EP3 receptor mediates calcium oxalate kidney stone formation via Ca²⁺/AKT and necroinflammation.

Kidney stone disease is a prevalent urological condition with rising incidence and high recurrence rates, yet effective pharmacological interventions remain lacking [1] Prostaglandin E₂ (PGE₂) and its receptor EP3 have been implicated in inflammatory kidney injury [1-3], but their role in nephrolithiasis is unclear. This study investigates the role and mechanism of EP3 in calcium oxalate (CaOx) stone formation using in vitro and in vivo models. EP3 expression was significantly upregulated in CaOx-stimulated human renal tubular epithelial cells (HK-2) and in kidney tissues of glyoxylate-induced nephrolithiasis mice. Treatment with the EP3 antagonist DG041 in vitro reduced crystal adhesion, decreased HMGB1 release, and suppressed IL-6 and TNF-α expression. Mechanistically, EP3 antagonism inhibited aberrant intracellular Ca2⁺ elevation and reduced AKT phosphorylation. Virtual screening identified Astragaloside A as a potential EP3 inhibitor. Astragaloside A recapitulated the protective effects of DG041, attenuating crystal adhesion, inflammation, and necroinflammation both in vitro and in vivo. These findings indicate that EP3 mediates crystal-induced renal tubular injury and inflammation via the Ca2⁺/AKT signaling axis. Targeting EP3 may interrupt the "crystal‑injury‑inflammation" cycle in nephrolithiasis, offering a new therapeutic strategy. This study is the first to demonstrate that the EP3 receptor mediates calcium oxalate kidney stone formation through the Ca2⁺/AKT signaling pathway and necroinflammation. EP3 antagonism reduces crystal adhesion, HMGB1 release, and inflammation. Virtual screening identifies astragaloside A as a novel EP3 inhibitor with protective effects in vitro and in vivo, highlighting EP3 as a promising therapeutic target.