Ultrasonic-Activated Electrical Stimulation for Regulating Stem Cell Differentiation and Its Molecular Stress Response Monitoring in the Endoplasmic Reticulum with the Plasmonic Piezoelectric SERS Platform

Precise and controllable electrical stimulation (ES) for regulating stem cell differentiation is promising and an emerging means for promoting tissue regeneration and repair. Herein, a wireless controllable ES at the subcellular level is developed to quickly promote the directed differentiation of the dental pulp stem cell (DPSC) to early osteogenesis, based on endoplasmic reticulum (ER)-targeting BaTiO₃@AgNPs (named as ET-BTO@AgNPs) under ultrasound stimulation (US). Ultrasonic-driven electrical stimulation (UES) can greatly shorten the DPSC differentiation time from the usual more than 10 days to only 3 days compared with traditional drug stimulation. The reason might be that ER stress can facilitate the mitochondrial membrane potential boosting during UES, promoting adenosine triphosphate (ATP) generation and, hence, accelerating DPSC differentiation. In the study, the developed ET-BTO@AgNPs played dual roles, which can generate a controllable voltage of about 26 mV under US to promote DPSC differentiation and also act as a superior surface-enhanced Raman spectroscopy (SERS) substrate for dynamic profiling monitoring the molecular stress responses of the ER at the single-cell level. The SERS spectra of the ER within the DPSC were recorded during the differentiation process, which revealed amino acid levels (tyrosine, phenylalanine, and hydroxyproline) associated with differentiation proteins, the contents of which were significantly boosted with extending the differentiation time. Notably, the levels of lipids within the ER were also markedly upregulated during DPSC differentiation. Our developed method has excellent potential for tooth regeneration and repair in future clinical applications.