维甲酸介导的线粒体片段化抑制肝癌细胞营养缺乏相关凋亡抵抗

Retinoic acid-mediated mitochondrial fragmentation inhibits apoptosis resistance related to nutrient deficiency in hepatocellular carcinoma cells

目的探索代谢物和线粒体如何在营养缺乏环境中调控肝癌细胞存活及其相关机制。方法通过分析肝动脉栓塞化疗(TACE)治疗敏感及耐受肝癌样本的转录组数据,鉴定肝癌细胞抵抗营养缺乏的关键通路;在营养剥夺耐受(HLF、Huh7)和敏感(Hep3B、HLE)细胞中检测维甲酸含量;利用维甲酸或维甲酸受体抑制剂处理肝癌细胞,MTS、流式细胞术等检测细胞在营养剥夺下的生长及凋亡情况;利用透射电镜和激光共聚焦显微镜观察维甲酸对肝癌细胞线粒体数目、形态变化的影响。结果转录组分析提示TACE治疗耐受与敏感肝癌组织差异基因主要富集在视黄醇代谢通路。进一步发现与营养剥夺敏感肝癌细胞相比,耐受肝癌细胞中视黄醇代谢产物维甲酸显著降低(P<0.01)。HLF细胞在营养剥夺情况下补充维甲酸,线粒体呈片段化且细胞死亡增加(P<0.01);而在Hep3B细胞中抑制维甲酸受体,线粒体片段化减少,细胞死亡减少(P<0.01)。结论补充维甲酸可通过诱导肝癌细胞线粒体片段化抑制肝癌细胞在营养缺乏环境下的凋亡抵抗,而低水平维甲酸可通过增强线粒体融合,促进营养缺乏诱导的肝癌细胞凋亡抵抗。

Objective To explore how metabolites and mitochondria regulate the survival of hepatocellular carcinoma(HCC)cells and its related mechanisms in a nutrient-deficient environment.Methods Transcriptomic data of sensitive and tolerant HCC samples treated by transarterial chemoembolization(TACE)were analyzed to identify the critical pathways for HCC cells to resist nutrient deficiency.The content of retinoic acid was detected in nutrient-deprived tolerant(HLF,Huh7)and sensitive(Hep3B,HLE)cells.HCC cells were treated with retinoic acid or retinoic acid receptor inhibitors.MTS assays and flow cytometry were used to detect the growth and apoptosis of the cells under nutrient deprivation.The influence of retinoic acid on the number and morphology of mitochondria in HCC cells was observed using transmission electron microscopy and laser scanning confocal microscopy.Results Transcriptomic analysis revealed that differentially expressed genes between TACE-treated tolerant and sensitive HCC tissues were predominantly enriched in retinol metabolism pathway.Furthermore,it was found that retinoic acid,a metabolite of retinol metabolism,was significantly reduced in tolerant HCC cells compared with sensitive HCC cells under nutrient-deprived conditions(P<0.01).Supplementation of retinoic acid in HLF cells subjected to nutrient deprivation led to mitochondrial fragmentation and increased cell death(P<0.01).In contrast,inhibition of retinoic acid receptor in Hep3B cells resulted in reduced mitochondrial fragmentation and decreased cell death(P<0.01).Conclusion Supplementation of retinoic acid can inhibit the apoptosis resistance of HCC cells under nutrient-deficient conditions by inducing mitochondrial fragmentation,while low levels of retinoic acid can promote apoptosis resistance of HCC cells induced by nutrient deficiency by enhancing mitochondrial fusion.