CAJ | 학술논문

目的研究大豆苷原( daidzein,DA)对成骨细胞雌激素受体(estrogen receptor,ER)和过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptor γ,PPARγ)表达的调节作用,并观察雌激素对这种调节的影响.方法小鼠成骨细胞MC3T3-E1体外低血清(α-MEM,含2％ FBS)培养,分别用0.1和10 μmol/L的DA处理,采用实时定量PCR或Western blot分析细胞ERα、ERβ和PPARγ的表达变化.加入浓度均为0.1μtmol/L的ER拮抗剂ICI182780或PPARγ拮抗剂GW9662,观察ER和PPARγ在DA调节中的作用.为观察雌激素的影响,采用无血清培养,在10 nmol/L 17β-雌二醇(E2)条件下研究DA对细胞受体表达的作用.结果　DA抑制体外培养成骨细胞ER表达,而刺激其PPARγ表达.0.1和10 μmol/L的DA分别下调ERα蛋白水平44％和38％ (P＜0.05),下调ERβ蛋白水平50％ (P＜0.05)和31％ (P＜0.05),上调PPARγ 74％和78％(P＜0.05).ICI182780可阻断DA对ERα转录水平的抑制作用,DA对成骨细胞ERβ mRNA水平的下调无统计学意义(P=0.087 4)；GW9662可阻断DA对PPARγ表达的上调作用,提示成骨细胞ER和PPARγ参与自身表达调节.在10 nmol/L 17β-雌二醇条件下,DA对无血清培养的成骨细胞ERα转录水平的抑制作用由28.0％～29.6％ (P＜0.05)增强至74.0％～82.8％(P＜0.01),其对ERβ表达的抑制作用也明显增强,而对PPARγ的上调作用几乎丧失.结论　DA可通过调节ERs和PPARγ等受体表达间接影响成骨细胞的药物反应,雌激素可明显影响DA的受体调节作用.DA对细胞受体表达的调节作用可能是其对成骨细胞时间相关双相调节的重要机制之一.
목적 연구대두감원( daidzein,DA)대성골세포자격소수체(estrogen receptor,ER)화과양화물매체증식물격활수체γ(peroxisome proliferator-activated receptor γ,PPARγ)표체적조절작용,병관찰자격소대저충조절적영향.방법 소서성골세포MC3T3-E1체외저혈청(α-MEM,함2％ FBS)배양,분별용0.1화10 μmol/L적DA처리,채용실시정량PCR혹Western blot분석세포ERα、ERβ화PPARγ적표체변화.가입농도균위0.1μtmol/L적ER길항제ICI182780혹PPARγ길항제GW9662,관찰ER화PPARγ재DA조절중적작용.위관찰자격소적영향,채용무혈청배양,재10 nmol/L 17β-자이순(E2)조건하연구DA대세포수체표체적작용.결과　DA억제체외배양성골세포ER표체,이자격기PPARγ표체.0.1화10 μmol/L적DA분별하조ERα단백수평44％화38％ (P＜0.05),하조ERβ단백수평50％ (P＜0.05)화31％ (P＜0.05),상조PPARγ 74％화78％(P＜0.05).ICI182780가조단DA대ERα전록수평적억제작용,DA대성골세포ERβ mRNA수평적하조무통계학의의(P=0.087 4)；GW9662가조단DA대PPARγ표체적상조작용,제시성골세포ER화PPARγ삼여자신표체조절.재10 nmol/L 17β-자이순조건하,DA대무혈청배양적성골세포ERα전록수평적억제작용유28.0％～29.6％ (P＜0.05)증강지74.0％～82.8％(P＜0.01),기대ERβ표체적억제작용야명현증강,이대PPARγ적상조작용궤호상실.결론　DA가통과조절ERs화PPARγ등수체표체간접영향성골세포적약물반응,자격소가명현영향DA적수체조절작용.DA대세포수체표체적조절작용가능시기대성골세포시간상관쌍상조절적중요궤제지일.
Objective To explore the effects of daidzein (DA) on the expressions of estrogen receptors (ER) and peroxisome proliferator-activated recepor γ (PPARγ) in osteoblasts and the influence of estrogen on these effects.Methods A mouse osteoblastic cell line MC3T3-E1 cultured in α-MEM containing 2％ FBS was treated by 0.1 and 10 μmol/L DA.ER antagonist ICI182780 and PPARγ antagonist GW9662 in 0.1 μmol/L was added as required,and an equivalent amount of phosphate buffer solution (PBS) was used as control.For the study on estrogen effect,the cells were treated by DA in the serum-free medium with or without 10 nmol/L 17β-estradiol (E2).The expressions of ERa,ERβ and PPARγ were determined by real-time RT-PCR and Western blot analysis,respectively.Results DA inhibited ER,expression but stimulated PPARγ expression in the cells at the concentration of 0.1 and 10 μmol/L.The down-regulation of ERα by DA could be blocked by ICI182780,whereas the up-regulation of PPARγ could be repressed by GW9662 in transcription levels.Furthermore,the inhibitory effect of DA on ERβ expression was markedly enhanced,while its stimulatory effect on PPARγ expression was almost lost in serum-free medium with 10 nmol/L 17βestradiol as determined by real-time RT-PCR.Conclusions Besides its direct roles in ERs and PPARγ mediated gene transcriptions,DA could exert indirect effect on cellular pharmacological responses by altering ER and PPARγ expressions.The predominant influence on receptors expression probably involved in the time-related biphasic effects of DA on osteogenesis,which was supposedly influenced by estrogen level.