title: Elucidating the individual effects of calcium and phosphate ions on hMSCs by using composite materials.
abstract: The biological performance of bone graft substitutes based on calcium phosphate bioceramics is dependent on a number of properties including chemical composition, porosity and surface micro- and nanoscale structure. However, in contemporary bioceramics these properties are interlinked, therefore making it difficult to investigate the individual effects of each property on cell behavior. In this study we have attempted to investigate the effects of calcium and inorganic phosphate ions independent from one another by preparing composite materials with polylactic acid (PLA) as a polymeric matrix and calcium carbonate or sodium phosphate salts as fillers. Clinically relevant bone marrow derived human mesenchymal stromal cells (hMSCs) were cultured on these composites and proliferation, osteogenic differentiation and ECM mineralization were investigated with time and were compared to plain PLA control particles. In parallel, cells were also cultured on conventional cell culture plates in media supplemented with calcium or inorganic phosphate to study the effect of these ions independent of the 3D environment created by the particles. Calcium was shown to increase proliferation of cells, whereas both calcium and phosphate positively affected alkaline phosphatase enzyme production. QPCR analysis revealed positive effects of calcium and of inorganic phosphate on the expression of osteogenic markers, in particular bone morphogenetic protein-2 and osteopontin. Higher levels of mineralization were also observed upon exposure to either ion. Effects were similar for cells cultured on composite materials and those cultured in supplemented media, although ion concentrations in the composite cultures were lower. The approach presented here may be a valuable tool for studying the individual effects of a variety of soluble compounds, including bioinorganics, without interference from other material properties.
authors: Department of Tissue Regeneration, University of Twente, P.O. Box 217, 7500 AE Enschede, The Netherlands.
authors: Charlène B S S
authors: CB
authors: Danoux
authors: Faculty of Dentistry, McGill University, 3640 University Street, Montreal, QC H3 A 2B2, Canada.
authors: David C
authors: DC
authors: Bassett
authors: Department of Tissue Regeneration, University of Twente, P.O. Box 217, 7500 AE Enschede, The Netherlands.
authors: Ziryan
authors: Z
authors: Othman
authors: Department of Tissue Regeneration, University of Twente, P.O. Box 217, 7500 AE Enschede, The Netherlands; 3B's Research Group, Universidade do Minho, AvePark, 4806-909 Caldas das Taipas, Portugal; ICVS/3B's - PT Government Associated Laboratory, Guimarães, Portugal.
authors: Ana I
authors: AI
authors: Rodrigues
authors: 3B's Research Group, Universidade do Minho, AvePark, 4806-909 Caldas das Taipas, Portugal; ICVS/3B's - PT Government Associated Laboratory, Guimarães, Portugal.
authors: Rui L
authors: RL
authors: Reis
authors: Faculty of Dentistry, McGill University, 3640 University Street, Montreal, QC H3 A 2B2, Canada.
authors: Jake E
authors: JE
authors: Barralet
authors: Department of Tissue Regeneration, University of Twente, P.O. Box 217, 7500 AE Enschede, The Netherlands.
authors: Clemens A
authors: CA
authors: van Blitterswijk
authors: Department of Tissue Regeneration, University of Twente, P.O. Box 217, 7500 AE Enschede, The Netherlands. Electronic address: p.habibovic@maastrichtuniversity.nl.
authors: Pamela
authors: P
authors: Habibovic
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D001672:Biocompatible Materials / Q000737:chemistry*; D055396:Bone Morphogenetic Protein 2 / Q000378:metabolism; D002118:Calcium / Q000737:chemistry*; D002130:Calcium Phosphates / Q000737:chemistry*; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002516:Ceramics; D006801:Humans; D007477:Ions; D019344:Lactic Acid / Q000737:chemistry; D008422:Materials Testing; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008855:Microscopy, Electron, Scanning; D010012:Osteogenesis; D053495:Osteopontin / Q000378:metabolism; D010316:Particle Size; D010710:Phosphates / Q000737:chemistry*; D011091:Polyesters; D011108:Polymers / Q000737:chemistry
chemical_list: C525718:BMP2 protein, human; D001672:Biocompatible Materials; D055396:Bone Morphogenetic Protein 2; D002130:Calcium Phosphates; D007477:Ions; D010710:Phosphates; D011091:Polyesters; D011108:Polymers; C506895:SPP1 protein, human; D053495:Osteopontin; D019344:Lactic Acid; C033616:poly(lactide); C020243:calcium phosphate; D000469:Alkaline Phosphatase; D002118:Calcium
keywords: Bone graft substitute; Calcium; Composite; Inorganic phosphate; hMSCs