蛋白质类泛素修饰角度解读亚低温对新生儿缺氧缺血性脑病的保护机制  

The role of SUMOylation in the protective mechanisms of therapeutic hypothermia on neonatal hypoxic-ischemic encephalopathy

作　　者：刘晓智[1] 徐盼盼 张春艳 高瑕 刘光碧 楚冬梅[3] 刘扬 田秀英[2] 郑军[2] Liu Xiaozhi;Xu Panpan;Zhang Chunyan;Gao Xia;Liu Guangbi;Chu Dongmei;Liu Yang;Tian Xiuying;Zheng Jun(Tianjin Fifth Central Hospital,Tianjin Key Laboratory of Epigenetics for Organ Development of Premature Infants,Tianjin 300450,China;Department of Neonatology,Tianjin Central Hospital of Gynecology Obstetrics,Tianjin 300199,China;Department of Pediatrics,Tianjin Fifth Central Hospital,Tianjin 300450,China;Department of Pediatric Surgery,Tianjin Fifth Central Hospital,Tianjin 300450,China)

机构地区：[1]天津市第五中心医院,天津市早产儿器官发育表观遗传重点实验室,天津300450 [2]天津市中心妇产科医院新生儿科,天津300199 [3]天津市第五中心医院儿科,天津300450 [4]天津市第五中心医院小儿外科,天津300450

出　　处：《中华新生儿科杂志（中英文）》2023年第5期294-300,共7页Chinese Journal of Neonatology

基　　金：天津市自然科学基金项目(21JCZDJC01270);天津市医学重点学科(专科)建设项目(TJYXZDXK-062B)。

摘　　要：目的从蛋白质类泛素化修饰角度解析亚低温对神经干细胞(neural stem cells,NSCs)的保护机制,为新生儿缺氧缺血性脑病(hypoxic ischemic encephalopathy,HIE)的亚低温治疗提供科学依据。方法原代分离培养小鼠NSCs,分为对照组、缺氧组、亚低温组、缺氧+亚低温组并予相应处理,Western Blot法检测小泛素相关修饰蛋白2/3(small ubiquitin-related modifier 2/3,SUMO2/3)、低氧诱导因子1 α(hypoxia inducible factor 1 α,HIF-1α)、过氧化物酶体增殖物激活受体γ辅激活因子1 α(peroxisome proliferator-activated receptor gamma coactivator 1 α,PGC-1α)和八聚体结合转录因子4(octamer-binding transcription factor 4,Oct4)的蛋白水平;比较NSCs克隆球直径变化,酶联免疫吸附法检测乳酸脱氢酶(lactate dehydrogenase,LDH)含量,流式细胞术检测细胞凋亡率,免疫荧光法检测NSCs向神经元细胞分化情况。结果与对照组相比,缺氧组NSCs中SUMO2/3、HIF-1α和PGC-1α的蛋白水平分别升高至1.33、2.26和2.40倍,亚低温组NSCs中SUMO2/3和PGC-1α的蛋白表达水平分别升高至1.52倍和6.36倍,差异均有统计学意义(P<0.05);与缺氧组相比,缺氧+亚低温组NSCs中SUMO2/3、HIF-1α、PGC-1α和Oct4的蛋白表达水平分别升高至1.44、1.40、3.30和1.59倍,差异均有统计学意义(P<0.05)。缺氧组、亚低温组、缺氧+亚低温组细胞克隆球直径明显小于对照组,缺氧+亚低温组细胞克隆球直径明显小于缺氧组,差异均有统计学意义(P<0.05)。亚低温组与对照组LDH水平比较差异无统计学意义(P>0.05),缺氧+亚低温组LDH水平明显低于缺氧组(P<0.05)。亚低温组细胞死亡率与对照组比较差异无统计学意义(P>0.05),缺氧+亚低温组细胞死亡率明显低于缺氧组(P<0.05)。与对照组相比,缺氧组和亚低温组巢蛋白表达水平均有所升高,但神经元特异性烯醇化酶(neuron-specific enolase,NSE)水平明显下降(P<0.05);与缺氧组和亚低温组相比,缺氧+亚低温Objective To study the role of SUMOylation in the process of therapeutic hypothermia on neural stem cells(NSCs)in neonatal hypoxic-ischemic encephalopathy.Methods SUMOylation is an essential post-translational modification involving small ubiquitin-like modifiers(SUMOs).Primary-cultured NSCs from mice were assigned into four groups:control group,hypoxia group,hypothermia group and hypoxia+hypothermia group.Western Blot was used to detect the protein levels of SUMO2/3,hypoxia-inducible factor-1α(HIF-1α),peroxisome proliferator-activated receptorγcoactivator factor 1α(PGC-1α)and octamer binding transcription factor 4(Oct4).The diameters of NSCs were compared.ELISA was used to detect lactate dehydrogenase(LDH)level.Apoptosis was examined using flow cytometry.Immunofluorescence method was used to measure the differentiation of NSCs into neuronal cells.Results Compared with the control group,the levels of SUMO2/3,HIF-1αand PGC-1αin NSCs of the hypoxia group increased 33%,126%and 140%,respectively(P<0.05).Compared with the control group,the levels of SUMO2/3 and PGC-1αin NSCs of the hypothermia group increased 52%and 536%,respectively(P<0.05).Compared with the hypoxia group,the levels of SUMO2/3,HIF-1α,PGC-1αand Oct4 in the hypoxia+hypothermia group increased 44%,40%,230%and 59%,respectively(P<0.05).The diameters of NSCs in hypoxia group,hypothermia group and hypoxia+hypothermia group were smaller than control group,and hypoxia+hypothermia group smaller than hypoxia group(P<0.05).No significant differences existed in LDH levels between hypothermia group and control group(P>0.05).LDH level in hypoxia+hypothermia group were significantly lower than hypoxia group(P<0.05).No significant differences existed in the cell death rates between hypothermia group and control group(P>0.05).The cell death rate in hypoxia+hypothermia group was significantly lower than hypoxia group(P<0.05).Compared with the control group,the expressions of Nestin in both hypoxia group and hypothermia group were increased,but neuron specific

关 键 词：缺氧缺血性脑病 小泛素相关修饰蛋白 亚低温疗法 神经干细胞 

分 类 号：R722.1[医药卫生—儿科]