基于FOXO1的抗氧化作用探讨六味地黄方对去卵巢大鼠及H_(2)O_(2)诱导的氧化损伤MC3T3⁃E1细胞的干预作用  

Intervention effect of Liuwei Dihuang Decoction on ovariectomized rats and H_(2)O_(2)⁃induced oxidative damaged MC3T3⁃E1 cells based on antioxidant effect of FOXO1

作　　者：陶乐维[1] 韩煦 陈清光[1] 徐隽斐[1] 陆灏[1] TAO Lewei;HAN Xu;CHEN Qingguang;XU Juanfei;LU Hao(Department of Endocrinology,Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)

机构地区：[1]上海中医药大学附属曙光医院内分泌科,上海201203

出　　处：《上海中医药杂志》2023年第7期13-20,共8页Shanghai Journal of Traditional Chinese Medicine

基　　金：上海市自然科学基金项目(19ZR1458300);上海市科委科研计划项目(21S21900700);上海市中医糖尿病专科联盟建设项目(ZY〔2018-2020〕-FWTX-4011);上海市临床重点专科项目(shslczdzk05401);上海市中医临床重点实验室项目(20DZ2272200);上海市浦东新区中医联合体建设项目(PDZY-2019-0601)。

摘　　要：目的观察六味地黄方对去卵巢(OVX)大鼠氧化应激相关指标、骨转换标志物及股骨叉头框蛋白O1(FOXO1)表达的干预作用,并探讨其能否通过FOXO1来发挥抗氧化作用及其作用机制。方法(1)动物实验。将雌性Wistar大鼠分为正常组、假手术组、模型组、补佳乐组、N-乙酰半胱氨酸(NAC)组、六味地黄方大剂量及小剂量组,药物干预8周后检测骨髓活性氧(ROS)、血清抗氧化指标[超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-PX)、还原型谷胱甘肽(GSH)]、骨转换标志物[骨碱性磷酸酶(BALP)、骨钙素(OCN)、血清I型胶原羧基末端肽(CTX-I)]及股骨FOXO1蛋白表达。(2)细胞实验。制备对照组含药血清和低、中、高剂量含药血清。①将胚胎成骨细胞前体细胞(MC3T3-E1)分为正常组、模型组、对照含药血清组,以及低、中、高剂量含药血清组和NAC组,检测各组细胞FOXO1蛋白表达。②沉默MC3T3-E1细胞的FOXO1基因表达,以Western blot法验证后,将细胞分为正常组、模型组、NAC组、高剂量含药血清组、siNC组、siFOXO1组、siFOXO1+高剂量含药血清组,检测各组细胞的增殖及ROS水平。③将MC3T3-E1细胞分为正常组、模型组、NAC组、对照含药血清组,以及高剂量含药血清组,检测各组细胞p-FOXO1蛋白表达。结果(1)动物实验:与假手术组比较,模型组SOD、CAT、GSH-PX、GSH、BALP、OCN水平均明显降低(P<0.05),CTX-I、ROS水平及FOXO1蛋白表达均明显升高(P<0.05);与模型组比较,六味地黄方大剂量组SOD、CAT、GSH-PX、GSH、BALP、OCN水平均明显升高(P<0.05),CTX-I、ROS水平及FOXO1蛋白表达均明显降低(P<0.05)。(2)细胞实验:①与正常组比较,经H_(2)O_(2)处理的MC3T3-E1细胞FOXO1蛋白表达明显增加(P<0.05);含药血清各剂量组FOXO1蛋白表达均低于模型组(P<0.05);②沉默MC3T3-E1细胞FOXO1基因后,siFOXO1组在48 h及72 h细胞增殖均明显低于模型组和siNC组(P<0.05),ROS�Objective To observe the intervention effect of Liuwei Dihuang Decoction on oxidative stress-related indices,bone turnover markers and the protein expression of FOXO1 in femoral in ovariectomized(OVX)rats.And to explore whether Liuwei Dihuang Decoction can exert antioxidant effect through FOXO1 and its mechanism.Methods(1)In animal experiment:Female Wistar rats were divided into normal group,sham-operated group,model group,Progynova group,N-acetylcysteine(NAC)group,Liuwei Dihuang Decoction high-and low-dose groups.Reactive oxygen species of bone marrow,serum antioxidant indices including SOD,CAT,GSH-PX,GSH and bone turnover markers including BALP,OCN,CTX-I,and the protein expression of FOXO1 in femoral were detected after 8 weeks of intervention.(2)In cell experiments,the decoction of Liuwei Dihuang was prepared and then the medicated serum of Liuwei Dihuang Decoction was collected,including medicated serum of control group,medicated serum groups with low-,medium-and high-dose.①MC3T3-E1 cells were divided into normal group,model group,control medicated serum group,low-,medium-and high-dose medicated serum group of Liuwei Dihuang Decoction and NAC group.Protein expression of FOXO1 in each group was detected to verify the results of animal experiment.②FOXO1 gene expression of MC3T3-E1 cells was silenced.After verified by Western blot,the cells were divided into normal group,model group,NAC group,high-dose medicated serum group of Liuwei Dihuang Decoction,siNC group,siFOXO1 group,and siFOXO1+high-dose group of Liuwei Dihuang Decoction medicated serum.The proliferation and ROS levels of cells in each group were detected.③MC3T3-E1 cells were divided into normal group,model group,NAC group,control medicated serum group and high-dose medicated serum group of Liuwei Dihuang Decoction,and the protein expression of p-FOXO1 in each group was detected.Results(1)In animal experiment:Compared with the sham operation group,SOD,CAT,GSH-PX,GSH,BALP and OCN levels were significantly decreased(P<0.05),while CTX-I,ROS levels a

关 键 词：骨质疏松症 六味地黄方 氧化应激 大鼠模型 经典名方 中药研究 

分 类 号：R285.5[医药卫生—中药学]