Development of a Surface Plasmon Resonance Biosensor for Real-Time Detection of Osteogenic Differentiation in Live Mesenchymal Stem Cells

Surface plasmon resonance (SPR) biosensors have been recognized as a useful tool and widely used for real-time dynamic analysis of molecular binding affinity because of its high sensitivity to the change of the refractive index of tested objects. The conventional methods in molecular biology to evaluate cell differentiation require cell lysis or fixation, which make investigation in live cells difficult. In addition, a certain amount of cells are needed in order to obtain adequate protein or messenger ribonucleic acid for various assays. To overcome this limitation, we developed a unique SPR-based biosensing apparatus for real-time detection of cell differentiation in live cells according to the differences of optical properties of the cell surface caused by specific antigen-antibody binding. In this study, we reported the application of this SPR-based system to evaluate the osteogenic differentiation of mesenchymal stem cells (MSCs). OB-cadherin expression, which is up-regulated during osteogenic differentiation, was targeted under our SPR system by conjugating antibodies against OB-cadherin on the surface of the object. A linear relationship between the duration of osteogenic induction and the difference in refractive angle shift with very high correlation coefficient was observed. To sum up, the SPR system and the protocol reported in this study can rapidly and accurately define osteogenic maturation of MSCs in a live cell and label-free manner with no need of cell breakage. This SPR biosensor will facilitate future advances in a vast array of fields in biomedical research and medical diagnosis

Innovative Scaffold Solution for Bone Regeneration Made of Beta-Tricalcium Phosphate Granules, Autologous Fibrin Fold, and Peripheral Blood Stem Cells

The drawbacks of traditional bone defect treatments have prompted the exploration of bone tissue engineering. The use of porous biomaterial scaffolds from calcium, bio-ceramic, and other different polymers to induce and increase bone cell and tissue growth is a present hot topic. In bone transplantation, the use of biomaterials may be a solution to avoid the lack of donor sites for autografts and the risk of rejection with allograft procedures. Challenges and efforts involve the use of engineered biomaterials that can mimic both the mechanical and biological properties of real bone tissue, supporting the vascularization of the implanted site. β-Tricalcium phosphate (β-TCP) has been used by dentists and clinicians for a decade in clinical applications on over a thousand patients with different bone pathologies including mandibular and maxillary reconstruction. This study aimed to explore suitable combination of β-TCP granules, autologous fibrin from human peripheral blood (hPB), and autologous peripheral blood stem cells (PB-SCs) for the realization of a bioscaffold (Compact Bio-BoneR) for bone regeneration and identify an efficient method to establish it as effective osteo-regenerators. It has been assessed that human PB is an exceptional source of multiple type of stem cells including mesenchymal (MSCs), neural (NSCs), hematopoietic (HSCs), and embryonic like (ESCs) which may differentiate into different cell phenotypes such as osteoblasts, chondrocytes, adipocytes, myocytes, cardiomyocytes, and neurons. Isolated PB-SCs were induced into osteoblasts using β-TCP granules. Cultured PB-SCs were directly transferred and seeded into the scaffolds and induced to differentiate into osteoblasts. β-TCP granules with diameters of 1 mm and 1–2.5 mm were embedded in a fibrin gel matrix and PB-SCs were added successively. The bioscaffold was poured in culture with serum-free medium (SFM) for a period of 7–10 days. Improved proliferation of PBSCs was assessed by the expression of multipotent and pluripotent stem cell biomarkers performed by flow cytometry analysis as CD34, CD45, CD90, CD105, and SSEA3; osteoblasts were assessed by the positive expression of immune stain as alizarin red (AR), von Kossa (VK), and alkaline phosphatase (ALP). This study provides an alternative to biofunctionalized scaffold that exhibits improved osteogenesis that can be extremely beneficial in dentistry and orthopedics