Effects Of Fluoride On Proliferated Activity And Expression Of Vascular Endothelial Growth Factor In Fibroblast

It is possible that fluoride can induce expression of osteogenic phenotype and enhance the potent osteogenesis function of FB. This may play an important role in the development of extraperiosteal calcification and ossification in skeletal fluorosis. In this paper, we studied the effects of fluoride on the proliferated activity of 3D cultured FB, and research the expression of VEGF and VEGF mRNA in both FB and OB from 2D and 3D sides.1. The effects of fluoride on the proliferated activity of 3D cultured FBWe cultured FB in eight F- dose (0,0.0001,0.0002,0.001,0.1,1,10,20mol/L) and at seven times (1,2,4,24,48,72,96h) via make fibroblasts populated collagen lattice (FPCL) use the rat tail collagen. The MTT method was used to detect the proliferated activity of FB. Results showed that fluoride increased the proliferated activity of FB at certain time (24,48,72h) to compare with the control group, but the results didn't reach the significant level of statistics; the others was decreased or obviously decreased. Though the effect was fairly slow and the degree was relatively feeble to contrast with it in 2D cultured FB, we think they are semblable. It can be concluded that FB have been proliferated under the role of fluoride.2. The expression of VEGF in 2D cultured FB and OB under the influence of fluorideBy using the methods of IHC and Elisa, we investigated the expression of VEGF in FB and OB following the treatment with fluoride. The IHC results showed that the expression of VEGF increased significantly (FB:10mg/L at 24h; .001mg/L at 48h; OB: 0.001,1,10mg/L at 48h ). The Elisa results showed that the expression of VEGF in culture medium supernatant of FB and OB step up obviously, too (FB: 0.1,1,10mg/L at 48h; OB: 0.001mg/L at 24h; 1mg/L at 72h ) . The results suggest that fluoride can stimulate the functions of FB and OB in the expression of VEGF, and the same tendency was found.RT-PCR method was used to study the expression of VEGF mRNA in FB and OB following the treatment with fluoride at 48h , we detected that there are small increased on the expression of VEGF mRNA (0.0001,10mg/L in FB; 0.0001mg/L in OB ). The tendency of it in two cells is semblable.The results above suggest that the expression of VEGF both in FB and OB are stimulated by fluoride, and they have the same change, it hint that FB have been transformed into OB under the influence of fluoride. 3. The expression of VEGF in 3D cultured FB under the influence of fluorideWe studied the expression of VEGF in 3D cultured FB in seven F- concentrates (0,0.0001,0.001,0.1,1,10,20mol/L) and at four times (4,24,48,72h) by using the methods of IHC. We found fluoride can stimulate the expression of VEGF in 3D FB, too (0.001,0.1,1 mg/L at 48h; 0.001,0.1 mg/L at 72h) . Results of Elisa showed that the expression of VEGF in culture medium supernatant have increased in different degree, more than it in 2D cells. Contrast to the control group, follows are changed significantly : 0.1,1,20 mg/Lat 2h, 0.001,0.1,1 mg/L at 24h, 0.1,1 mg/L at 48h and 72h . Result of RT-PCR tell us the expression of VEGF mRNA in 3D FB following the treatment with fluoride (0.1 mg/L) at 48h raise up notablely.Both VEGF and VEGF mRNA are expressed increase in 3D cultured cells more than they are expressed in 2D cultured cells. It implies application of fluoride in 3D FB is significative.In this paper, we studied the effects of fluoride on the proliferated activity of 3D cultured FB. Results showed that 3D cultured FB can be proliferated under the influence of fluoride. Contrast to 2D cultured cells, it develop slowly and have thinness level. It may because fluoride can't get in touch with cells. Meanwhile, we researched the expression of VEGF and VEGF mRNA in both FB and OB from 2D and 3D sides, and detected fluoride can stimulate the functions of FB and OB in the expression of VEGF, both in 2D and 3D cultured cells, and visibly in 3D cultured cells. But the exact mechanism needed more study. The same tendency in FB and OB was found, it consistent with our hypothesis that FB have been transformed into OB under the influence of fluoride. We've discovered fluoride can increase the expression of cbfa1 in FB in past years, and there is a report says excess expression of cbfa1 can stimuted excess expression of VEGF and VEGF mRNA, so we infer fluoride affect osteogenic phenotype of FB by contribution to excess expression of cbfa1. It point out a new way to study the osteogenic phenotype of FB under the influence of fluoride.