Experimental Research on Ectopic Osteogenesis of BMP2-derived Peptide P24 Combined with PLGA Copolymers

To experimentally evaluate the ectopic osteogenetic capacity of synthesized BMP2-derived peptide P24 combined with poly lactic-co-glycolic acid （PLGA）, Wistar rats were divided into two groups： group A, in which BMP2-derived peptide P24/PLGA complex was implanted, and group B which received simple PLGA implant. The complex was respectively implanted into the back muscles of rats. Samples were taken the 1st, 4th, 8th, and the 12th week after the implantation. Their bone formation was detected by X-ray examination, and tissue response was histologically observed. Western blotting was used for the detection of the expression of collagen Ⅰ （Col- Ⅰ ） and osteopontin （OPN）. There was acute inflammation in the tissue around both types of implants at early stage. The cartilage was found around implant areas 4 weeks after the implantation of BMP2-derived peptide p24/PLGA complex, 8 weeks after the implantation, osteoblasts were found, and 12 weeks after the implantation, typical trabecular bone structure was observed. In group B, after 12 weeks, no osteoblasts were found. It is concluded that PLGA is an ideal scaffold material for bone tissue engineering. BMP2-derived peptide can start endochondral ossification and is more effective in inducing ectopic osteogenesis.

Vascularization and osteogenesis in ectopically implanted bone tissue-engineered constructs with endothelial and osteogenic differentiated adipose-derived stem cells

BACKGROUND A major problem in the healing of bone defects is insufficient or absent blood supply within the defect.To overcome this challenging problem,a plethora of approaches within bone tissue engineering have been developed recently.Bearing in mind that the interplay of various diffusible factors released by endothelial cells(ECs)and osteoblasts(OBs)have a pivotal role in bone growth and regeneration and that adjacent ECs and OBs also communicate directly through gap junctions,we set the focus on the simultaneous application of these cell types together with platelet-rich plasma(PRP)as a growth factor reservoir within ectopic bone tissue engineering constructs.AIM To vascularize and examine osteogenesis in bone tissue engineering constructs enriched with PRP and adipose-derived stem cells(ASCs)induced into ECs and OBs.METHODS ASCs isolated from adipose tissue,induced in vitro into ECs,OBs or just expanded were used for implant construction as followed:BPEO,endothelial and osteogenic differentiated ASCs with PRP and bone mineral matrix;BPUI,uninduced ASCs with PRP and bone mineral matrix;BC(control),only bone mineral matrix.At 1,2,4 and 8 wk after subcutaneous implantation in mice,implants were extracted and endothelial-related and bone-related gene expression were analyzed,while histological analyses were performed after 2 and 8 wk.RESULTS The percentage of vascularization was significantly higher in BC compared to BPUI and BPEO constructs 2 and 8 wk after implantation.BC had the lowest endothelial-related gene expression,weaker osteocalcin immunoexpression and Spp1 expression compared to BPUI and BPEO.Endothelial-related gene expression and osteocalcin immunoexpression were higher in BPUI compared to BC and BPEO.BPEO had a higher percentage of vascularization compared to BPUI and the highest CD31 immunoexpression among examined constructs.Except Vwf,endothelial-related gene expression in BPEO had a later onset and was upregulated and well-balanced during in vivo incubation that induced late onset of Spp1 expression and pronounced osteocalcin immunoexpression at 2 and 8 wk.Tissue regression was noticed in BPEO constructs after 8 wk.CONCLUSION Ectopically implanted BPEO constructs had a favorable impact on vascularization and osteogenesis,but tissue regression imposed the need for discovering a more optimal EC/OB ratio prior to considerations for clinical applications.

Ectopic osteogenesis and scaffold biodegradation of tissue engineering bone composed of chitosan and osteo-induced bone marrow mesenchymal stem cells in vivo

Background Chitosan （CS） scaffolds combined with osteogenically induced bone marrow mesenchymal stem cells （BMSCs） have been proved to be promising substitutes for repairing bone defects.Nevertheless,the bone-forming and scaffold-biodegrading processes are seldom studied.This study aimed to determine the osteogenic ability of CS/osteoinduced BMSC composites by observing the bone-forming process and explore the relationship between bone formation and scaffold biodegradation.Methods The CS/osteo-induced BMSC composites （CS＋cells group） and the CS scaffolds （CS group） were,respectively,implanted into SD rat thigh muscles.At 2,4,6,8,and 12 weeks postoperatively,the rat femurs were scanned by CT,and the CT values of the implants were measured and comparatively analyzed.Subsequently,the implants were harvested and stained with hematoxylin and eosin and Masson trichrome,and the percentages of bone area,scaffold area,and collagen area were calculated and compared between the two groups.Results The imaging results showed that the densities of implants of the two groups gradually increased along with time,but the CT values of implants in the CS＋cells group were much higher than in the CS group at the same time point （P ＜0.05）.The histological results showed that the de novo bone and collagen formed in the pores of the scaffolds and gradually increased since 2 weeks postoperation in both groups,and the scaffold gradually degraded along with the boneforming process.However,the comparative analysis results showed that the CS＋cells group gained more de novo bone and collagen formation and had less scaffold than the CS group at the same time point （P ＜0.05）.Conclusion The CS/osteo-induced BMSC composites are excellent bone tissue engineering substitutes,and the scaffold biodegradation is accordant with the bone formation.

Effect of covalent-binding modes of osteogenic-related peptides with artificial carriers on their biological activities in vivo

Covalent binding between bioactive substances and materials in different ways can significantly improve the bone inductivity and biological activity of bone repair materials.However,there is a lack of systematic understanding of how these binding modes affect biological activities of the active substances.In this study,four kinds of functionalized Multi-walled carbon nanotubes(MWCNTs)were prepared,ensuring the same grafting rate of different functional groups.Subsequently,two kinds of osteogenic-related peptides,bone morphogenetic protein-2 mimicking peptides and osteogenic growth mimicking peptides,were covalently bound to functionalized MWCNTs,ensuring the same molar mass of peptides bound to different functionalized MWCNTs in this process.Then the same amount of functionalized MWC-NTs/Peptides composites were introduced into the scaffolds,and through the ectopic osteogenesis model in rats and calvarial defect model in rabbits,ectopic osteogenesis and bone repair ability of the composites were analyzed.Furthermore,the effects of different covalent binding modes on peptide-induced osteogenesis and bone repair were studied.The results showed that the negative influencing trend of different covalent binding modes of osteogenic-related peptides with artificial carriers on their biological activities was in the order as follows:amide binding(carboxyl)>silane coupling>dopamine bind-ing>amide binding(amino),whose mechanism might be mainly that the covalent binding of peptides with different functional groups resulted in different charges.We believe that the results of this study have important guiding significance for the research and development of bone repair materials covalently bound with bioactive substances.