordeaux, 33076 Bordeaux, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0032-0085","authenticated-orcid":false,"given":"Herv\u00e9","family":"Duval","sequence":"additional","affiliation":[{"name":"Laboratoire de G\u00e9nie des Proc\u00e9d\u00e9s et Mat\u00e9riaux, CentraleSup\u00e9lec, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5520-8613","authenticated-orcid":false,"given":"Bertrand","family":"David","sequence":"additional","affiliation":[{"name":"Laboratoire de M\u00e9canique Paris-Saclay, CNRS, CentraleSup\u00e9lec, ENS Paris-Saclay, Universit\u00e9 Paris-Saclay, 91190 Gif-sur-Yvette, France"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,10,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"136","DOI":"10.1007\/s11657-013-0136-1","article-title":"Osteoporosis in the European Union: Medical Management, Epidemiology and Economic Burden","volume":"8","author":"Hernlund","year":"2013","journal-title":"Arch. 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