摘要
目的 探讨氟暴露对于人成骨细胞系Saos-2细胞(简称Saos-2细胞)微小RNA(miR)-200c表达及其靶点的影响。方法 采用DMEM/F-12培养液,体外培养Saos-2细胞。按培养液中氟化钠(NaF)剂量将Saos-2细胞分为对照组(0 mg/L)、染氟组(4 mg/L),染氟培养48 h后收集细胞。采用实时荧光定量PCR(qRT-PCR)检测Saos-2细胞中miR-200c表达,碱性磷酸酶(ALP)、骨钙素(BGP)及miR-200c作用靶点人第10号染色体缺失磷酸酶(PTEN)、双特异性磷酸酶(DUSP1)mRNA的表达;采用蛋白免疫印迹法(Western blot)检测PTEN、DUSP1蛋白表达。结果 染氟组Saos-2细胞中ALP、BGP mRNA和miR-200c表达(23.60 ± 1.87、9.41 ± 0.94、8.61 ± 0.26)高于对照组(1.00 ± 0.11、1.00 ± 0.07、1.00 ± 0.12),组间比较差异有统计学意义(t = - 24.084、- 18.388、- 8.687,P均 〈 0.05);PTEN和DUSP1 mRNA表达(0.63 ± 0.02、0.38 ± 0.02 )低于对照组(1.02 ± 0.24、1.02 ± 0.24),组间比较差异有统计学意义(t = 3.327、5.454,P均 〈 0.05)。染氟组Saos-2细胞PTEN、DUSP1蛋白表达(1.19 ± 0.10、0.83 ± 0.07)低于对照组(1.81 ± 0.14、1.44 ± 0.25),组间比较差异有统计学意义(t = 6.250、4.171,P均 〈 0.05)。结论 氟暴露能够使Saos-2细胞miR-200c表达水平升高,通过miR-200c作用于PTEN和DUSP1,下调PTEN、DUSP1 mRNA与蛋白表达水平。
Objective To investigate the effect of fluoride exposure on expression of miRNA(miR)-200c and its target in human osteoblast Saos-2 cells. Methods Saos-2 cells were cultured in DMEM/F-12 medium and treated with fluoride (sodium fluoride, NaF). There were two groups including: control group (0 mg/L) and fluoride group (4 mg/L). Cells were harvested after 48 hours of culture with fluoride. The expression of miR-200c, the mRNA of alkaline phosphatase (ALP), osteocalcin (BGP), the target phosphatase and tensin homolog deleted on chromosome ten (PTEN) and dual-specific phosphatase 1(DUSP1) of miR-200c was detected by qRT-PCR. The protein expression of PTEN and DUSP1 was detected by Western blotting. Results The expressions of ALP, BGP mRNA and miR-200c in Saos-2 cells in the fluoride group (23.60 ± 1.87, 9.41 ± 0.94, 8.61 ± 0.26) were higher than those in the control group (1.00 ± 0.11, 1.00 ± 0.07, 1.00 ± 0.12). The differences were statistically significant (t = - 24.084, - 18.388, - 8.687, P 〈 0.05). The mRNA expressions of PTEN and DUSP1 in the fluoride group (0.63 ± 0.02, 0.38 ± 0.02) were lower than those in the control group (1.02 ± 0.24, 1.02 ± 0.24). The differences were statistically significant (t = 3.327, 5.454, P 〈 0.05). The protein expressions of PTEN and DUSP1 in Saos-2 cells in the fluoride group (1.19 ± 0.10, 0.83 ± 0.07) were lower than those in the control group (1.81 ± 0.14, 1.44 ± 0.25). The differences were statistically significant (t = 6.250, 4.171, P 〈 0.05). Conclusion Exposure to fluorine may increase the expression of miR-200c in Saos-2 cells, and fluorine may act on PTEN and DUSP1 through miR-200c, downregulates the mRNA and protein expression levels of PTEN and DUSP1.
作者
王红格
姜宇婷
闫盼
邵汉文
卜野
高彦辉
孙殿军
Wang Hongge, Jiang Yuting, Yan Pan, Shao Hanwen, Bu Ye, Gao Yanhui, Sun Dianjun(1Institute for Endemic Fluorosis Control, Center for Endemic Disease Control, Chinese Center for Disease Control and Prevention, Harbin Medical University, Harbin 150081, China ;2 Heilongjiang Provincial Key Laboratory of Trace Elements and Human Health, Harbin 150081, China)
出处
《中华地方病学杂志》
CAS
CSCD
北大核心
2018年第5期366-369,共4页
Chinese Journal of Endemiology
基金
国家自然科学基金(81673110)
黑龙江省杰出青年基金(JC2015018)