CAJ | 학술논문

目的：了解雌二醇与铁对小鼠原代成骨细胞活性的影响及活性氧（ ROS）的作用。方法提取新生小鼠颅骨原代成骨细胞，经差速贴壁法纯化后传代，选取3～4代进行实验，随机分为对照组，枸橼酸铁铵（ FAC）组和FAC＋17β-雌二醇（ FAC ＋E2）组，分别用2.5μmol/L FAC和10 nmol/L E2干预7 d后收集细胞，用CCK-8法检测各组细胞增殖情况；用碱性磷酸酶（ ALP）活性试剂盒检测各组细胞碱性磷酸酶活性；荧光定量PCR （ Q-PCR）检测成骨相关基因（ Runx2、 osterix、 Bglap、 IBSP）表达；荧光酶标仪检测各组ROS水平并在荧光显微镜下观察。结果细胞活性氧水平检测结果显示， FAC组活性氧水平明显高于其余各组， FAC＋E2组与FAC组相比ROS水平降低（ P＜0.05）； FAC干预后细胞增殖能力及ALP活性明显下降（ P＜0.05）， FAC＋E2组细胞增殖能力较FAC 组上升（ P＜0.05）， ALP活性上升，但差异无统计学意义（ P＞0.05）； Q-PCR结果显示，与对照组相比， FAC组Runx2、 osteorix表达水平明显下降（ P＜0.05）， FAC＋E2组与FAC组相比，各基因表达水平均显著升高（均P＜0.05）。结论 FAC可能通过升高ROS水平抑制成骨细胞生物活性；雌二醇可能通过清除FAC诱导生成的ROS下调该抑制作用。
목적：료해자이순여철대소서원대성골세포활성적영향급활성양（ ROS）적작용。방법제취신생소서로골원대성골세포，경차속첩벽법순화후전대，선취3～4대진행실험，수궤분위대조조，구연산철안（ FAC）조화FAC＋17β-자이순（ FAC ＋E2）조，분별용2.5μmol/L FAC화10 nmol/L E2간예7 d후수집세포，용CCK-8법검측각조세포증식정황；용감성린산매（ ALP）활성시제합검측각조세포감성린산매활성；형광정량PCR （ Q-PCR）검측성골상관기인（ Runx2、 osterix、 Bglap、 IBSP）표체；형광매표의검측각조ROS수평병재형광현미경하관찰。결과세포활성양수평검측결과현시， FAC조활성양수평명현고우기여각조， FAC＋E2조여FAC조상비ROS수평강저（ P＜0.05）； FAC간예후세포증식능력급ALP활성명현하강（ P＜0.05）， FAC＋E2조세포증식능력교FAC 조상승（ P＜0.05）， ALP활성상승，단차이무통계학의의（ P＞0.05）； Q-PCR결과현시，여대조조상비， FAC조Runx2、 osteorix표체수평명현하강（ P＜0.05）， FAC＋E2조여FAC조상비，각기인표체수평균현저승고（균P＜0.05）。결론 FAC가능통과승고ROS수평억제성골세포생물활성；자이순가능통과청제FAC유도생성적ROS하조해억제작용。
Objective To explore the effects of iron and estradiol on the biological activity of osteoblasts and the influence of reactive oxidative stress ( ROS) on this process.Methods Primary osteoblasts, extracted from suckling mice's cranium and purified with differential adhesion method , were divided into 3 groups: control group, FAC group, FAC+E2 group.Cells were intervened by FAC and estradiol .Harvested cells after 7d and then detected cell proliferative capacity using CCK-8 kit.Runx2, osterix, Bglap, and IBSP mRNA expression were detected by q-PCR.Alkaline phos-phatase ( ALP) activity was measured using ALP kit .ROS level was detected using a fluorescence microplate reader .Re-sults The level of ROS in FAC group was significantly higher than that in control group ( P<0.05 ) , and level in FAC+E2 group were lower than that in FAC group (P<0.05).Cell proliferative capacity and alkaline phosphatase activity were reduced with the treatment of FAC ( P<0.05 ) , while estradiol improved the inhibition of cell proliferative capacity and alkaline phosphatase activity induced by iron accumulation .Q-PCR results showed that FAC decreased Runx 2 and osteorix mRNA expression (P<0.05).Runx2, osterix, IBSP, and Bglap mRNA expression were increased in FAC +E2 group, comparing to those in FAC group ( P<0.05 ) .Conclusion Iron accumulation might increase ROS level and then reduce the biological activity of osteoblasts .This process might be inhibited by estradiol .