Periodontal ligament (PDL), connecting alveolar bone and tooth root, is crucial for mechanical stabilization and mastication-related forces absorption. The capacities deteriorate if PDL is affected by periodontitis, which is ranked as the sixth most prevalent disease worldwide, affecting approximately 10% of adults. It is characterized by the progressive degeneration of periodontium, eventually leading to the detachment of teeth from their alveolar cavity. In the field of periodontal research, studies have demonstrated that PDL cells possess the ability to regenerate periodontal tissue when exposed to specific physiological stimuli (e.g., tension, compression, and shear stress) in an ex vivo environment. Indeed, tissue engineering could represent a promising strategy for treating periodontitis; however, due to the PDL complexity, any regeneration strategy should entail studying specific signals that guide the formation and remodeling of the PDL.