Per2基因调控p53信号通路促人脑胶质母细胞瘤干细胞凋亡的分子机制及放疗敏感性研究

Glioblastoma stem cells (GSCs) are endowed with highly aggressive, self-renewal ability, multi-potent nature and result in glial tumor occurrence and proliferation rapidly. These features of GSCs are attributed to presence of a higher heterogeneity, which are regarded as initial tumorigenic ability glial cells with conventional chemotherapy and radiotherapy resistant and often show poor patient prognosis due to frequent recurrence. We have reported that Per1 and Per2 were expressed with a circadian rhythm in glioma and normal brain tissue in rats and their expression profiles differed in the two types of tissue, Per2 played a key role in regulating the ATM-p53-MDM2 signaling pathway increasing apoptosis of glioma. Therefore, it has become an urgent problem to understand the molecular mechanisms in GSCs, which plays a core role in occurrence and progression of tumor. In this research, we discuss the peak time interval of circadian rhythm in Per2 mediated positive CD133 and p53 signaling pathway increasing the cell-cycle, apoptosis, proliferation and radiosensitivity in human GSCs by x-irradiation. Furthermore, it is important to understand the mechanisms and its key impact on the occurrence and progression of GSCs, and elucidate the molecular mechanisms and core action sites of regulated expression of Per2 in GSCs apoptosis in Human. Finally, our research will contribute to find potential accurate molecular target sites of glioblastoma therapy in human.

胶质母细胞瘤干细胞是具有启动肿瘤快速增殖、侵袭、转移等生物学特性并有完整成瘤能力的一群肿瘤内异质性的初始细胞，对常规放化疗抗拒的特点，从而造成胶质母细胞瘤复发率高、预后差。课题组前期研究已证实Per1、Per2基因在人脑胶质瘤细胞与周围正常组织细胞存在明显不同的生物节律性表达；发现Per2基因通过调控ATM-p53-MDM2信号通路促进胶质瘤细胞凋亡过程中起到关键作用。探索在肿瘤发生、发展中关键作用的肿瘤干细胞分子机制成为迫切研究问题，我们拟研究Per2基因介导CD133阳性人脑胶质母细胞瘤干细胞生物节律表达峰值时段，在射线干预下调控通过p53基因信号通路促进胶质母细胞瘤干细胞的生长、凋亡机制及放射敏感性作用，更好的认识人脑胶质母细胞瘤干细胞产生肿瘤的本质和关键作用，阐明Per2 基因调控胶质母细胞瘤干细胞凋亡分子机制及核心位点作用，寻找人脑胶质母细胞瘤真正有效的潜在精准分子治疗靶点。

研究背景：胶质瘤干细胞（GSCs）启动肿瘤快速增殖、侵袭、转移、抗拒治疗，可能是胶质瘤发生、复发和预后差的根源。Per2基因和肿瘤干细胞的生物学行为密切相关，但和胶质瘤干细胞的相关性机制研究甚少。本项目旨在研究、阐明Per2基因调控GSCs凋亡的作用及机制，寻找胶质瘤治疗的有效靶点。.主要研究内容：（1）对U87胶质瘤细胞系分离、培养获得胶质瘤干细胞干细胞并行干性鉴定，同时研究干细胞的Per2节律特点；（2）研究Per2基因过表达对U87，U251胶质瘤干细胞干性及生物学行为的影响及其对放疗敏感性的影响；（3）探索Per2对胶质瘤干细胞干性调控及放疗敏感性影响的分子机制。 .关键数据及重要结果：（1）U87细胞分离培养获得繁殖力强、多向分化的GSCs；测GSCs干性标记物CD133、SOX2、NESTIN表达和自我更新能力；Per2基因在U87及GCSs均呈节律表达。（2）转染Per2过表达慢病毒构建Per2过表达胶质瘤干细胞株，上调γH2AX表达、增强U87胶质瘤干细胞放疗敏感性。（3）U87、U251胶质瘤GCSs、胶质瘤细胞和正常胶质细胞Per2表达显示在胶质瘤干细胞中Per2 mRNA和蛋白水平均下调，与GSCs增殖率和转移侵袭力及细胞周期密切相关。（4）建立裸鼠异种移植瘤模型发现Per2过表达的干细胞成瘤能力明显下降并和放疗敏感性相关。（5）进一步通过高通量测序、发现在GCSs中Wnt/β-Catenin信号通路是Per2发挥作用的重要下游信号途径，同时Per2和干性相关基因ID1和ID3密切相关。.科学意义：胶质瘤干细胞中Per2的下调；上调Per2通过抑制Wnt/β-Catenin信号通路抑制GSCs的干性和恶性表型及增强胶质瘤干细胞的放疗敏感作用。Per2有望成为增强胶质瘤放化疗敏感性、降低胶质瘤复发的新的分子靶点，为肿瘤时辰治疗提供新的理论依据和思路。