烟酰胺单核苷酸对乙醇诱导L02细胞DNA损伤的保护作用研究  

Protective effects of nicotinamide mononucleotide on ethanol-induced DNA damage in L02 cells

作　　者：狄春红[1] 阴洁[1] 钟文英[1] 张盈盈 曹月佳 谭晓华[2] DI Chunhong;YIN Jie;ZHONG Wenying;ZHANG Yingying;CAO Yuejia;TAN Xiaohua(Department of Laboratory Testing,Affiliated Hospital of Hangzhou Normal University,Hangzhou,Zhejiang 310015,China;Hangzhou Normal University,Hangzhou,Zhejiang 311121,China)

机构地区：[1]杭州师范大学附属医院检验科,浙江杭州310015 [2]杭州师范大学,浙江杭州311121

出　　处：《预防医学》2024年第6期548-552,共5页CHINA PREVENTIVE MEDICINE JOURNAL

基　　金：杭州市生物医药和健康产业发展扶持科技专项项目(2021WJCY144,2022WJC016);浙江省自然科学基金项目(LY23H190001,LQ18H190003)。

摘　　要：目的探究烟酰胺单核苷酸(NMN)对乙醇诱导L02细胞DNA损伤的保护作用,为NMN辅助治疗酒精性肝病提供依据。方法采用不同浓度NMN(1、2、4和8 mmol/L)预处理L02细胞6 h后,暴露于0.4%乙醇12 h,分为对照组、0.4%乙醇组和不同浓度NMN组。采用台盼蓝染色分析细胞活力,确定NMN作为保护剂的浓度;通过碱性彗星实验、H2组蛋白家族X(γH2AX)免疫荧光检测和活性氧(ROS)检测评估NMN对乙醇诱导L02细胞DNA损伤的影响。L02细胞暴露于0.4%乙醇12 h后,采用含保护浓度NMN的培养基继续培养,分为PBS对照组和NMN组,分别在0、2、4、8、16和32 h检测细胞活力;通过碱性彗星实验评估NMN对乙醇诱导L02细胞DNA损伤修复的影响。结果与对照组比较,0.4%乙醇组L02细胞活力降低;与0.4%乙醇组比较,各浓度NMN组L02细胞活力升高(均Ρ<0.05);4 mmol/L及以上NMN组L02细胞活力与对照组差异无统计学意义(均Ρ>0.05),故选用4 mmol/L为NMN作为保护剂的浓度。与对照组比较,0.4%乙醇组L02细胞尾矩增加,γH2AX免疫荧光相对强度和ROS相对水平升高(均Ρ<0.05);与0.4%乙醇组比较,4 mmol/L及以上NMN组L02细胞尾矩缩短,γH2AX免疫荧光相对强度和ROS相对水平下降(均Ρ<0.05)。随着4 mmol/LNMN干预时间增加,L02细胞活力逐渐升高,尾矩逐渐缩短;与PBS对照组比较,干预4 h及以上NMN组L02细胞活力较高,尾矩较短(均Ρ<0.05)。结论NMN以剂量依赖方式减轻DNA损伤,并以时间依赖方式促进DNA损伤的修复,NMN对乙醇诱导肝细胞DNA损伤具有保护作用。Objective To investigate protective effects of nicotinamide mononucleotide(NMN)on ethanol-induced DNA damage in L02 cells,so as to provide the evidence for adjuvant therapy of NMN on alcoholic liver diseases.Methods L02 cells were pretreated with different concentrations of NMN(0,1,2,4 and 8 mmol/L)for 6 h,and then were exposed to 0.4%ethanol for 12 h.The treated cells were divided into the control group,0.4%ethanol group and different concentrations of NMN groups.Cell viability was analyzed using trypan blue staining for determining the concentration of NMN as a protective agent.The effects of NMN on ethanol-induced DNA damage in L02 cells were evaluated using immunofluorescence detection and reactive oxygen species(ROS)assay.L02 cells were exposed to 0.4%ethanol for 12 h,cultured in a medium containing a protective concentration of NMN,and divided into PBS group and NMN group.Cell viability was detected at 0,2,4,8,16 and 32 h,and the effects of NMN on repairing ethanol-induced DNA damage were evaluated by alkaline comet assay.Results The cell viability was lower in 0.4%ethanol group than than in the control group,and was higher in different concentrations of NMN groups than in 0.4%ethanol group(all P<0.05),with no significant difference in the cells viability between 4 mmol/L and higher concentrations of NMN groups and the control group(all P>0.05).Therefore,4 mmol/L NMN was selected as a protective agent.The cell tail moments,relative immunofluorescence intensities ofγH2AX and relative levels of ROS were higher in 0.4%ethanol group than in the control group,and lower in 4 mmol/L and higher concentrations of NMN groups than in 0.4%ethanol group(all P<0.05).The cell viability was increased and the cell tail moment was shortened with the increase of 4 mmol/L NMN intervention time;and the cell viability in 4 h and more of NMN groups were higher,and the cell tail moment were lower than that in PBS group(all P<0.05).Conclusions NMN attenuates DNA damage in a dose-dependent manner and promotes the repair of DNA dama

关 键 词：烟酰胺单核苷酸 乙醇 DNA损伤 活性氧 

分 类 号：R575[医药卫生—消化系统]