Quantitative analysis of factors influencing tissue-engineered bone formation by detecting the expression levels of alkaline phosphatase and bone γ-carboxyglutamate protein 2

Abstract

Bone tissue engineering is a promising alternative approach that permits the efficient reconstruction of bone defects. There are four elements involved in bone tissue engineering technology, including the seed cells, growth factors, scaffolds and culture environment. The aim of the present study was to evaluate the effect of these factors on bone formation in tissue engineering technology by analyzing the expression of osteogenetic markers using polymerase chain reaction (PCR). Bone marrow mesenchymal stem cells (BMSCs) were extracted from the bone marrow of the bilateral tibial platform of New Zealand white rabbits. In addition, platelet-rich plasma (PRP) samples were prepared from blood extracted from the ear vein of the rabbits. A perfusion bioreactor was used to provide the culture environment, and β-tricalcium phosphate (β-TCP) was used to build the scaffolds. The β-TCP scaffolds were divided into five groups and each group was treated with a different combination of the factors. Next, the composites were implanted into the rabbits. After three months, the expression levels of the new bone formation markers, alkaline phosphatase and bone γ-carboxyglutamate protein 2, were detected using quantitative reverse transcription-PCR analysis. The expression levels of the markers in the experimental groups were higher compared with the negative control group. Comparisons between the experimental groups also revealed statistical significance. Scanning electron microscopy revealed good adhesion and distribution of the BMSCs on the β-TCP scaffold. In conclusion, the PCR results indicated that PRP, BMSCs and the bioreactor exhibited a promoting effect on bone formation.