Prostaglandin E2 (PGE2), a major cyclooxygenase-2 (COX-2) product, is highly secreted by the osteoblast lineage in the subchondral bone tissue of osteoarthritis (OA) patients. However, NSAIDs, including COX-2 inhibitors, have severe side effects during OA treatment. Therefore, the identification of novel drug targets of PGE2 signaling in OA progression is urgently needed. Osteoclasts play a critical role in subchondral bone homeostasis and OA-related pain. However, the mechanisms by which PGE2 regulates osteoclast function and subsequently subchondral bone homeostasis are largely unknown. Here, we show that PGE2 acts via EP4 receptors on osteoclasts during the progression of OA and OA-related pain. Our data show that while PGE2 mediates migration and osteoclastogenesis via its EP2 and EP4 receptors, tissue-specific knockout of only the EP4 receptor in osteoclasts (EP4^LysM) reduced disease progression and osteophyte formation in a murine model of OA. Furthermore, OA-related pain was alleviated in the EP4^LysM mice, with reduced Netrin-1 secretion and CGRP-positive sensory innervation of the subchondral bone. The expression of platelet-derived growth factor-BB (PDGF-BB) was also lower in the EP4^LysM mice, which resulted in reduced type H blood vessel formation in subchondral bone. Importantly, we identified a novel potent EP4 antagonist, HL-43, which showed in vitro and in vivo effects consistent with those observed in the EP4^LysM mice. Finally, we showed that the Gαs/PI3K/AKT/MAPK signaling pathway is downstream of EP4 activation via PGE2 in osteoclasts. Together, our data demonstrate that PGE2/EP4 signaling in osteoclasts mediates angiogenesis and sensory neuron innervation in subchondral bone, promoting OA progression and pain, and that inhibition of EP4 with HL-43 has therapeutic potential in OA.

前列腺素 E2 (PGE2) 是一种主要的环氧合酶-2 (COX-2) 产物，由骨关节炎 (OA) 患者的软骨下骨组织中的成骨细胞谱系高度分泌。然而，非甾体抗炎药，包括 COX-2 抑制剂，在 OA 治疗期间具有严重的副作用。因此，迫切需要鉴定 OA 进展中 PGE2 信号传导的新药物靶点。破骨细胞在软骨下骨稳态和 OA 相关疼痛中起关键作用。然而，PGE2 调节破骨细胞功能和随后的软骨下骨稳态的机制在很大程度上是未知的。在这里，我们显示 PGE2 在 OA 和 OA 相关疼痛的进展过程中通过破骨细胞上的 EP4 受体起作用。我们的数据表明，虽然 PGE2 通过其 EP2 和 EP4 受体介导迁移和破骨细胞生成，EP4 ^LysM ) 减少了 OA 小鼠模型中的疾病进展和骨赘形成。此外，与 OA 相关的疼痛在EP4 ^LysM小鼠中得到缓解，Netrin-1 分泌减少和软骨下骨的 CGRP 阳性感觉神经支配。EP4 ^LysM小鼠中血小板衍生生长因子-BB (PDGF-BB) 的表达也较低，这导致软骨下骨中 H 型血管的形成减少。重要的是，我们发现了一种新型强效 EP4 拮抗剂 HL-43，其在体外和体内的作用与在EP4 ^{LysM中观察到的一致。}老鼠。最后，我们发现 Gαs/PI3K/AKT/MAPK 信号通路是破骨细胞中通过 PGE2 激活 EP4 的下游。总之，我们的数据表明破骨细胞中的 PGE2/EP4 信号传导介导软骨下骨中的血管生成和感觉神经元神经支配，促进 OA 进展和疼痛，并且用 HL-43 抑制 EP4 在 OA 中具有治疗潜力。