Dental pulp stem cells (DPSCs) which are derived from the neural crest are promising tools in stem cell therapy in the clinic. However, they tend to lose their multi-lineage potency spontaneously during long-term expansion in vitro. A micropattern-based system is developed to train DPSCs with 1024 & mu;m(2) and aspect ratios of 1:2 and 1:4, which enhances their stemness and differentiation potential. This mechanical memory induced by micropattern would also rescue the pluripotency of long-term expansion DPSCs. Micropatterns remodel the cytoskeleton and nuclear morphology, soften the cells, and induce the mitochondria into an immature state. Micropattern also activates extracellular related kinase, signal transducer, and activator of transcription 3, and inhibits Yes-associated protein nuclear localization to enhance the stemness of DPSCs. In vivo implantation proves that DPSCs mechanically trained by micropattern could promote nerve regeneration in rats with peripheral nerve injury. A micropattern-based approach is provided as a stem cell gym that absents exogenous growth factors to enhance the stemness and regenerative properties of DPSCs. It may shed light on future stem cell therapy.