氧含量通过调节蛋白质类泛素化修饰影响肝癌化疗药物敏感性的研究  

Study of oxygen content affecting the chemosensitivity of liver cancer by regulating protein SUMOylation

作　　者：闫玉兰[1] 张春艳 邵志江[2] 陈贺凯 张树朋[2] 赵徽[2] 吴问汉 YAN Yulan;ZHANG Chunyan;SHAO Zhijiang;CHEN Hekai;ZHANG Shupeng;ZHAO Hui;WU Wenhan(Department of Hematology and Oncology,Tianjin Fifth Central Hospital,Tianjin 300450,China;Department of General Surgery,Tianjin Fifth Central Hospital,Tianjin 300450,China;Tianjin Key Laboratory of Epigenetic Organ Development of Preterm Infants,Tianjin 300450,China;Department of General Surgery,Peking University First Hospital,Beijing 100034,China)

机构地区：[1]天津市第五中心医院血液肿瘤科,天津300450 [2]天津市第五中心医院普通外科,天津300450 [3]天津市早产儿器官发育表观遗传重点实验室,天津300450 [4]北京大学第一医院普通外科,北京100034

出　　处：《现代肿瘤医学》2023年第23期4295-4300,共6页Journal of Modern Oncology

基　　金：国家自然科学基金项目(编号:82272880);天津市卫生健康委员会科技项目(编号:TJWJ2022XK041,TJWJ2022XK042,TJWJ2022MS051,TJWJ2022QN101);天津市医学重点学科(专科)建设项目资助(编号:TJYXZDXK-079D)。

摘　　要：目的:从蛋白质类泛素化修饰角度解读氧含量对肝癌化疗药物敏感性的影响,为未来肝癌患者的化疗增敏治疗提供实验依据。方法:以人源性肝癌细胞株Hep3B为实验对象,分别置于乏氧、常氧及高氧培养环境,Western Blot法检测小泛素样相关修饰蛋白1(small ubiquitin-like-related modified protein 1,SUMO1)、SUMO2/3、性别决定区Y-box2(sex-determining region Y-box2,Sox2)和八聚体结合转录因子4(octamer-binding transcription factor 4,Oct4)的蛋白表达水平;观察Hep3B细胞克隆球形成率;然后在Hep3B细胞培养基中加入顺铂,MTT法检测肿瘤细胞半数致死量,ELISA法检测乳酸脱氢酶(lactate dehydrogenase,LDH)含量,流式细胞术检测细胞凋亡率。结果:乏氧及高氧均不影响共价态SUMO1产物的增加(P>0.05),乏氧能够明显增加共价态SUMO2/3产物,但高氧则短暂升高共价态SUMO2/3产物后又快速下调;乏氧能够增加肿瘤细胞克隆球形成率及细胞干性维持蛋白Sox2和Oct4的蛋白表达(P<0.05),而高氧则降低克隆球形成率及Sox2和Oct4的蛋白表达(P<0.05);乏氧能够提高Hep3B细胞对顺铂的半数致死量(P<0.05),降低LDH水平及细胞凋亡率(P<0.05),高氧则能够降低Hep3B细胞对顺铂的半数致死量(P<0.05),提高LDH水平及细胞凋亡率(P<0.05)。结论:乏氧能够通过提高蛋白质SUMO化修饰水平增加肝癌细胞干性,降低化疗药物敏感性;高氧则能够通过降低蛋白质SUMO化修饰水平降低肝癌细胞干性,提高化疗药物敏感性。Objective:To analyze the effect of oxygen content on the chemosensitivity of liver cancer from the perspective of protein SUMOylation modification,and provide experimental basis for chemotherapy sensitization of liver cancer patients in the future.Methods:The human-derived hepatoma cell line Hep3B was cultured in hypoxia,normoxia and hyperoxia,and Western Blot was used to detect the protein expression level of small ubiquitin-like-related modified protein 1(SUMO1),SUMO2/3,and sex-determining region Y-box2(Sox2)and octamer-binding transcription factor 4(Oct4).The formation rate of Hep3B cell clone spheres was observed.Then,cisplatin was added to Hep3B cell culture medium,3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide(MTT)method was used to detect the median lethal dose of tumor cells.The content of lactate dehydrogenase(LDH)was detected by enzyme linked immunosorbent assay(ELISA),and the apoptosis rate was detected by flow cytometry.Results:Both hypoxia and hyperoxia did not affect the increase of covalent SUMO1 products(P>0.05).Hypoxia could significantly increase covalent SUMO2/3 products,but hyperoxia temporarily increased covalent SUMO2/3 products and then decreased again.Hypoxia can increase the formation rate of clonal spheres and the protein expressions of cell stemness maintenance proteins Sox2 and Oct4(P<0.05),while hyperoxia decreased the formation rate of clonal spheres and the protein expressions of Sox2 and Oct4(P<0.05).Hypoxia can increase the median lethal dose of Hep3B cells to cisplatin(P<0.05),reduce the level of LDH and apoptosis rate(P<0.05),and hyperoxia can reduce the median lethal dose of Hep3B cells to cisplatin(P<0.05),increased LDH level and apoptosis rate(P<0.05).Conclusion:Hypoxia can increase the stemness of liver cancer cells and reduce the sensitivity of chemotherapy by increasing the level of protein SUMOylation.Hyperoxia can reduce the stemness of liver cancer cells and improve the sensitivity of chemotherapy by reducing the level of protein SUMOylation.

关 键 词：肝癌 小泛素相关修饰蛋白 乏氧 高氧 化疗敏感性 

分 类 号：R735.7[医药卫生—肿瘤]