Effect On The Proliferation Of Osteoblasts Cultured In Vitro And The Expression Of TM9SF1 Gene After Exposed To Fluoride

Objective: This project explored proliferating of osteoblasts of mice on the basis of different doses of fluoride after we developed the culture method of osteoblasts in vitro;detected influences of TM9SF1 proteins and distribution of TM9SF1mRNA after fluoride worked; checked the change of expression quantity of TM9SF1mRNA of osteoblasts cultured in vitro. Methods: We improved the culture method of osteoblasts of mice in vitro, and identified the origination of osteoblasts by using AKP and calcium nodes dyeing. There are five groups in experiment according to different doses of fluoride: 0, 2.5,5.0,10.0,20.0mg/L. And there are 10 groups in experiment according todifferent doses of MTT:0,2.5,5.0,10.0,20.0,40.0,60.0,80.0,100.0,120.0 mg/L. We observed the curves of the proliferation of osteoblasts when the doses increased. We detected the distribution of TM9SF1 mRNA of osteoblasts exposed to fluoride by ISH; We detected the quantity of TM9SF1 mRNA of osteoblasts exposed to fluoride by RT-PCR; We got quantity changes of TM9SF1 mRNA of osteoblasts in human exposed to fluoride by Real-time PCR. Results: The culture method that we developed in experiment made osteoblast stably in vitro. And the experiment demonstrated that fluoride influenced the proliferation of osteoblasts: Sodium fluoride significantly increased the proliferation of osteoblasts in low doses and decreased it in high doses. There was positive expression of TM9SF1 gene in all groups after exposed to fluoride, but positive degree and area in fluoride groups were stronger and bigger than control group. Calculating the numbers of positive osteoblasts showed that it had statistic difference between fluoride groups and control one. We discovered that the numbers was higher than control group and other groups which were lower than control group under the certification dose of 2.5mg/L through TM9SF1 mRNA by RT-PCR.There appeared TM9SF1 mRNA in all osteoblasts of human cell, but fluoride influenced the quantity of TM9SF1 mRNA in human cell: the quantity were highest under the doses2.5, 5.0mg/l and 1.04 and 1.03 times of control group, respectively. Conclusions: Fluorine of low dose improved the proliferation of osteoblasts, while high dose showed depressant effect. Fluoride had an effect on the proteins expression quantity of TM9SF1mRNA, namely, low dose improved the proteins expression and high doses weaken the expression. We could conclude, that is to say, that there were the same tendancy in the gene expression and quantity of TM9SF1mRNA and the proliferation of osteoblasts. Maybe the reason of improvement was that fluoride make TM9SF1 exist in the osteoblasts. Surely we should continue to do more research to prove it.