In Vitro Evaluation of Recombinant Bone Morphogenetic Protein-2 Bioactivity for Regenerative Medicine

• Published in: Tissue engineering. Part C, Methods Vol. 25; no. 9; pp. 553 - 559
• Main Authors: Fung, Stephanie L., Wu, Xiaohuan, Maceren, Julian P., Mao, Yong, Kohn, Joachim
• Format: Journal Article
• Language: English
• Published: United States Mary Ann Liebert, Inc., publishers 01.09.2019
• Subjects: Animals; Biological Assay; Bone Morphogenetic Protein 2 - biosynthesis; Bone Morphogenetic Protein 2 - chemistry; Bone Morphogenetic Protein 2 - genetics; Bone Morphogenetic Protein 2 - pharmacology; Cell Line; Drug Evaluation; Humans; Methods Articles; Mice; Protein Stability; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - pharmacology; Regenerative Medicine; bone regeneration; alkaline phosphatase activity; bioactivity; BMP-2
• ISSN: 1937-3384; 1937-3392
• DOI: 10.1089/ten.tec.2019.0156

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is a commonly used growth factor in bone regeneration due to its high potency and ability to induce osteogenic differentiation of osteoblasts and osteoblast precursors. When designing delivery systems for rhBMP-2, the activity of the loaded and released protein is an important consideration. The variability in the experimental design parameters used to measure rhBMP-2 activity in vitro has precluded comparative analysis. Here, for the first time, we report a direct comparison of the assay parameters used in rhBMP-2 bioactivity assays in the literature and an evaluation of commercially available rhBMP-2 obtained from different vendors. Most published rhBMP-2 assays use W-20-17 (mouse stromal), MC3T3 (preosteoblast), or C2C12 (myoblast) cell lines. We found that each model cell line has an optimal concentration range over which it is most sensitive to rhBMP-2 induction. Therefore, it is difficult to find one single bioassay protocol that could be universally used. In addition, we established a correlation between protein concentration (as measured by enzyme-linked immunosorbent assay) and protein activity (as measured by alkaline phosphatase induction). We found that the expression system used to produce the rhBMP-2 had the greatest effect on its activity and stability in vitro . Establishing a standard method of measuring rhBMP-2 activity in vitro is the first step toward developing an in vitro–in vivo correlation between measured activity and clinical outcomes.