Despite the evidences of elevated expression of Mer tyrosine kinase (MerTK) in multiple human cancers, mechanisms underlying the oncogenic roles of MerTK in hepatocellular carcinoma (HCC) remains undefined. We explored the functional effects of MerTK and N-Glycosylated MerTK on HCC cell survival and tumor growth. Here, we show that MerTK ablation increases reactive oxygen species (ROS) production and promotes the switching from glycolytic metabolism to oxidative phosphorylation in HCC cells, thus suppressing HCC cell proliferation and tumor growth. MerTK is N-glycosylated in HCC cells at asparagine 294 and 454 that stabilizes MerTK to promote oncogenic transformation. Moreover, we observed that nuclear located non-glycosylated MerTK is indispensable for survival of HCC cells under stress. Pathologically, tissue microarray (TMA) data indicate that MerTK is a pivotal prognostic factor for HCC. Our data strongly support the roles of MerTK N-glycosylation in HCC tumorigenesis and suggesting N-glycosylation inhibition as a potential HCC therapeutic strategy.

目的 构建PMLA216T-RARα过表达质粒,并研究雄黄对其蛋白产物的影响及机制.方法 通过资料库及文献获得PML、RARα的mRNA序列,以及PML-RARα[的融合位点.设计引物后,通过聚合酶链反应获得目的 片段,通过无缝克隆技术连接两个片段,插入突变,并与载体质粒相连接,测序验证.转染质粒至HL-60细胞,血清饥饿法处理24 h后用Western blot观察PML-RARα、p-mTOR、p62、LC3B水平变化;不同浓度的雄黄处理24 h后,用Western blot观察PML-RARα、p-mTOR、p62、LC3B水平变化.结果 测序结果显示质粒构建成功,序列符合预期设计.1、2、4、8μmol/L雄黄处理后PMLWT-RARα和PMLA216T-RARα水平低于未经雄黄处理(0μmol/L)的PMLWT-RARα和PMLA216T-RARα水平,差异有统计学意义(P＜0.05);0.5 μmol/L雄黄处理后PMLWT-RARα水平低于未经雄黄处理的PMLWT-RARα水平,差异有统计学意义(P＜0.05).相同浓度雄黄处理后PMLA216T-RARα水平高于PMLWT-RARα水平,差异有统计学意义(P＜0.05).5％ FBS和未经FBS处理的PMLA216T-RARα、p-mTOR水平低于10％ FBS处理的PMLA216T-RARα、p-mTOR水平,p62、LC3B水平高于10％ FBS处理的PMLA216T-RARα水平,差异有统计学意义(P＜0.05).4、8、16、32、64 μmol/L雄黄处理后PMLA216T-RARα水平低于未经雄黄处理的PMLA216T-RARα水平,差异有统计学意义(P＜ 0.05);8、16、32、64μmol/L雄黄处理后p-mTOR、p62水平低于未经雄黄处理的p-mTOR、p62水平,LC3B水平高于未经雄黄处理的LC3B水平,差异有统计学意义(P＜0.05);16 μmol/L雄黄处理后p-mTOR、p62水平低于8μmol/L雄黄处理后的p-mTOR、p62水平,LC3B水平高于8 μmol/L雄黄处理后的LC3B水平,差异有统计学意义(p＜0.05).结论 错义突变型PMLA216T-RARα过表达质粒构建成功,高浓度雄黄可能通过选择性自噬下调PMLA216T-RARα.

该文初步探讨了环吡酮胺(ciclopirox olamine,CPX)抑制人胶质瘤细胞SHG44生长的作用和机制.该研究用梯度浓度的CPX处理人胶质瘤细胞SHG44后,通过MTT实验检测药物半抑制浓度IC50和细胞增殖能力;应用平板克隆实验检测细胞克隆形成能力;Transwell小室和细胞划痕实验检测细胞侵袭和迁移能力;流式细胞术检测细胞内活性氧(reactive oxygen species,ROS)、线粒体内ROS以及细胞凋亡的变化;Seahorse XF96 Flux analysis分析仪检测细胞耗氧率(oxygen comsumption rate,OCR);实时荧光定量PCR(qPCR)和Western blot技术检测细胞内mRNA和蛋白的变化.结果 表明:SHG44对CPX药物敏感;CPX能使STAT3在mRNA转录水平和蛋白表达水平下降,抑制SHG44的迁移和侵袭,同时,促进线粒体ROS的累积,最终破坏线粒体功能抑制细胞生长并促进细胞凋亡.该研究结果提示,CPX具有一定的抗胶质瘤细胞SHG44生长的作用,有望成为一种治疗胶质瘤的药物.

The primary liver cancer (PLC) is one of the leading causes of cancer-related death worldwide. The predominant form of PLC is hepatocellular carcinoma (HCC), which accounts for about 85% of all PLC. Artemisinin (ART) was clinically used as anti-malarial agents. Recently, it was demonstrated to inhibit cell growth and migration in multiple cancer types. However, the molecular mechanism underlying these anti-cancer activity remains largely unknown. Herein, it is discovered that ART dramatically suppresses HCC cell growth in vitro through arresting cell cycle progression, and represses cell migration and invasion via regulating N-cadherin-Snail-E-cadherin axis. In addition, the disruption of cellular bioenergetics contributed to ART-caused cell growth, migration and invasion inhibition. Moreover, ART (100 mg/kg, intraperitoneally) substantially inhibits HCC xenograft growth in vivo. Importantly, Hippo-YAP signal transduction is remarkably inactivated in HCC cells upon ART administration. Collectively, these data reveal a novel mechanism of ART in regulating HCC cell growth, migration, and invasion, which indicates that ART could be considered as a potential drug for the treatment of HCC.