Smart Surface-Enhanced Resonance Raman Scattering Nanoprobe for Monitoring Cellular Alkaline Phosphatase Activity during Osteogenic Differentiation.

High-efficiency induction of bone marrow mesenchymal stem cells (BMSCs) to osteogenic differentiation in vitro can help solve a series of bone diseases such as bone injury, fracture repair, and osteoporosis. In order to explore the optimal conditions for different chemical inducers to promote BMSCs differentiation and the possible differentiation mechanisms, we developed a smart nanoprobe that can achieve in situ alkaline phosphatase (ALP) activity detection during osteogenic differentiation in cells. The smart nanoprobe (Au@BCIP) was designed as the surface decoration of gold nanoparticles (AuNPs) with 5-bromo-4-chloro-3-indolyl phosphate (BCIP). The nanoprobe was co-cultured with differentiated BMSCs at different stages to monitor ALP activity based on an ALP-catalyzed hydrolysis reaction with BCIP as a substrate. The product can be quickly oxidized by dissolved oxygen to obtain a Raman-active species (5,5'-dibromo-4,4'-dichloro-1H,1H-[2,2'] biindolylidene-3,3'-dione). The SERS sensitivity was greatly improved by resonating the excitation wavelength of 632.8 nm. It is a new strategy for tracing bone disease-related ALP activity in an in vivo model with high sensitivity and selectivity and non-invasion. By using this nanoprobe, osteogenic differentiation of cells under osteogenic supplements was assessed and the p38 MAPK signaling pathway for osteogenic differentiation was experimentally evidenced, which are of significance for understanding BMSCs and regulating their osteogenic differentiation process.