Abstract
BackgroundA novel biodegradable scaffold including gelatin (G), chitooligosaccharide (COS), and demineralized bone matrix (DBM) could play a significant part in bone tissue engineering. The present study aimed to investigate the biological characteristics of composite scaffolds in combination of G, COS, and DBM for in vitro cell culture and in vivo animal bioassays.MethodsThree-dimensional scaffolds from the mixture of G, COS, and DBM were fabricated into 3 groups, namely, G, GC, and GCD using a lyophilization technique. The scaffolds were cultured with mesenchymal stem cells (MSCs) for 4 weeks to determine biological responses such as cell attachment and cell proliferation, alkaline phosphatase (ALP) activity, calcium deposition, cell morphology, and cell surface elemental composition. For the in vivo bioassay, G, GC, and GCD, acellular scaffolds were implanted subcutaneously in 8-week-old male Wistar rats for 4 weeks and 8 weeks. The explants were assessed for new bone formation using hematoxylin and eosin (H&E) staining and von Kossa staining.ResultsThe MSCs could attach and proliferate on all three groups of scaffolds. Interestingly, the ALP activity of MSCs reached the greatest value on day 7 after cultured on the scaffolds, whereas the calcium assay displayed the highest level of calcium in MSCs on day 28. Furthermore, weight percentages of calcium and phosphorus on the surface of MSCs after cultivation on the GCD scaffolds increased when compared to those on other scaffolds. The scanning electron microscopy images showed that MSCs attached and proliferated on the scaffold surface thoroughly over the cultivation time. Mineral crystal aggregation was evident in GC and greatly in GCD scaffolds. H&E staining illustrated that G, GC, and GCD scaffolds displayed osteoid after 4 weeks of implantation and von Kossa staining confirmed the mineralization at 8 weeks in G, GC, and GCD scaffolds.ConclusionThe MSCs cultured in GCD scaffolds revealed greater osteogenic differentiation than those cultured in G and GC scaffolds. Additionally, the G, GC, and GCD scaffolds could promote in vivo ectopic bone formation in rat model. The GCD scaffolds exhibited maximum osteoinductive capability compared with others and may be potentially used for bone regeneration.
Highlights
Demineralized bone matrix (DBM) is well-known as an osteoinductive biomaterial utilized for tissue engineering
The mesenchymal stem cells (MSCs) cultured in G/ COS/DBM (GCD) scaffolds revealed greater osteogenic differentiation than those cultured in G and GC scaffolds
The G, GC, and GCD scaffolds could promote in vivo ectopic bone formation in rat model
Summary
Demineralized bone matrix (DBM) is well-known as an osteoinductive biomaterial utilized for tissue engineering. When DBM is combined with osteoprogenitors and implanted into animals, it could support and induce new bone formation in in vivo [1]. DBM could trigger osteoinductivity both in vitro and in vivo. Alidadi and colleagues evaluated DBM combined with chitosan and polymethylmethacrylate in radial bone defect of rat model, showing that DBM provided significantly superior biocompatibility, biodegradability, osteoconductivity, and osteoinductivity to the scaffold [3]. DBM significantly improved bone healing close to the autologous cortical bone graft. A novel biodegradable scaffold including gelatin (G), chitooligosaccharide (COS), and demineralized bone matrix (DBM) could play a significant part in bone tissue engineering. The present study aimed to investigate the biological characteristics of composite scaffolds in combination of G, COS, and DBM for in vitro cell culture and in vivo animal bioassays
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