六味地黄丸含药血清通过诱导自噬对氧化应激状态下成骨细胞增殖的影响  

Effect of Liuwei Dihuang pill-containing serum on osteoblast proliferation under oxidative stress through induction of autophagy

作　　者：郭澜 李莉 葛继荣 黄景文 柴昊 谢丽华 陈娟 许鹏超 GUO Lan;LI Li;GE Jirong;HUANG Jingwen;CHAI Hao;XIE Lihua;CHEN Juan;XU Pengchao(Fujian Academy of Chinese Medical Sciences,Fuzhou 350003,China;Fujian Key Laboratory of Integrated Traditional Chinese and Western Medicine for the Prevention and Treatment of Osteoporosis,Fuzhou 350003,China)

机构地区：[1]福建省中医药科学院,福建福州350003 [2]福建省中西医结合防治骨质疏松重点实验室,福建福州350003

出　　处：《中国骨质疏松杂志》2023年第1期11-16,共6页Chinese Journal of Osteoporosis

基　　金：国家自然科学基金项目(81873323);福建省自然科学基金(2020J01725);福建省卫生健康青年科研课题(2019-1-76)。

摘　　要：目的观察六味地黄丸含药血清对氧化应激状态下MC3T3-E1细胞增殖、活性氧及自噬水平的影响。方法用过氧化氢(H 2O 2)模拟氧化应激状态后采用不同浓度的六味地黄丸含药血清干预MC3T3-E1细胞,CCK-8法检测细胞增殖情况;通过细胞活性氧(reactive oxygen species,ROS)检测细胞ROS水平;qRT-PCR检测自噬相关基因Beclin 1、LC3B mRNA表达情况;蛋白印迹法检测自噬相关蛋白Beclin 1、LC3B表达情况。结果①CCK-8结果显示,H 2O 2会抑制成骨细胞的增殖,当其浓度为0.10 mmol/L时具有相对较小程度的抑制增殖效果;六味地黄丸含药血清可以促进氧化应激状态下成骨细胞的增殖,且当体积浓度为20%时促进增殖的效果最好。②细胞活性氧检测结果显示,六味地黄丸含药血清组ROS水平降低(P<0.05)。③qRT-PCR结果显示,六味地黄丸组Beclin 1、LC3B mRNA表达上调(P<0.05),且该作用会被自噬抑制剂所抑制。④蛋白印迹法结果显示,六味地黄丸组Beclin 1蛋白表达上调(P<0.05),蛋白LC3Ⅱ/Ⅰ比值增高(P<0.05)。结论六味地黄丸含药血清可以促进氧化应激状态下成骨细胞增殖、降低细胞ROS水平,其机制可能与诱导细胞自噬相关。Objective To observe the effects of Liuwei Dihuang pill-containing serum on the proliferation and reactive oxygen species and autophagy levels of MC3T3-E1 cells under oxidative stress.Methods MC3T3-E1 cells were intervened with different concentrations of Liuwei Dihuang pill-containing serum after simulating oxidative stress with hydrogen peroxide(H 2O 2).Cell proliferation was detected with CCK-8 method.Cell ROS levels were detected with cell reactive oxygen species(ROS).mRNA expressions of autophagy-related genes Beclin 1 and LC3B were detected with qRT-PCR.The expressions of autophagy-related proteins Beclin 1 and LC3B were detected with Western blotting.Results(1)CCK-8 result showed that H 2O 2 inhibited the proliferation of osteoblasts,and it had a relatively small inhibitory effect when its concentration was 0.10 mmol/L.Liuwei Dihuang pill-containing serum promoted the proliferation of osteoblasts under oxidative stress,and the best effect was achieved when the volume concentration was 20%.(2)The result of cell ROS detection showed that the ROS level in Liuwei Dihuang pill-containing serum group decreased(P<0.05).(3)The result of qRT-PCR showed that Beclin 1 and LC3B mRNA expressions were up-regulated in the Liuwei Dihuang pill group(P<0.05),and the effect was inhibited by autophagy inhibitors.(4)Western blotting result showed that expression of Beclin 1 was up-regulated and LC3II/Ⅰratio was increased(P<0.05).Conclusion Liuwei Dihuang pill-containing serum promotes osteoblast proliferation and reduces cellular ROS levels under oxidative stress.The mechanism may be related to the induction of cellular autophagy.

关 键 词：六味地黄丸 成骨细胞 氧化应激 自噬 

分 类 号：R681[医药卫生—骨科学] R285.5[医药卫生—外科学]