Effects of gene interference binding immunoglobulin protein on stimulation of osteoblast differentiation induced by fluoride
10.3760/cma.j.issn.2095-4255.2015.05.007
- VernacularTitle:基因干扰免疫球蛋白结合蛋白对氟刺激成骨细胞作用的影响
- Author:
Zhitao ZHAO
;
Chen YANG
;
Yan WANG
;
Guangsheng LI
;
Hui XU
- Publication Type:Journal Article
- Keywords:
Fluorides;
Osteoblast;
RNA interference
- From:
Chinese Journal of Endemiology
2015;34(5):335-339
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observe the effect of RNA interference binding immunoglobulin protein (BiP) on expression of bone markers and keytranscription factors in osteoblast exposed to fluoride.Methods MC3T3-E1 cells were used as osteoblast model in vitro.The cell viability was test with cell counting Kit-8 after cells were administrated with varying concentrations of fluoride [0.0 (control),0.1,1.0,2.0,4.0,8.0,16.0,20.0,32.0 and 64.0 mg/L] for different duration.Cells transfected with small interfering RNA (siRNA) BiP were exposed to fluoride (2.0,8.0 and 20.0 mg/L) for 2 days.Real-time PCR and Western blotting technique were used to determine the gene and protein levels of BiP.Meantime,the mRNA expression of bone markers and key transcription factors was investigated by real-time PCR.Results The difference of all viability in fluoride-dose groups was statistically significant exposed for 1,3,7 and 14 days (F =46.7,118.6,214.6,325.6,all P ＜ 0.05).Expression of BiP significantly increased in cells exposed to 20.0 mg/L compared to that of control (11.22 ± 3.25 vs.7.94 ± 1.31,P ＜ 0.05).The expression of alkaline phosphatase (ALP) elevated in cells exposed to 2.0 mg/L of fluoride (12.81 ± 3.62 vs.6.86 ± 2.13,P ＜ 0.01);conversely,it significantly reduced in cells exposed to 20.0 mg/L of fluoride (0.89 ± 0.17 vs.6.87 ± 2.14,P ＜ 0.01).Cells transfected with siRNA BiP significantly decreased the ALP expression in cells exposed to fluoride compared to that of cells only exposed to the same concentration of fluorine (12.81 ± 3.62 vs.7.43 ± 2.06;5.92 ± 2.38 vs.3.96 ± 0.21,all P ＜ 0.05).Cells transfected with siRNA BiP and administrated with 2.0 mg/L significantly reduced the osteocalcin expression (4.29 ± 0.99 vs.1.29 ± 0.86,P ＜ 0.01).Similarly,expression of runt-related transcription factor 2 (Runx2) significantly increased in cells exposed to 2.0 mg/L.However,expression of Runx2 significantly decreased in cells transfected with siRNA BiP and administrated with fluoride.Cells transfected with siRNA BiP significantly decreased the Runx2 expression in cells exposed to 2.0 mg/L and 20.0 mg/L compared to that of cells only exposed to the same concentration of fluorine (1.13 ± 0.22 vs.6.61 ± 0.48;0.02 ± 0.02 vs.1.50 ± 0.38,all P ＜ 0.01).As the downstream of Runx2,the expression of osterix in cells treated with different concentrations of fluoride was similar to that of Runx2.Conclusion BiP is not directly involved in the process of osteoblast differentiation induced by fluoride;instead,it affects the expression of bone markers and key transcription factors in osteoblast exposed to fluoride.
