A phenotypic comparison of osteoblast cell lines versus human primary osteoblasts for biomaterials testing E. M. Czekanska, 1,2 M. J. Stoddart, 1 J. R. Ralphs, 2 R. G. Richards, 1,2 J. S. Hayes 1,3 1 AO Research Institute Davos, Davos Platz, Switzerland 2 Cardiff School of Biosciences, Cardiff University, Cardiff, Wales, Great Britain 3 The Department of Orthobiologics, Regenerative Medicine Institute, National Centre for Biomedical Engineering Science, National University of Ireland, Galway, Ireland Received 11 June 2013; revised 26 July 2013; accepted 21 August 2013 Published online 00 Month 2013 in Wiley Online Library (wileyonlinelibrary.com). DOI: 10.1002/jbm.a.34937 Abstract: Immortalized cell lines are used more frequently in basic and applied biology research than primary bone- derived cells because of their ease of access and repeatability of results in experiments. It is clear that these cell models do not fully resemble the behavior of primary osteoblast cells. Although the differences will affect the results of biomaterials testing, they are not clearly defined. Here, we focused on comparing proliferation and maturation potential of three osteoblast cell lines, SaOs2, MG-63, and MC3T3-E1 with pri- mary human osteoblast (HOb) cells to assess their suitability as in vitro models for biomaterials testing. We report similar- ities in cell proliferation and mineralization between primary cells and MC3T3-E1. Both, SaOs2 and MG-63 cells demon- strated a higher proliferation rate than HOb cells. In addition, SaOs2, but not MG-63, cells demonstrated similar ALP activi- ty, mineralization potential and gene regulation to HOb’s. Our results demonstrate that despite SaOs-2, MG63, and MC3T3 cells being popular choices for emulating osteoblast behav- ior, none can be considered appropriate replacements for HOb’s. Nevertheless, these cell lines all demonstrated some distinct similarities with HOb’s, thus when applied in the cor- rect context are a valuable in vitro pilot model of osteoblast functionality, but should not be used to replace primary cell studies. V C 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 00A:000–000, 2013. Key Words: osteoblast, differentiation, primary cell, cell line, In vitro cell model How to cite this article: Czekanska EM, Stoddart MJ, Ralphs JR, Richards RG, Hayes JS. 2013. A phenotypic comparison of osteoblast cell lines versus human primary osteoblasts for biomaterials testing. J Biomed Mater Res Part A 2013: 00A: 000–000. INTRODUCTION The cost relating to the treatment and management of bone related orthopedic disorders accounts for billions annually. 1 Preclinical models, although not perfect, offer a prescreening method for testing novel biomaterials. 2 However, to ethically rationalize the use of animals and to further our detailed knowledge, in vitro testing is required. In terms of bone research, a plethora of osteoblast cell models have emerged to tackle the surge in research in this area and to compen- sate to some degree for the ever present problem relating to sustainable supply of primary cells from either healthy or diseased donors. These include cells isolated from various species, 3–7 immortalized cell lines, 8,9 malignant cells, 10–12 and more recently induced pluripotent stem cells. 13,14 All of these cell models are employed for basic bone biology investigations, as well as assessment of novel drugs and bio- materials. Immortalized cell lines are mostly used due to the availability of unlimited number of cells, ease of culture and higher phenotypical stability compared to primary cells. 15 Moreover, in the case of human-derived cell lines, both normal and malignant, the concern about species dif- ferences can be avoided. 16 In contrast, the reported drawbacks of cell lines are many. 8,17,18 One of the main issues regarding cell lines is their stage specific differentiation profile which essentially limits comprehensive study of phenotypic features of associ- ated with primary human osteoblast cells. 15,19 Thus, their use as prescreening in vitro models is somewhat limited to specific research questions, though often interpretations beyond their scope are given in publications, which can cause confusion. Furthermore, aberrations in cell mitotic processes limits their use in long term in vitro investiga- tions as the lack of growth inhibition has negative influence on later stages of osteoblast cell phenotype development. 20 Despite being characterized as phenotypically stable and homogenous, previous reports suggest cell proliferation rate and phenotype changes between cells from early and late passages. 17,18,21 The potential of isolating clones from cell lines contradicts the opinion of their homogenous phenotype. 8,22,23 Correspondence to: M. J. Stoddart; e-mail: martin.stoddart@aofoundation.org Contract grant sponsor: EU FP7 Biodesign V C 2013 WILEY PERIODICALS, INC. 1