Palmitoylethanolamine alleviates osteoclast activation and reduces bone loss in osteoporosis by regulating mitochondrial function.

Abstract

Osteoclasts are responsible for bone resorption, and their excessive activation causes bone loss and structural damage in osteoporosis. Palmitoylethanolamide (PEA), an endogenous PPAR-α agonist, exerts anti-inflammatory effects, but its role in osteoporosis remains incompletely understood. Primary bone marrow-derived macrophages (BMMs) were isolated, and the cytotoxicity of PEA was assessed via CCK-8 analysis. Osteoclast differentiation of BMMs was determined by TRAP staining, osteoclast markers (NFATc1, CTSK, TRAP) and F-actin ring staining. Mitochondrial function was evaluated by MitoSOX, JC-1, ATP and NAD⁺/NADH ratio. Molecular interactions were validated with chromatin immunoprecipitation, dual luciferase assay and RNA immunoprecipitation. Micro-CT and histological staining analyses were performed to evaluate bone loss in ovariectomized mice. PEA dose-dependently suppressed RANKL-induced osteoclast differentiation, F-actin ring formation, and osteoclast marker gene expression. PEA suppressed mitochondrial function (reduced mitochondrial membrane potential, mitochondrial ROS, ATP, NAD⁺/NADH) in RANKL-induced BMMs. However, activation of NF-κB signaling or blockade of PPAR-α dramatically reversed these effects of PEA. Mechanistically, PEA promoted YTHDC1 transcription in a PPAR-α/RXRA-dependent manner. Moreover, YTHDC1 bound to m6A-modified RELA mRNA, leading to its nuclear export and recognition by YTHDF2, which in turn led to its degradation and consequent inactivation of NF-κB signaling. Rescue experiments demonstrated that YTHDC1 knockdown reversed PEA-mediated suppression of osteoclast differentiation and mitochondrial function. PEA impairs mitochondrial function in RANKL-induced BMMs, thereby repressing osteoclast differentiation via the YTHDC1/m6A-RELA/NF-κB signaling axis, which provides a novel molecular mechanism of PEA in osteoporosis treatment.