A novel gelatin/chitooligosaccharide/demineralized bone matrix composite scaffold and periosteum-derived mesenchymal stem cells for bone tissue engineering

Abstract

Background 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. Methods Three-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. Results The 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. Conclusion The 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.

Comparative evaluation of mesenchymal stromal cell growth and osteogenic differentiation on a shape memory polymer scaffold

“…Unlike mineralization, the maximum ALP activity in osteogenic differentiation media was relatively similar for each cell type, albeit more transient, with all cell types reaching a similar level, although at different time points. This is consistent with previous studies, which have demonstrated transient levels of ALP activity on scaffolds, with levels increasing at one time‐point but then subsequently decreasing 29–33 . Overall, in terms of viability, ALP activity, and mineralization assays, the data strongly supports BMSCs, ADSCs, and CHSCs as promising candidates for successful in vivo bone healing with the SMP scaffold.…”

“…This is consistent with previous studies, which have demonstrated transient levels of ALP activity on scaffolds, with levels increasing at one time-point but then subsequently decreasing. [29][30][31][32][33] Overall, in terms of viability, ALP activity, and mineralization assays, the data strongly supports BMSCs, ADSCs, and CHSCs as promising candidates for successful in vivo bone healing with the SMP scaffold.…”

Mineralization in a Critical Size Bone-Gap in Sheep Tibia Improved by a Chitosan-Calcium Phosphate-Based Composite as Compared to Predicate Device

“…In a review paper, titanium surfaces functionalized with chitosan were concluded to have greater osseointegration capacity than uncoated controls [27]. Thitiset et al [28], used chitoologosaccharide for preparation of composites that did show promising results in cell culture, as well as for new bone formation in vivo. The results from Thitiset et al are remarkable in the light of our ideas on the mechanism of action of chitosan in bone regeneration where chitooligosachharides play a central role [17].…”

A review on background, process and application of electrospun nanofibers for tissue regeneration

“…The scaffold triggered the proliferation and differentiation of osteoblast cells and proved to be an efficient bone graft with excellent osteogenic potency for bone tissue engineering applications. 66 Chitosan. Chitosan is a polysaccharide that is extracted by deacetylation of chitin.…”