Chemical Engineering Journal 426 (2021) 131321 Available online 14 July 2021 1385-8947/© 2021 Elsevier B.V. All rights reserved. Effect of zinc-doped hydroxyapatite/graphene nanocomposite on the physicochemical properties and osteogenesis differentiation of 3D-printed polycaprolactone scaffolds for bone tissue engineering Hossein Maleki-Ghaleh a , M. Hossein Siadati b , Ali Fallah c, d , Ali Zarrabi d , Ferdows Afghah c, d , Bahattin Koc c, d , Elaheh Dalir Abdolahinia a , Yadollah Omidi e , Jaleh Barar a, f , Ali Akbari-Fakhrabadi g , Younes Beygi-Khosrowshahi h , Khosro Adibkia a, f, * a Research Center for Pharmaceutical Nanotechnology, Biomedicine Institute, Tabriz University of Medical Sciences, Tabriz, Iran b Faculty of Materials Science and Engineering, K. N. Toosi University of Technology, Tehran, Iran c Faculty of Engineering and Natural Sciences, Sabanci University, Istanbul, Turkey d Nanotechnology Research and Application Center (SUNUM), Sabanci University, Istanbul, Turkey e Department of Pharmaceutical Sciences, College of Pharmacy, Nova Southeastern University, Fort Lauderdale, FL, USA f Department of Pharmaceutics, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran g Advanced Materials Laboratory, Department of Mechanical Engineering, University of Chile, Santiago, Chile h Department of Chemical Engineering, Faculty of Engineering, Azarbaijan Shahid Madani University, Tabriz, Iran A R T I C L E INFO Keywords: 3D printed bone scaffold Polycaprolactone Zinc doped hydroxyapatite Graphene Osteogenesis ABSTRACT Processing and composition can signifcantly affect the mechanobiology, biodegradability, and cellular behavior of polymer-based bone scaffolds to replace damaged bone tissue. In this research, hydroxyapatite (HA), zinc- doped HA (ZnHA), and ZnHA-graphene (ZnHA-rGO) nanoparticles are composed in a polycaprolactone (PCL) matrix. After compositing PCL with nanoparticles, 3D bone scaffolds were built by a custom-built 3D printing system. The characterization of nanoparticles was extensively investigated by TEM, EDX-MAP, XRD, and ATR- FTIR. Simultaneously, 3D-printed scaffolds with different compositions were studied in terms of structure, morphology, thermogravimetry, biodegradability, and mechanical behaviors. The FE-SEM images of the scaf- folds showed a highly regular structure and good printability of the developed material system. Moreover, the stiffness modulus of the samples increased due to the presence of the nanoparticles, especially in the ZnHA-rGO nanocomposite. In vitro cell assessment of 3D bone scaffolds was investigated by cell viability tests, cell attachment, and alizarin red staining via mesenchymal stem cells (MSCs). For differentiation capacity of the developed scaffolds, stem cell osteogenesis differentiation was studied by RT-PCR to analyze the ALP, RUNX2, BMP2, TGFβ, and OCN genes. The cellular assessments revealed an increase in PCL scaffold’s cell osteogenesis due to the HA nanoparticles in the scaffold matrix. Zinc doping in the HA nanoparticles and rGO addition signifcantly increased the osteogenesis of MSCs. In particular, the nanocomposite of ZnHA-rGO in PCL scaffold matrix signifcantly improved the osteogenic differentiation and, thus, it is a viable option for effective regen- eration of damaged bone tissue. 1. Introduction Hard tissue damages due to accidents and diseases such as cancerous sarcoma tumors are one of the most common health problems [1,2]. The human body’s capacity to produce new bone tissue depends on the size of the damaged area [3]. A replacement tissue is needed to solve this problem, and the tissue can be transferred from other hard tissues of the patient’s body, requiring secondary surgery and subsequent issues [4]. A major goal of tissue engineering is to provide a suitable solution for replacing damaged tissues such as bone [5]. Bioceramics are known as one of the most remarkable materials in bone replacement and recon- struction [6]. The similarity of calcium phosphate compounds to the mineral phase of bone tissues and their biocompatibility has led to the widespread use of these substances in the body [7]. Hydroxyapatite * Corresponding author. E-mail address: adibkia@tbzmed.ac.ir (K. Adibkia). Contents lists available at ScienceDirect Chemical Engineering Journal journal homepage: www.elsevier.com/locate/cej https://doi.org/10.1016/j.cej.2021.131321 Received 19 May 2021; Received in revised form 3 July 2021; Accepted 11 July 2021