Zhu Zhenyu, Kong Fanqi, Jiang Feng, Jiang Jialin, Quan Danni, Guo Jiazheng, Sun Kaiqiang, Shi Jiangang, Wang Changnan, Zhuang Chunlin, Xu Ximing
Department of Orthopedic Surgery, Changzheng Hospital, Navy Medical University, No. 415 Fengyang Rd, Shanghai, 200003, People's Republic of China.
School of Materials and Chemistry, University of Shanghai for Science and Technology, Shanghai, 200093, People's Republic of China.
Apoptosis. 2025 Sep 16. doi: 10.1007/s10495-025-02169-y.
Intervertebral disc degeneration (IVDD) is a major contributor to lumbar diseases, including low back pain, herniation, and stenosis. Despite significant efforts, there have been limited improvements in treatments to alleviate IVDD. The nucleus pulposus (NP) is a crucial component of the intervertebral disc (IVD), responsible for secreting aggrecan, collagen II, and other extracellular matrix components. Programmed cell death (PCD) of NP cells is believed to play a central role in IVDD. RIPK1 is a key mediator of PCD and recently reported PANoptosis, playing essential role in kidney injury, arteriosclerosis, and acute or chronic inflammation-related diseases. We collected varied degenerated human IVD specimens to examine the expression of RIPK1 and downstream cell death-related markers, including GSDMD, Caspase3, and MLKL, which are indicative of pyroptosis, apoptosis, necroptosis, or the recently denominated PANoptosis. In vitro, we performed RIPK1 knockdown and overexpression to study their effects on IVDD. in vivo, we constructed RIPK1 conditional knockout (CKO) mice to confirm the role of RIPK1 in IVDD. We also utilized a small molecule targeted inhibitor to explore its effects on IVDD in vitro and in vivo. Phosphorylated RIPK1 (p-RIPK1) was significantly increased during IVDD in both human and mouse models. Knockout of RIPK1 effectively alleviated IVDD, as evidenced by the RIPK1 cko mice. Further pathological staining and western blot analysis revealed the overexpression of GSDMD, Caspase3, and MLKL, indicating that RIPK1-mediated PANoptosis plays a crucial role in IVDD. in vitro, overexpression of RIPK1 in NP cells exacerbated PANoptosis and degeneration, while RIPK1 knockdown inhibited these processes. We developed a RIPK1-targeted small molecular inhibitor, compound 3-47, which demonstrated superior efficacy in inhibiting p-RIPK1. Both in vitro and in vivo, 3-47 showed remarkable effects in alleviating IVDD by inhibiting RIPK1-mediated PANoptosis. RIPK1-mediated PANoptosis of NP cells plays a critical role in IVDD. The molecular inhibitor 3-47 could effectively delay IVDD progression in mice, highlighting its therapeutic potential.
椎间盘退变（IVDD）是腰椎疾病的主要原因，包括腰痛、椎间盘突出和椎管狭窄。尽管付出了巨大努力，但缓解IVDD的治疗方法进展有限。髓核（NP）是椎间盘（IVD）的关键组成部分，负责分泌聚集蛋白聚糖、胶原蛋白II和其他细胞外基质成分。NP细胞的程序性细胞死亡（PCD）被认为在IVDD中起核心作用。RIPK1是PCD的关键介质，最近报道其与PANoptosis有关，在肾损伤、动脉硬化以及急性或慢性炎症相关疾病中起重要作用。我们收集了各种退变的人类IVD标本，以检测RIPK1及下游细胞死亡相关标志物的表达，包括GSDMD、Caspase3和MLKL，这些标志物分别指示焦亡、凋亡、坏死性凋亡或最近命名的PANoptosis。在体外，我们进行了RIPK1基因敲低和过表达实验，以研究它们对IVDD的影响。在体内，我们构建了RIPK1条件性敲除（CKO）小鼠，以确认RIPK1在IVDD中的作用。我们还利用一种小分子靶向抑制剂，探索其在体外和体内对IVDD的影响。在人类和小鼠模型的IVDD过程中，磷酸化RIPK1（p-RIPK1）均显著增加。如RIPK1 CKO小鼠所示，敲除RIPK1可有效缓解IVDD。进一步的病理染色和蛋白质印迹分析显示GSDMD、Caspase3和MLKL过表达，表明RIPK1介导的PANoptosis在IVDD中起关键作用。在体外，NP细胞中RIPK1的过表达加剧了PANoptosis和退变，而RIPK1基因敲低则抑制了这些过程。我们开发了一种靶向RIPK1的小分子抑制剂化合物3-47，它在抑制p-RIPK1方面表现出卓越的效果。在体外和体内，3-47通过抑制RIPK1介导的PANoptosis，在缓解IVDD方面均显示出显著效果。NP细胞中RIPK1介导的PANoptosis在IVDD中起关键作用。分子抑制剂3-47可有效延缓小鼠IVDD的进展，突出了其治疗潜力。
