氟对大鼠成骨细胞Runx2表达的影响

Influence of fluoride on Runx2 mRNA and protein expression in rat osteoblasts

目的研究氟对体外培养大鼠乳鼠成骨细胞中Runx2表达的影响。方法体外培养Wistar大鼠乳鼠成骨细胞，按染氟剂量分为0（对照组）、1、2、4mg／L4个水平，分别在48、72h后，提取RNA，采用反转录聚合酶链反应（RT—PCR）方法检测Runx2 mRNA表达；采用酶联免疫吸附试验（EUSA）方法检测培养液上清中的Runx2蛋白质。结果Wistar大鼠乳鼠成骨细胞在体外染氟培养48h后，1、2、4mg／L染氟水平下Runx2的mRNA表达（0．613±0．055、0．773±0．070、0．775±0．070）与对照组（0．482±0．043）比较，差异均有统计学意义（P〈0．05）；在染氟培养72h后，1、2、4mg／L染氟水平和对照组的Runx2 mRNA表达量分别为0．969±0．048、1．229±0．061、1．255±0．063、0．724±0．036，任意两组间比较，差异均有统计学意义（P〈0．05）。各染氟组Runx2 mRNA表达，72h组均高于48h组（P〈0．05）；染氟剂量和染氟时间有交互作用（F=189．20，P〈0．05）。在染氟培养48h后，1、2、4mg／L染氟水平下的Runx2蛋白质表达（0．141±0．007、0．143±0．008、0．143±0．011）与对照组（0．129±0．012）比较，差异有统计学意义（P〈0．05）；在染氟培养72h后，1、2、4mg／L染氟水平下的Runx2蛋白质表达（0．156±0．014、0．168±0．018、0．162±0．010）与对照组（0．137±0．016）比较，差异有统计学意义（P〈0．05）；各染氟剂量Runx2蛋白质表达，72h组均高于48h组（P〈0．05），染氟剂量和染氟时间之间无交互作用（F=2．22，P〉0．05）。结论氟可促进成骨细胞Runx2的mRNA和蛋白质表达，与染氟的剂量以及作用时间有关。

Objective To study the influence of fluoride on the expression of Runx2 in suckling rat osteoblasts. Methods Osteoblasts obtained from calvarium of suckling Wistar rats were cultured in the media supplemented with NaF at different doses（0, 1,2 and 4 mg/L）, and Runx2 mRNA expression and protein expression were evaluated by RT-PCR and ELISA, respectively. Results Runx2 mRNA expression in suckling rat osteoblasts cultured in vitro significantly increased after exposure to NaF for 48 h at different doses （0.613 ± 0.055, 0.773 ± 0.070 and 0.775 ± 0.070 for 1,2 and 4 mg/L,respectively） compared to the control （0.482 ± 0.043,P 〈 0.05）. Runx2 mRNA expression further increased after 72 h exposure to NaF（0.969 ± 0.048,1.229 ± 0.061,1.255 ± 0.063 for 1,2 and 4 mg/L, respectively）,which is significantly higher than the control（0.724 ± 0.036,P 〈 0.05） and corresponding groups at 48 h. NaF doses and exposure time exhibited a significant synergistic effect on Runx2 mRNA expression （P 〈 0.05）. Similarly, NaF also enhanced Runx2 protein expression in suckling rat osteoblasts cultured in vitro. Significant differences were observed between groups exposed to NaF （1,2 and 4 rag/L） and control at 48 h post-exposure （0.141 ± 0.007, 0.143 ± 0.008, 0.143 ± 0.011 vs 0.129 ± 0.012, P 〈 0.05） as well as 72 h post-exposure（0.156 ± 0.014, 0.168 ± 0.018, 0.162 ± 0.0100 vs 0.137 ± 0.016, P〈 0.05）. In addition, Runx2 protein expression at 72 h post-exposure was significantly higher than that at 48 h. Conclusions The results suggested that NaF could increase Runx2 expression in suckling rat osteoblasts with a synergistic effect between the doses and exposure time.