<p>Osteoporosis, characterized by diminished bone density and compromised microstructure, presents a significant healthcare challenge, particularly in the aging population. The primary approach in addressing osteoporosis involves the use of anti-resorptive agents as well as medications that promote bone formation. However, these therapies have limitations, prompting the exploration of novel therapeutic targets. Prolyl Endopeptidase (PREP), an endopeptidase with diverse roles in neuronal peptide metabolism and various physiological processes, has emerged as a potential player in osteoporosis, though its mechanistic involvement remains largely uncharted. This study delves into the role of PREP in osteoporosis, aiming to unravel its underlying mechanisms and therapeutic potential. Utilizing murine models and cellular experiments, we systematically investigate how PREP influences osteoblast differentiation and osteoporosis pathogenesis. Our results suggest activation of the Wnt pathway counteracts the inhibitory effects of PREP deletion on osteoblast differentiation. Additionally, we observe that PREP affects osteoclastogenesis, influencing osteoclast differentiation and bone resorption capacity. Moreover, our investigation extends to age-related osteoporosis, demonstrating PREP’s potential therapeutic efficacy beyond estrogen-deficiency-induced osteoporosis. In summary, this study advances our understanding of PREP’s multifaceted role in osteoporosis pathogenesis. It underscores PREP as a potential therapeutic target for osteoporosis, offering fresh perspectives on its etiology and treatment.</p>