摘要
目的探讨不同浓度吡格列酮(PIO)对正常糖浓度培养下MC3T3-E1小鼠早期成骨细胞增殖、凋亡及功能蛋白分泌表达的影响。方法体外培养MC3T3-E1细胞,分为正常对照组、不同浓度PIO干预组(5μmol/L、10μmol/L、20μmol/L、40μmol/L),分别干预24、48 h。CCK-8检测细胞增殖,流式细胞仪检测细胞凋亡率,RIA法和ELISA法分别检测相关功能蛋白骨钙素(OCN)、碱性磷酸酶(ALP),RT-PCR检测过氧化物酶体增殖物激活受体γ(PPARγ)、成骨因子runt相关基因2(Runx2)及骨形成蛋白-2(BMP-2)mRNA表达水平。结果 (1)与对照组比较,MC3T3-E1细胞增殖能力在5μmol/L PIO干预组增加显著(P<0.05)、在PIO大于5μmol/L～10μmol/L干预组则降低(P<0.05)。干预时间由24 h延长至48 h,各组细胞增殖能力呈不同程度增加,但在20、40μmol/L PIO干预的两组增加无统计学差别。(2)对照组及各PIO浓度干预干预24 h,细胞凋亡率分别为:1.97%、0.43%、13.0%、48.30%、81.00%;(3)随着PIO浓度从0～40μmol/L范围内的增加,MC3T3-E1细胞的PPARγmRNA表达水平呈增加趋势(P<0.05);Runx2 mRNA表达量在5μmol/L PIO浓度组时较对照组增加,在剂量较高时(PIO≥10μmol/L)随浓度升高表达下降。(4)细胞功能蛋白ALP、OCN的分泌及BMP-2的表达在5μmol/L PIO浓度组时最高,PIO≥10μmol/L时,上述蛋白量逐渐下降(P<0.05);随干预时间从24～48 h延长,各PIO浓度干预组ALP以及BMP-2量增加(P<0.05),而OCN无显著改变(P>0.05)。结论 PIO对正常糖浓度培养的MC3T3-E1细胞具有双向作用:低浓度促进细胞增殖,高浓度则促进凋亡。PPARγ适当激活可促使成骨细胞通过Runx2增加功能蛋白的合成;过度激活,则表现出细胞毒性。
Objective To investigate the effects of pioglitazone （PIO） on proliferation, apoptosis and expressions of functional proteins in mouse preosteoblastic cells MC3T3-E1 exposed to normal glucose concentration. Methods MC3T3-EI cells were cultured and divided into control group, PIO administration group with different PIO concentrations （5, 10, 20 and 40μmol/L, respectively） and were exposed for 24 or 48 hours. Cell proliferation was tested with CCK-8. Cell apoptosis was detected using quantitative flow cytometry. The secretions of osteocalcin （OCN） and alkaline phosphatase （ALP） were measured using RIA or ELISA, respectively. The mRNA expressions of peroxisome proliferator-activated receptor gamma （PPARγ） , osteogenic factor runt related gene 2 （Runx2） , and bone morphogenetic protein-2 （BMP-2） was detected using semi-quantitative RT-PCR. Results （1） Compared to the control group, cell proliferation was promoted in PIO 5 μmol/L group （P 〈 0.05 ）. In cultures administered with PIO concentration higher than 5-10μmol/L, cell proliferation was decreased significantly compared to that in control group （P 〈 0. 05）. Extending the interfering time from 24 h to 48 h, cell proliferation in each group increased in a certain extent; however the increase was not statistically significant in 20 and 40 μmol/L PIO groups. （2） In cultures of control and PIO groups exposed for 24 h, the apoptotic rates were 1.97% , 0. 43% , 13.0% , 48.30% , and 81.00% , respectively. （3） In cultures with PIO concentration from 0 to 40 μmol/L, PPART mRNA expression levels of the cell showed a dose-dependent increase （P 〈 0. 05 ）. Runx2 mRNA expression increased in PIO 5μmol/L group, but it decreased in groups with PIO doses higher than 10 μmol/L. （4） The secretions of ALP, OCN, and the expression of BMP-2 were the highest in PIO 5 μmol/L group. Those were decreased dose-dependently when PIO concentrations were more than 10 μmol/L （P 〈 0. 05 ）. Expressions of ALP and BMP-2 in
出处
《中国骨质疏松杂志》
CAS
CSCD
2011年第2期96-101,共6页
Chinese Journal of Osteoporosis
基金
福建省自然科学基金资助项目(2009J01135)
