Alpha B-Crystallin改变肝癌能量代谢影响细胞上皮间质转化及sorafenib耐药研究

Energy metabolism plays a critical role in cancers, However, a systematic study for the role of energy metabolism in hepatocellular carcinoma is lacked. Previously, we detected alpha B-Crystallin (Cryab) overexpression induced epithelial-mesenchymal transition (EMT), promoting liver cancer cell invasion and metastasis. The process of tumor invasion and metastasis needs glucose for energy. Therefore, we hypothesized that Cryab not only induce EMT, and may regulate energy metabolism. To confirm these assumptions, the study intends to investigate glucose metabolic changes during Cryab-induced EMT process and subsequent epithelial mesenchymal transformation process by Cryab down-regulation. Using Gene microarray technology can detect Cryab-regulated genome-wide distribution patterns, and then we combine the data with bioinformatics analysis and find key protein regulating glucose metabolism. Using gene transfection regulates the level of key protein, changes of cell invasion, metastasis biological behavior and morphology are observed, while the relationship between the key protein and EMT, metabolism is investigated in orthotopic liver cancer model. Moreover，we investigated the relationship between the energy metabolism the above key protein regulated and sorafenib resistance.

能量代谢在肿瘤发生发展中起重要作用，但肝癌中能量代谢的研究还较欠缺。我们前期已揭示了alpha B-Crystallin（Cryab）过表达诱导肝癌上皮间质转化(EMT)及sorafenib耐药，促进侵袭转移。而肿瘤侵袭转移过程中需要糖类供能。故我们推测Cryab不仅诱导EMT，而且可调节能量代谢。为了证实以上假设，本项目拟检测Cryab诱导EMT过程中及随后Cryab下调诱导间质上皮转化过程中糖类代谢变化；利用表达谱芯片技术结合生物信息学分析并找到Cryab调控糖代谢关键蛋白，采用糖代谢关键蛋白基因转染和干扰技术，双向调节Cryab调控的糖代谢关键蛋白的表达水平，观察细胞侵袭、转移及形态等生物学行为变化调查Cryab调控的能量代谢与EMT关系，并且在肝癌组织及肝癌原位模型验证筛选蛋白与EMT、代谢的关系。而且我们进一步调查筛选蛋白调节的能量代谢能否影响sorafenib耐药。

近年，上皮间质转化诱导的肿瘤(包括肝癌)侵袭转移受到极大关注。我们前期证实：热休克蛋白Cryab是影响肝癌上皮间质转化的重要分子。异常的葡萄糖代谢参与许多癌症的肿瘤发生和进展。 一些肿瘤细胞即使在具有充足氧气的条件下优先依赖于糖酵解而不是氧化磷酸化（“有氧糖酵解”）。 虽然以前的研究已经非常注意这种异常的糖酵解，但很少研究了解糖原异生的作用。果糖-1,6-二磷酸酶（FBP1）是糖异生中的限速酶之一，催化果糖1,6二磷酸（F-1,6-P2）水解为果糖-6-磷酸（F-6-P）和无机磷酸盐。最近，据报道，FBP1在几种癌症中发挥肿瘤抑制作用。然而，失调的FBP1在HCC中的作用和机制仍然不清楚。 本课题在前期工作基础上，对肝细胞癌上皮间质转化行进一步的研究， 证实：（1）调查了肝癌中糖异生关键酶果糖-1,6-二磷酸酶（FBP1）与核转录因子Snail的表达关系，证实了snail可下调FBP1的水平。 （2）过表达FBP1水平诱导肿瘤细胞侵袭与转移作用明显下调；过表达FBP1水平可使snail诱导的细胞上皮-间质转化细胞间质上皮转化（3）动物实验也证实FBP1过表达后肿瘤生长减慢。（4）临床上snail高表达与FBP1低表达预后最差。FBP1的表达与AFP水平，卫星灶，门静脉癌栓和晚期肿瘤分期显着相关。生存分析显示FBP1是总体生存和无复发生存的独立预后因素。（5）我们通过chip-sequence 验证了snail可结合FBP1的启动子抑制FBP1的表达。