A biaxial rotating bioreactor for the culture of fetal mesenchymal stem cells for bone tissue engineering Zhi-Yong Zhang a, b , Swee Hin Teoh b, c , Woon-Shin Chong d , Toon-Tien Foo d , Yhee-Cheng Chng d , Mahesh Choolani e , Jerry Chan e, * a Graduate Program in Bioengineering (GPBE), National University of Singapore, Singapore b Centre for Biomedical Materials Applications and Technology (BIOMAT), Department of Mechanical Engineering, Faculty of Engineering, National University of Singapore, Singapore c National University of Singapore Tissue Engineering Programme (NUSTEP), National University of Singapore, Singapore d Bioengineering Laboratory, Technology Centre for Life Sciences, Singapore Polytechnic, Singapore e Experimental Fetal Medicine Group, Department of Obstetrics and Gynaecology, Yong Loo Lin School of Medicine, National University of Singapore and National University Hospital, Singapore article info Article history: Received 2 November 2008 Accepted 13 January 2009 Available online xxx Keywords: Bioreactor Fetal mesenchymal stem cell Bone tissue engineering Polycaprolactone NOD/SCID mice abstract The generation of effective tissue engineered bone grafts requires efficient exchange of nutrients and mechanical stimulus. Bioreactors provide a manner in which this can be achieved. We have recently developed a biaxial rotating bioreactor with efficient fluidics through in-silico modeling. Here we investigated its performance for generation of highly osteogenic bone graft using polycaprolactone– tricalcium phosphate (PCL–TCP) scaffolds seeded with human fetal mesenchymal stem cell (hfMSC). hfMSC scaffolds were cultured in either bioreactor or static cultures, with assessment of cellular viability, proliferation and osteogenic differentiation in vitro and also after transplantation into immunodeficient mice. Compared to static culture, bioreactor-cultured hfMSC scaffolds reached cellular confluence earlier (day 7 vs. day 28), with greater cellularity (2, p < 0.01), and maintained high cellular viability in the core, which was 2000 mm from the surface. In addition, bioreactor culture was associated with greater osteogenic induction, ALP expression (1.5 p < 0.01), calcium deposition (5.5, p < 0.001) and bony nodule formation on SEM, and in-vivo ectopic bone formation in immunodeficient mice (3.2, p < 0.001) compared with static-cultured scaffolds. The use of biaxial bioreactor here allowed the maintenance of cellular viability beyond the limits of conventional diffusion, with increased proliferation and osteogenic differentiation both in vitro and in vivo, suggesting its utility for bone tissue engineering applications. Ó 2009 Elsevier Ltd. All rights reserved. 1. Introduction Bone tissue engineering provides a promising approach to address the significant drawbacks of existing bone grafts, which include: firstly, the limited availability of bone tissue and donor-site morbidity associated with autografts [1,2]; secondly, the significant risk of disease transmission and immune reaction arising from the use of allografts [3]; and lastly, the lack of remodeling and subse- quent fatigue-associated graft failure with the use synthetic grafts [4]. In bone tissue engineering, biodegradable porous scaffolds are seeded with osteogenic cell types to develop an in-vitro-matured engineered cellular bone graft, which can be fashioned into different shapes and sizes, stimulate bone healing and remodel accordingly [5]. Three dimensional (3D) scaffolds provide the necessary support for cells to attach, proliferate and differentiate, and define the overall shape of the tissue engineered transplant [6]. Scaffolds made of polycaprolactone (PCL) have recently been shown to possess favorable properties for load bearing bone tissue engi- neering application compared with other materials such as PLGA [7]. They have a slower degradation speed with structural degra- dation kinetics of only 7% over a six month period in vivo [8], maintaining a sustained period of mechanical support. In addition the slow degradation kinetics would result in a reduced risk of acidosis from the rapid accumulation of acidic by-products, sug- gesting its utility for bone tissue engineering applications. Mesenchymal stem cells (MSCs) are increasingly being used as a cellular source for osteogenic tissue engineering applications, due to their ease of isolation and well defined osteogenic differentiation pathways [9,10]. However, the use of human adult MSC has been * Corresponding author. Experimental Fetal Medicine Group, Department of Obstetrics and Gynaecology, Yong Loo Lin School of Medicine, National University of Singapore, 5 Lower Kent Ridge Road, Singapore 119074. Tel.: þ65 6772 2672; fax: þ65 6779 4753. E-mail address: jerrychan@nus.edu.sg (J. Chan). Contents lists available at ScienceDirect Biomaterials journal homepage: www.elsevier.com/locate/biomaterials ARTICLE IN PRESS 0142-9612/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.biomaterials.2009.01.028 Biomaterials xxx (2009) 1–11 Please cite this article in press as: Zhang Z-Yet al., A biaxial rotating bioreactor for the culture of fetal mesenchymal stem cells for bone tissue engineering, Biomaterials (2009), doi:10.1016/j.biomaterials.2009.01.028