Autologous platelet-rich plasma induces bone formation of tissue-engineered bone with bone marrow mesenchymal stem cells on beta-tricalcium phosphate ceramics

Abstract

Background: The purpose of the study is to investigate whether autologous platelet-rich plasma (PRP) can serve as bone-inducing factors to provide osteoinduction and improve bone regeneration for tissue-engineered bones fabricated with bone marrow mesenchymal stem cells (MSCs) and beta-tricalcium phosphate (β-TCP) ceramics. The current study will give more insight into the contradictory osteogenic capacity of PRP.

Methods: The concentration of platelets, platelet-derived growth factor-AB (PDGF-AB), and transforming growth factor-β1 (TGF-β1) were measured in PRP and whole blood. Tissue-engineered bones using MSCs on β-TCP scaffolds in combination with autologous PRP were fabricated (PRP group). Controls were established without the use of autologous PRP (non-PRP group). In vitro, the proliferation and osteogenic differentiation of MSCs on fabricated constructs from six rabbits were evaluated with MTT assay, alkaline phosphatase (ALP) activity, and osteocalcin (OC) content measurement after 1, 7, and 14 days of culture. For in vivo study, the segmental defects of radial diaphyses of 12 rabbits from each group were repaired by fabricated constructs. Bone-forming capacity of the implanted constructs was determined by radiographic and histological analysis at 4 and 8 weeks postoperatively.

Results: PRP produced significantly higher concentration of platelets, PDGF-AB, and TGF-β1 than whole blood. In vitro study, MTT assay demonstrated that the MSCs in the presence of autologous PRP exhibited excellent proliferation at each time point. The results of osteogenic capacity detection showed significantly higher levels of synthesis of ALP and OC by the MSCs in combination with autologous PRP after 7 and 14 days of culture. In vivo study, radiographic observation showed that the PRP group produced significantly higher score than the non-PRP group at each time point. For histological evaluation, significantly higher volume of regenerated bone was found in the PRP group when compared with the non-PRP group at each time point.

Conclusions: Our study findings support the osteogenic capacity of autologous PRP. The results indicate that the use of autologous PRP is a simple and effective way to provide osteoinduction and improve bone regeneration for tissue-engineered bone reconstruction.

Keywords: Autologous; Beta-tricalcium phosphate; Osteogenic; Platelet-rich plasma; Scaffold; Tissue-engineered bone.

Fig. 1

Cell proliferation in vitro. The PRP group demonstrated significantly higher absorbance values compared with the non-PRP group at each time point. (Data in mean ± SD, n = 6, *p < 0.05)

Fig. 2

ALP activity for cell osteogenic differentiation in vitro. Significantly higher ALP activity was detected for the PRP group when compared to that for the non-PRP group at 7 and 14 days after seeding. (Data in mean ± SD, n = 6, *p < 0.05)

Fig. 3

OC content for cell osteogenic differentiation in vitro. The PRP group exhibited significantly higher OC content than the non-PRP group on days 7 and 14 after being cultured. (Data in mean ± SD, n = 6, *p < 0.05)

Fig. 4

Representative radiographs of critical-sized bone defects repair by PRP group constructs (a, c) and Non-PRP group constructs (b, d) at 4 weeks (a, b) and 8 weeks (c, d) postoperatively. At 4 weeks postoperatively, the radioopaque areas of implants and radiolucent zone at the interface between the implant and host bone was visible in both groups. However, the boundary of constructs in the PRP group became more cloudy. At 8 weeks postoperatively, a decrease in radiopacity related to the new bone formation and material degradation was more obvious in the PRP group. The radiolucent zone at the interfacial area almost disappeared in the PRP group. The arrow indicates the implanted construct

Fig. 5

Histological evaluation of regenerated bone of PRP group constructs (a, c) and non-PRP group constructs (b, d) at 4 weeks (a, b) and 8 weeks (c, d) postoperatively. The PRP group demonstrated more extensive bone formation with the degradation of implanted constructs than the non-PRP group at each time point. (HE staining × 100. TCP, tricalcium phosphate; TB, tissue-engineered bone)

Fig. 6

Histological evaluation of interfacial area between the implant and the host bone of PRP group constructs (a, c) and non-PRP group constructs (b, d) at 4 weeks (a, b) and 8 weeks (c, d) postoperatively. The newly formed bone grew and merged at interfacial area for both groups with implantation period prolonging. However, the amount and rate of the new bone formation of the non-PRP group was less and slower than that of PRP group (HE staining; × 40. TCP, tricalcium phosphate; TB, tissue-engineered bone; NB, native bone; MC, medullary cavity. The dotted rectangles indicate the interfacial area)