The idea that somatic stem cells are localized in periodontal ligament (PDL) tissues as PDL stem cells (PDLSCs) responsible for construction and reconstruction of the periodontium has been widely accepted. Many dental scientists have attempted to clarify the identity of these PDLSCs, but the number of PDLSCs localized in PDL tissues is too small to be routinely and conveniently analyzed. Therefore, researchers have been attempting to develop undifferentiated PDL cell lines by transducing them with genes that are suitable for immortalization. The present authors were the first to succeed in establishing two clonal human PDL stem/progenitor cell lines that possessed multipotency derived from PDL tissues and that expressed PDL-related molecules as well as neural crest- and embryonic stem-related markers. The differentiation stages of these cell lines appeared to vary based on their potential to differentiate into other lineage cells, their response to tissue regeneration-related cytokines, and their behavior when transplanted into immunodeficient rats. This review describes the phenotypes of these cell lines compared with reported PDLSCs or other MSCs and discusses contemporary circumstances related to PDL regenerative medicine. Differential analyses between these two clones will reveal the mechanism of differentiation of PDLSCs as well as their phenotypes. The results will also allow for the acquisition of a mass population of PDLSCs or other stem cells directed toward PDL-lineage cells and to develop an unmet treatment needed for construction and reconstruction of PDL tissues based on tissue engineering techniques.