Department of Neurosurgery, Penn State College of Medicine, Hershey, Pennsylvania, United States of America.
Department of Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania, United States of America.
PLoS One. 2019 Sep 6;14(9):e0221952. doi: 10.1371/journal.pone.0221952. eCollection 2019.
Elevated expression of the iron regulatory protein, ferritin heavy chain 1 (FTH1), is increasingly being associated with high tumor grade and poor survival outcomes in glioblastoma. Glioma initiating cells (GICs), a small population of stem-like cells implicated in therapeutic resistance and glioblastoma recurrence, have recently been shown to exhibit increased FTH1 expression. We previously demonstrated that FTH1 knockdown enhanced therapeutic sensitivity in an astrocytoma cell line. Therefore, in this study we developed a liposomal formulation to enable the in vitro delivery of FTH1 siRNA in patient xenograft derived GICs from glioblastomas with pro-neural and mesenchymal transcriptional signatures to interrogate the effect of FTH1 downregulation on their radiation sensitivity. Transfection with siRNA decreased FTH1 expression significantly in both GICs. However, there were inherent differences in transfectability between pro-neural and mesenchymal tumor derived GICs, leading us to modify siRNA: liposome ratios for comparable transfection. Moreover, loss of FTH1 expression resulted in increased extracellular lactate dehydrogenase activity, executioner caspase 3/7 induction, substantial mitochondrial damage, diminished mitochondrial mass and reduced cell viability. However, only GICs from pro-neural glioblastoma showed marked increase in radiosensitivity upon FTH1 downregulation demonstrated by decreased cell viability, impaired DNA repair and reduced colony formation subsequent to radiation. In addition, the stemness marker Nestin was downregulated upon FTH1 silencing only in GICs of pro-neural but not mesenchymal origin. Using liposomes as a siRNA delivery system, we established FTH1 as a critical factor for survival in both GIC subtypes as well as a regulator of radioresistance and stemness in pro-neural tumor derived GICs. Our study provides further evidence to support the role of FTH1 as a promising target in glioblastoma.
铁调节蛋白，铁蛋白重链 1（FTH1）的表达升高与胶质母细胞瘤中的高肿瘤分级和不良生存结果越来越相关。神经胶质瘤起始细胞（GICs），一种被认为与治疗抵抗和胶质母细胞瘤复发有关的干细胞样细胞的小群体，最近已被证明表现出 FTH1 表达增加。我们之前证明了 FTH1 敲低可增强星形细胞瘤细胞系的治疗敏感性。因此，在这项研究中，我们开发了一种脂质体制剂，以能够在具有神经前体和间质转录特征的源自胶质母细胞瘤的患者异种移植物衍生的 GIC 中体外递送 FTH1 siRNA，以研究 FTH1 下调对其辐射敏感性的影响。siRNA 转染可使两种 GIC 中的 FTH1 表达显著降低。然而，神经前体细胞和间质肿瘤衍生的 GIC 之间在转染能力上存在固有差异，导致我们修改了 siRNA：脂质体比例以实现可比的转染。此外，FTH1 表达的丧失导致细胞外乳酸脱氢酶活性增加、效应器半胱氨酸蛋白酶 3/7 诱导、线粒体严重损伤、线粒体质量减少和细胞活力降低。然而，只有源自神经前体胶质母细胞瘤的 GIC 在 FTH1 下调后表现出明显的放射敏感性增加，表现为细胞活力降低、DNA 修复受损和随后的辐射后集落形成减少。此外，只有源自神经前体的 GIC 中，FTH1 沉默会导致干性标志物 Nestin 下调，而源自间质的 GIC 则不会。使用脂质体作为 siRNA 递送系统，我们确定 FTH1 是两种 GIC 亚型存活的关键因素，也是神经前体细胞来源的 GIC 中辐射抗性和干性的调节因子。我们的研究为 FTH1 作为胶质母细胞瘤有前途的靶点提供了进一步的证据。
