葛根素通过核因子E2相关因子2通路对骨髓间充质干细胞氧化损伤的保护作用  

Puerarin regulates the apoptosis and osteogenic differentiation of bone marrow mesenchymal stem cells under oxidative stress through nuclear factor E2 related factor 2

作　　者：安晋阳 蒋娜 刘进进 杨宽 关聪会[1,2] 汤旭磊[2] AN Jin-yang;JIANG Na;LIU Jin-jin;YANG Kuan;GUAN Cong-hui;TANG Xu-lei(The First Clinical Medical College,Lanzhou University,Lanzhou 730000,Gansu Province,China;Department of Endocrinology,The First Hospital of Lanzhou University,Lanzhou 730000,Gansu Province,China)

机构地区：[1]兰州大学第一临床医学院,甘肃兰州730000 [2]兰州大学第一医院内分泌科,甘肃兰州730000

出　　处：《中国临床药理学杂志》2023年第5期684-688,共5页The Chinese Journal of Clinical Pharmacology

基　　金：国家自然科学基金资助项目(81370970)。

摘　　要：目的研究葛根素(Pue)对过氧化氢(H_(2)O_(2))诱导的骨髓间充质干细胞(BMSC)的氧化损伤、凋亡和成骨分化作用的影响,及核因子E2相关因子2(NRF2)信号通路的作用。方法选择4周龄SD雄性大鼠,体外分离培养BMSC,收集第4代细胞。将BMSC随机分为5组,分别为对照组(未作处理)、模型组(300μmol·L^(-1)H_(2)O_(2))、Pue组(0.1μmol·L^(-1)Pue)、Pue预处理组(0.1μmol·L^(-1)Pue预处理后,300μmol·L^(-1)H_(2)O_(2)处理)、NRF2抑制剂组(10μmol·L^(-1)的ML385预处理,后同Pue预处理组)。以微板法测丙二醛(MDA)、还原型谷胱甘肽(GSH)含量;以蛋白质印迹法测胞核、胞质中NRF2、B淋巴细胞瘤-2相关蛋白x(Bax)、Runt相关转录因子2(Runx2)蛋白表达;通过茜素红半定量检测各组细胞矿化情况。结果对照组、模型组、Pue组、Pue预处理组、NRF2抑制剂组中MDA含量分别为(0.50±0.02)、(0.89±0.02)、(0.45±0.01)、(0.57±0.01)、(0.72±0.02)nmol·g·prot^(-1);这5组的细胞核NRF2蛋白表达水平分别为1.00±0.11、1.10±0.05、1.37±0.05、1.90±0.13、0.68±0.11;这5组的Bax蛋白表达水平分别为1.00±0.07、2.90±0.21、0.58±0.07、1.93±0.11、2.44±0.09;这5组的Runx2蛋白表达水平分别为1.00±0.05、0.42±0.02、1.97±0.06、0.87±0.09、0.22±0.03;这5组的矿化结节溶解后的光密度值分别为2.21±0.05、1.36±0.04、3.15±0.09、1.54±0.08、0.83±0.05。上述指标,模型组与对照组比较,差异有统计学意义(P<0.05);Pue预处理组与模型组比较,差异有统计学意义(P<0.05);NRF2抑制剂组与Pue预处理组比较,差异有统计学意义(P<0.01)。结论Pue可通过激活NRF2通路,减轻H_(2)O_(2)诱导的BMSC的氧化损伤,并在抗凋亡及促进成骨中起着关键作用。Objective To study the effects of puerarin(Pue)on hydrogen peroxide(H_(2)O_(2))-induced oxidative damage,apoptosis and osteogenic differentiation of bone marrow mesenchymal stem cells(BMSC)under oxidative stress,and the effects of nuclear factor E2related factor 2(NRF2)signaling pathway effect.Methods 4 weeks old SD male rats were selected,BMSC were isolated and cultured in vitro,and the fourth passage cells were collected.BMSC were randomly divided into 5 groups,control group(untreated),hydrogen peroxide group(300μmol·L^(-1)H_(2)O_(2)),Pue group(0.1μmol·L^(-1)Pue),and Pue pretreatment group(0.1μmol·L^(-1)Pue pretreatment,after treatment,300μmol·L^(-1)H_(2)O_(2)treatment),NRF2 inhibitor group(10μmol·L^(-1)ML385pretreatment,and later the same as puerarin pretreatment group).The content of malondialdehyde(MDA)and reduced glutathione(GSH)was measured by microplate method;Western blotting was used to measure NRF2,B-lymphoma-2 related protein X(Bax),Runt-related transcription factor 2(Runx2)protein expression.The mineralization of each group was detected semi-quantitatively by Alizarin red.Results The contents of MDA in the control group,hydrogen peroxide group,puerarin group,puerarin pretreatment group,and NRF2 inhibitor group were(0.50±0.02),(0.89±0.02),(0.45±0.01),(0.57±0.01),(0.72±0.02)nmol·g·prot^(-1);the nuclear NRF2 protein expression in these five groups were 1.00±0.11,1.10±0.05,1.37±0.05,1.90±0.13,0.68±0.11;the Bax protein expression in these five groups were 1.00±0.07,2.90±0.21,0.58±0.07,1.93±0.11,2.44±0.09;the protein expression levels of Runx2 in these five groups were 1.00±0.05,0.42±0.02,1.97±0.06,0.87±0.09,0.22±0.03.The optical density values of mineralized nodules in these five groups after dissolution were 2.21±0.05,1.36±0.04,3.15±0.09,1.54±0.08,0.83±0.05.The above indicators:the difference between model group and the control group was statistically significant(P<0.05);the difference between the Pue pretreatment group and the model group was statistically significan

关 键 词：葛根素 氧化应激 骨髓间充质干细胞 核因子E2相关因子2 成骨分化 

分 类 号：R28[医药卫生—中药学]