Synthesis and Characterization of Hydroxyl-Functionalized Caprolactone Copolymers and Their Effect on Adhesion, Proliferation, and Differentiation of Human Mesenchymal Stem Cells Hajar Seyednejad, † Tina Vermonden, † Natalja E. Fedorovich, ‡ Roel van Eijk, † Mies J. van Steenbergen, † Wouter J. A. Dhert, ‡,§ Cornelus F. van Nostrum, † and Wim E. Hennink* ,† Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Faculty of Science, Utrecht University, P.O. Box 80082, 3508 TB Utrecht, The Netherlands, Department of Orthopaedics, University Medical Center Utrecht, P.O. Box 85500, 3508 GA Utrecht, The Netherlands, and Faculty of Veterinary Medicine, Utrecht University, P.O. Box 80163, 3508 TD Utrecht, The Netherlands Received June 17, 2009; Revised Manuscript Received August 31, 2009 The aim of this study was to develop new hydrophilic polyesters for tissue engineering applications. In our approach, poly(benzyloxymethyl glycolide-co-ε-caprolactone)s (pBHMG-CLs) were synthesized through melt copolymer- ization of ε-caprolactone (CL) and benzyl-protected hydroxymethyl glycolide (BHMG). Deprotection of the polymers yielded copolymers with pendant hydroxyl groups, poly(hydroxymethylglycolide-co-ε-caprolactone) (pHMG-CL). The synthesized polymers were characterized by GPC, NMR, and DSC techniques. The resulting copolymers consisting of up to 10% of HMG monomer were semicrystalline with a melting temperature above body temperature. Water contact angle measurements of polymeric films showed that increasing HMG content resulted in higher surface hydrophilicity, as evidenced from a decrease in receding contact angle from 68° for PCL to 40° for 10% HMG-CL. Human mesenchymal stem cells showed good adherence onto pHMG-CL films as compared to the more hydrophobic PCL surfaces. The cells survived and were able to differentiate toward osteogenic lineage on pHMG-CL surfaces. This study shows that the aforementioned hydrophilic polymers are attractive candidates for the design of scaffolds for tissue engineering applications. 1. Introduction Tissue engineering is a multidisciplinary field, which com- bines the principles of engineering and life sciences toward generating replacements for biological tissues and organs that have lost their function due to a disease or an accident. 1,2 The key elements for the success of tissue regeneration are cells, scaffold, cell/matrix interactions, along with signaling and therapeutic molecules. 3 Scaffolds, those porous, three-dimen- sional, temporary structures, play an important role in manipu- lating cell function and guidance of new organ formation. 4 The scaffolds reported so far are mainly based on biodegradable polymers among which are aliphatic polyesters such as PLA, PGA, PCL, and their copolymers. 1,5-8 While the ester groups in the main chain of these polymers ensure biodegradation due to hydrolysis, the absence of pendant functional groups limits the versatility of these materials. 9 Besides, these polymers are mostly hydrophobic materials and their application is limited when cell adhesion is considered. There are different methods to improve the surface character- istics of polymeric scaffolds such as plasma treatment, 10-12 surface entrapment of a second polymer, 13-15 and partial surface hydrolysis by acid or base treatment, 16 but these methods have some limitations, such as a limited amount of penetration into pores of the scaffold or causing the degradation of the aliphatic polyester scaffolds. 17 The introduction of functional groups in these polymers is an important alternative to tailor the physical and chemical properties such as hydrophilicity and degradation rate 18-21 and also opens opportunities for further functional- ization with biologically active molecules 22-25 to enhance cell/ matrix interaction. The goal of this study was to develop a new functionalized polymer with appropriate physical properties and controlled cell response. Therefore, we recently introduced a novel function- alized dilactone with protected hydroxyl groups, benzyloxy- methyl glycolide. 26 The deprotected homopolymers of this monomer are not suitable for making scaffolds for tissue engineering due to its rapid degradation. 27 Hence, in the present work we synthesized and deprotected random copolymers of benzyl protected hydroxymethyl glycolide (BHMG) and ε-ca- prolactone (CL). CL was chosen because poly(ε-caprolactone) is a biocompatible and biodegradable polymer widely used in biomedical applications. However, poly(ε-caprolactone) de- grades very slowly, 28 with a degradation rate ranging from 2 to 4 years, 29 and the polymer is not favorable for cell adhesion and proliferation due to its intrinsic hydrophobicity and lack of bioactive functional groups. 30,31 In this study, we investigated the effect of chemical structure of these functionalized polyesters on their physical properties as well as on adhesion, survival, and osteogenic differentiation of human mesenchymal stem cells (hMSCs), which are important parameters to design scaffolds for tissue engineering applications. 2. Materials and Methods 2.1. Materials. All chemicals used in this study were purchased from Aldrich and used as received, unless stated otherwise. All solvents were purchased from Biosolve (Valkenswaard, The Netherlands) except * To whom correspondence should be addressed. E-mail: w.e.hennink@uu.nl. † Department of Pharmaceutics. ‡ Department of Orthopaedics. § Faculty of Veterinary Medicine. Biomacromolecules 2009, 10, 3048–3054 3048 10.1021/bm900693p CCC: $40.75  2009 American Chemical Society Published on Web 10/06/2009