Micro- and nanostructured hydroxyapatite–collagen microcarriers for bone tissue-engineering applications † R. A. Perez 1,2 , G. Altankov 3,4 , E. Jorge-Herrero 5 and M. P. Ginebra 1,2 * 1 Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgy, Technical University of Catalonia (UPC), Avda. Diagonal 647, E-08028 Barcelona, Spain 2 Biomedical Research Networking Centre in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), María de Luna 11, Ed. CEEI, 50118 Zaragoza, Spain 3 Institute for Bioengineering of Catalonia, Scientific Park, Josep Samitier 1-5, 08028 Barcelona, Spain 4 Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain 5 U. Biomateriales-Ingeniería Tisular, Hospital Puerta del Hierro Majadahonda, Manuel de Falla 2, 28222 Majadahonda, Madrid, Spain Abstract Novel hydroxyapatite (HA)–collagen microcarriers (MCs) with different micro/nanostructures were devel- oped for bone tissue-engineering applications. The MCs were fabricated via calcium phosphate cement (CPC) emulsion in oil. Collagen incorporation in the liquid phase of the CPC resulted in higher MC sphericity. The MCs consisted of a porous network of entangled hydroxyapatite crystals, formed as a result of the CPC setting reaction. The addition of collagen to the MCs, even in an amount as small as 0.8wt%, resulted in an improved interaction with osteoblast-like Saos-2 cells. The micro/nanostructure and the surface texture of the MCs were further tailored by modifying the initial particle size of the CPC. A synergistic effect between the presence of collagen and the nanosized HA crystals was found, resulting in significantly enhanced alkaline phosphatase activity on the collagen-containing nanosized HA MCs. Copyright © 2012 John Wiley & Sons, Ltd. Received 30 December 2010; Revised 15 July 2011; Accepted 26 September 2011 Keywords hydroxyapatite; microcarrier; collagen; calcium phosphate cement; bone regeneration; cell response 1. Introduction Microcarriers (MCs) were originally developed for cultur- ing anchorage-dependent cells in suspension (Van Wezel, 1967). Apart from their biotechnological applications, how- ever, recent studies have shown their potential for tissue- engineering purposes, as vectors for the delivery of cells or drug release vehicles (Freiberg and Zhu, 2004; Brandon et al., 2007; Ng et al., 2008; Malda and Frondoza, 2006; Malda et al., 2003). Suspensions of MCs can easily be injected or implanted in the diseased tissue, such as bone, and if they are colonized with cells, MCs can function as scaffolds for bone tissue repair with osteogenic potential (Malda and Frondoza, 2006). For this specific application, it is important to fabricate MCs of a material that has the ability to enhance bone ingrowth in vivo. In this context, the development of MCs of osteoconductive and osteoin- ductive materials, such as calcium phosphates, can be espe- cially useful (Cancedda et al., 2007; Fischer et al., 2003; Yuan et al., 2010). Hydroxyapatite (HA)/collagen-based materials have been extensively used as substrates for bone tissue engi- neering, as they mimic the extracellular bone matrix. Interesting results have been reported for this kind of material, combining the osteoconductivity of HA with an enhanced recognition from the cells provided by collagen incorporation (Schliephake et al., 2004; Kikuchi et al., 2004; Chaozong et al., 2009; Shunji et al., 2007; Miyamoto et al., 1998; Tamimi et al., 2008; Moreau et al., 2009). Microspheres made of collagen and HA have been previ- ously obtained by emulsion of a collagen solution in oil (Hsu et al., 1999; Wu et al., 2004; Kim et al., 2007). In these studies, however, the collagen was the continuous phase, its content in the range 35–70 wt%, and the stabilization * Correspondence to: M. P. Ginebra, Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgy, Technical University of Catalonia (UPC), Avda. Diagonal 647, E-08028 Barcelona, Spain. E-mail: maria.pau. ginebra@upc.edu † This study did not have direct ethical implications and therefore was not submitted to ethical committee approval. Copyright © 2012 John Wiley & Sons, Ltd. JOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE RESEARCH ARTICLE J Tissue Eng Regen Med (2012) Published online in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/term.530