Periodontal ligament cell (PDLC) sheets have been shown to contribute to periodontal tissue regeneration. Dental follicle cells (DFCs), acknowledged as the precursor cells of PDLCs, have demonstrated stemness, embryonic features, heterogeneity, and pluripotency. Therefore, we hypothesized that DFC sheets might be more effective and suitable for periodontal tissue regeneration than PDLC sheets. In this study, we compared the biological characteristics of DFC sheets and PDLC sheets in vitro. To investigate the potential for periodontal tissue regeneration in vivo, complexes composed of two types of cell sheets combined with dentin matrix were implanted subcutaneously into nude mice for 6 weeks. Our results showed that, when forming cell sheets, DFCs secreted richer extracellular matrix than PDLCs. And compared to DFCs, DFC sheets expressed high levels of calcification-related genes, including alkaline phosphatase (alp), bone sialoprotein (bsp), osteopontin (opn), runt-related transcription factor (runx2), as well as the periodontal ligament-specific genes collagen III (col III) and periostin, while the gene expression of bsp, osteocalcin (ocn), and opn were greatly increased in PDLC sheets, when compared to PDLCs. col I expression did not change significantly. However, cementum protein 23 (cp-23) expression increased several fold in PDLC sheets compared to PDLCs but decreased in DFC sheets compared to DFCs. DFC and PDLC sheets were both positive for Collagen I (Col I), cementum attachment protein (CAP), ALP, BSP, OCN, and OPN protein expression, and Col I, ALP, BSP, and OPN expression were increased after cell sheets were formed. Furthermore, the levels of laminin and fibronectin were higher in DFCs and DFC sheets than that of PDLCs and PDLC sheets, respectively. In vivo, DFC and PDLC sheets could both regenerate periodontal tissue-like structures, but DFC sheets demonstrated stronger periodontal regeneration potential than PDLC sheets. Therefore, DFC sheets derived from discarded dental follicle tissue after tooth extraction may be more advantageous for clinical periodontal tissue regeneration in the future.