髓核细胞老化及老化表型改变对局部微环境的旁分泌效应研究

髓核细胞在维持椎间盘内环境稳定中发挥着核心作用，而细胞老化及老化后的表型改变可潜在导致髓核细胞数量减少和功能异常。本研究通过体外连续培养传代、氧化应激干预人正常髓核细胞，分别建立复制性老化和应激诱导的过早老化模型；探索P53-p21-Rb和p16INK4A-Rb通路在髓核细胞老化过程中的信号介导与交通；比较老化髓核细胞在炎症因子、生长因子、基质降解酶、细胞外基质合成四大方面的表型改变；观察老化髓核细胞旁分泌作用于正常髓核细胞后的致退变效应。从"细胞启动老化信号通路→形成复制性老化或过早老化→表达老化分泌表型→恶化邻近细胞生存微环境"角度认识髓核细胞老化及其表型改变在椎间盘退行性变中的病因学作用，寻找髓核细胞老化的关键调控信号，为生物学治疗更具针对性地选择抗老化靶点来有效修复椎间盘退变打开崭新思路。

Normal nucelus pulposus (NP) cells are essential for the maintance of homeostasis within intertebral disc(IVD). As NP cells gradually or prematurely become senescent and change to demonstrate senescent phenotypes, a significant loss of NP cells and their biological functions is expected to occur. In this study, replicative senescence and premature senescence are to be induced in human NP cells by serial subculturing and exposure to oxidative stresses respectively. The activation and crosstalk between the two senescent pathways, p53-p21-Rb and p16INK4A-Rb, will be detected and analyzed in senescent NP cells. By comparing the ability in producing inflammative cytokines, growth factors, matrix degrading enzymes, as well as the extracellular matrix between senescent and non-senescent NP cells, a comprehensive study of the senescent phenotypes of NP cells is made. Besides, normal NP cells will be co-cultured with senescent NP cells to observe the paracrine actions from the phenotype changes of senescent NP cells. Our studies aim to illuminate the role of NP cell senescence and the senescent phenotype during IVD degeneration. It is probable that aging and multiple stresses activate the p53-p21-pRB and/or p16INK4A-pRB senescent pathways to induce NP cell senescence, then the accumulation of senescent cells potentially deteriorates the disc microenvironment, which triggers and accelerates the IVD degeneration. By identifying the key element in the transduction of senescent pathways, our studies also support the feasiblity of anti-senescent approaches for IVD regeneration.