Xie Tian, Gu Xi, Pan Ruijie, Huang Wenzhuo, Dong Sheng
Department of Orthopedics, Wuhan Hospital of Traditional Chinese Medicine, No. 49 Lihuangpi Road, Jiang'an District, Wuhan, 430014 China.
College of Acupuncture and Bone Injury, Hubei University of Chinese Medicine, Wuhan, 430061 China.
Cytotechnology. 2024 Apr;76(2):153-166. doi: 10.1007/s10616-023-00605-y. Epub 2023 Dec 27.
Degradation of extracellular matrix (ECM), reactive oxygen species (ROS) production, and inflammation are critical players in the pathogenesis of intervertebral disc degeneration (IDD). Evodiamine exerts functions in inhibiting inflammation and maintaining mitochondrial antioxidant functions. However, the biological functions of evodiamine and its related mechanisms in IDD progression remain unknown. The IDD-like conditions in vivo were stimulated via needle puncture. Hematoxylin and eosin staining, Safranin O/Fast Green staining and Alcian staining were performed to determine the degenerative status. The primary nucleus pulposus cells (NPCs) were isolated from Sprague-Dawley rats and then treated with tert-butyl peroxide (TBHP) to induce cellular senescence and oxidative stress. The cell viability was assessed by cell counting kit-8 assays. The mitochondria-derived ROS in NPCs was evaluated by MitoSOX staining. The mitochondrial membrane potential in NPCs was identified by JC-1 staining and flow cytometry. The expression of collagen II in NPCs was measured by immunofluorescence staining. The levels of mRNAs and proteins were measured by RT-qPCR and western blotting. The Nrf2 expression in rat nucleus pulposus tissues was measured by immunohistochemistry staining. Evodiamine alleviated TBHP-induced mitochondrial dysfunctions in NPCs. The enhancing effect of TBHP on the ECM degradation was reversed by evodiamine. The TBHP-stimulated inflammatory response was ameliorated by evodiamine. Evodiamine alleviated the IDD process in the puncture-induced rat model. Evodiamine promoted the activation of Nrf2 pathway and inactivated the MAPK pathway in NPCs. In conclusion, evodiamine ameliorates the progression of IDD by inhibiting mitochondrial dysfunctions, ECM degradation and inflammation via the Nrf2/HO-1 and MAPK pathways.
细胞外基质(ECM)降解、活性氧(ROS)生成和炎症是椎间盘退变(IDD)发病机制中的关键因素。吴茱萸碱具有抑制炎症和维持线粒体抗氧化功能的作用。然而,吴茱萸碱在IDD进展中的生物学功能及其相关机制尚不清楚。通过针刺刺激建立体内类似IDD的状态。进行苏木精-伊红染色、番红O/固绿染色和阿尔辛蓝染色以确定退变状态。从Sprague-Dawley大鼠分离原代髓核细胞(NPCs),然后用叔丁基过氧化氢(TBHP)处理以诱导细胞衰老和氧化应激。通过细胞计数试剂盒-8检测评估细胞活力。通过MitoSOX染色评估NPCs中线粒体来源的ROS。通过JC-1染色和流式细胞术鉴定NPCs中的线粒体膜电位。通过免疫荧光染色测量NPCs中Ⅱ型胶原蛋白的表达。通过RT-qPCR和蛋白质印迹法测量mRNA和蛋白质水平。通过免疫组织化学染色测量大鼠髓核组织中Nrf2的表达。吴茱萸碱减轻了TBHP诱导的NPCs线粒体功能障碍。吴茱萸碱逆转了TBHP对ECM降解的增强作用。吴茱萸碱改善了TBHP刺激的炎症反应。吴茱萸碱减轻了穿刺诱导的大鼠模型中的IDD进程。吴茱萸碱促进了NPCs中Nrf2通路的激活并使MAPK通路失活。总之,吴茱萸碱通过Nrf2/HO-1和MAPK通路抑制线粒体功能障碍、ECM降解和炎症,从而改善IDD的进展。