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Materials (Basel). 2019 Mar 19;12(6). pii: E919. doi: 10.3390/ma12060919.

Bone Tissue Engineering in a Perfusion Bioreactor Using Dexamethasone-Loaded Peptide Hydrogel.

Author information

1
Department of Biology, Faculty of Science, University of Zagreb, 10 000 Zagreb, Croatia. ingam@biol.pmf.hr.
2
Center for Translational and Clinical Research, School of Medicine, University of Zagreb, 10000 Zagreb, Croatia. ingam@biol.pmf.hr.
3
Department of Biology, Faculty of Science, University of Zagreb, 10 000 Zagreb, Croatia. maja.antunovic@biol.pmf.hr.
4
Department of Biology, Faculty of Science, University of Zagreb, 10 000 Zagreb, Croatia. lidija.pribolsan@gmail.com.
5
Department of Histology and Embryology, School of Medicine, University of Zagreb, 10000 Zagreb, Croatia. alan.ivkovic@gmail.com.
6
Department of Orthopaedic Surgery, University Hospital Sveti Duh, 10000 Zagreb, Croatia. alan.ivkovic@gmail.com.
7
Department of Biotechnology, University of Rijeka, 51000 Rijeka, Croatia. alan.ivkovic@gmail.com.
8
Department of Material Chemistry, Rudjer Boskovic Institute, 10000 Zagreb, Croatia. gotic@irb.hr.
9
Department of Histology and Embryology, School of Medicine, University of Zagreb, 10000 Zagreb, Croatia. andreja_vukasovic@yahoo.com.
10
Department of Biology, Faculty of Science, University of Zagreb, 10 000 Zagreb, Croatia. katarina.caput.mihalic@biol.pmf.hr.
11
Department of Biology, Faculty of Science, University of Zagreb, 10 000 Zagreb, Croatia. maja.pusic@biol.pmf.hr.
12
Department of Material Chemistry, Rudjer Boskovic Institute, 10000 Zagreb, Croatia. Tanja.Jurkin@irb.hr.

Abstract

The main goal of this study was the formation of bone tissue using dexamethasone (DEX)-loaded [COCH₃]-RADARADARADARADA-[CONH₂] (RADA 16-I) scaffold that has the ability to release optimal DEX concentration under perfusion force. Bone-marrow samples were collected from three patients during a hip arthroplasty. Human mesenchymal stem cells (hMSCs) were isolated and propagated in vitro in order to be seeded on scaffolds made of DEX-loaded RADA 16-I hydrogel in a perfusion bioreactor. DEX concentrations were as follows: 4 × 10-3, 4 × 10-4 and 4 × 10-5 M. After 21 days in a perfusion bioreactor, tissue was analyzed by scanning electron microscopy (SEM) and histology. Markers of osteogenic differentiation were quantified by real-time polymerase chain reaction (RT-PCR) and immunocytochemistry. Minerals were quantified and detected by the von Kossa method. In addition, DEX release from the scaffold in a perfusion bioreactor was assessed. The osteoblast differentiation was confirmed by the expression analysis of osteoblast-related genes (alkaline phosphatase (ALP), collagen I (COL1A1) and osteocalcin (OC). The hematoxylin/eosin staining confirmed the presence of cells and connective tissue, while SEM revealed morphological characteristics of cells, extracellular matrix and minerals-three main components of mature bone tissue. Immunocytochemical detection of collagen I is in concordance with given results, supporting the conclusion that scaffold with DEX concentration of 4 × 10-4 M has the optimal engineered tissue morphology. The best-engineered bone tissue is produced on scaffold loaded with 4 × 10-4 M DEX with a perfusion rate of 0.1 mL/min for 21 days. Differentiation of hMSCs on DEX-loaded RADA 16-I scaffold under perfusion force has a high potential for application in regenerative orthopedics.

KEYWORDS:

RADA 16-I; dexamethasone; human mesenchymal stem cells; hydrogel; osteodifferentiation; perfusion bioreactor

PMID:
30893951
DOI:
10.3390/ma12060919
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