Evaluation of in vitro osteoblast and osteoclast differentiation from stem cell: a systematic review of morphological assays and staining techniques

Abstract

Background. Understanding human stem cell differentiation into osteoblasts and osteoclasts is crucial for bone regeneration and disease modeling. Numerous morphological techniques have been employed to assess this differentiation, but a comprehensive review of their application and effectiveness is lacking. Methods. Guided by the PRISMA framework, we conducted a rigorous search through the PubMed, Web of Science and Scopus databases, analyzing 254 articles. Each article was scrutinized against pre-defined inclusion criteria, yielding a refined selection of 14 studies worthy of in-depth analysis. Results. The trends in using morphological approaches were identified for analyzing osteoblast and osteoclast differentiation. The three most used techniques for osteoblasts were Alizarin Red S (mineralization; six articles), von Kossa (mineralization; three articles) and alkaline phosphatase (ALP; two articles) followed by one article on Giemsa staining (cell morphology) and finally immunochemistry (three articles involved Vinculin, F-actin and Col1 biomarkers). For osteoclasts, tartrate-resistant acid phosphatase (TRAP staining) has the highest number of articles (six articles), followed by two articles on DAPI staining (cell morphology), and immunochemistry (two articles with VNR, Cathepsin K and TROP2. The study involved four stem cell types: peripheral blood monocyte, mesenchymal, dental pulp, and periodontal ligament. Conclusion. This review offers a valuable resource for researchers, with Alizarin Red S and TRAP staining being the most utilized morphological procedures for osteoblasts and osteoclasts, respectively. This understanding provides a foundation for future research in this rapidly changing field.