| Objective:Fluoride is one of the most widely distributed elements in nature.It has a strong affinity for hard tissues and is easily deposited in teeth and bones.Fluoride has a dual effect on human health.The proper amount of fluoride is an essential trace element in human body,but long-term excessive intake can cause chronic systemic diseases characterized by dental fluorosis and skeletal fluorosis.Fluoride has a damaging effect on cartilage,which can cause chondrocyte necrosis,change cartilage proteoglycan metabolism,decrease collagen synthesis ability,and destructive cartilage enzyme activity.The growth plate is composed of chondrocytes and extracellular matrix(ECM).ECM includes collagen,proteoglycan(PG),elastin and structural glycoprotein,which are involved in chondrocyte proliferation,differentiation and migration.Li Guangsheng used fluoride to stain chondrocytes,and found that the concentration of fluorine was 0.2~1.4mmol/L to promote the synthesis of PG.When the concentration of fluoride was 2.0mmol/L,the formation of PG was inhibited.Yan Qunqun,Liu Ying and Pu Chunji found that the concentration of sodium fluoride at 200 mg/L could increase the rate of GAG synthesis in rat chondrocytes and increase the content.Glycosaminoglycan(GAG)includes hyaluronic acid(HA),4-chondroitin sulfate(C4S),6-chondroitin sulfate(C6S),keratan sulfate(KS),dermatan sulfate(DS),heparin(HP)And heparan sulfate(HS).Prince’s study found that the levels of total GAG,C4S and HA decreased with the increase of age,the content of DS remained unchanged,and the contents of C6S and DS were three and two times that of the control group,respectively.Lucas found that fluoride increased the conversion of glycosaminoglycans in the callus and backbone areas of rats.The cartilage osteogenesis is regulated by Ihh/PTHrP,FGFs and other signaling pathways.These signaling pathways together with the cartilage matrix regulate the intrachondral osteogenesis process.To this end,we established a rat model of fluorosis to observe the effect of fluoride on the content of GAG in the growth plate cartilage matrix,combined with the expression of related proteins in the growth plate chondrocytes,to explore whether fluoride inhibits cartilage by affecting cartilage matrix and related signaling pathways.The internal osteogenesis process provides clues for elucidating the mechanism of fluoride inhibition of long bone growth and provides a basis for the prevention and treatment of skeletal fluorosis.Methods:Thirty-two male SD rats(50~70g)were randomly divided into 4 groups and distilled water containing 0,50 ppm,100 ppm and 150 ppm F~-was used for feeding for 12 weeks.After 12 weeks,the rats were sacrificed.The body weight,body length,femur length,tibia length and blood fluoride content were measured.Paraffin sections were made on the left tibia and fluoride content was measured on the right tibia.The contents of HA,CS,HSPGs and KS were detected by aliquots of paraffin sections.The expressions of related proteins(PTHrP,Ihh,FGFR3,STAT1)in chondrocytes of growth plates were observed by immunohistochemistry.Results:1.The effect of fluoride on general condition,body weight,bone length,blood fluoride content and bone fluoride content in rats:the weight of rats in each fluoride group was lower than that in the control group,and the difference in 150 ppm F~-group was statistically significant(P<0.05);The length of the rats in the fluoride-treated group was lower than that in the control group,and the difference was statistically significant in the 150 ppm F~-group(P<0.05).The length of the tibia and femur in the fluoride-treated group was lower than that in the control group,and the difference in the 150 ppm F~-group was statistically significant(P<0.05);the content of fluoride and bone fluoride in each fluoride group was higher than that in the control group,and the difference of fluoride group was statistically significant(P<0.05).2.The effect of fluoride on the content of several main components in the growth plate of rat growth plate:the distribution of HA staining and the color depth in the growth plate of each fluoride group were not different from those in the control group;the distribution of CS staining in the growth plate of each fluoride group There was no significant difference in color depth compared with the control group;there was no KS expression in the growth plates of each group;the content of HS in the growth plates of each fluoride group was higher than that of the control group.3.The effect of fluoride on the expression of FGFR3 in rat growth plate chondrocytes:The expression of FGFR3 in the nucleus of the growth plate and the pre-mast zone of the rats in the fluoride-treated group increased,and the expression of 100 ppmF~-group and 150 ppm F~-group was significantly higher.In the control group,the difference was statistically significant(P<0.05).4.The effect of fluoride on STAT1 in rat growth plate chondrocytes:the proliferation of growth plate and the expression of STAT1 in the anterior hypertrophic chondrocytes of rats exposed to fluoride increased,and the expression of 100 ppmF~-group and 150 ppmF~-group was significantly higher than that of the control group.Statistically significant(P<0.05).5.The effect of fluoride on the expression of PTHrP in the growth plate chondrocytes of rats at 12 weeks:the proliferation of the growth plate and the expression of PTHrP in the hypertrophic chondrocytes increased in the fluoride-treated group.The difference between the fluoride-treated group and the control group was statistically significant.(P<0.05).6.The effect of fluoride on the expression of Ihh in rat growth plate chondrocytes:The proliferation of growth plate and the expression of Ihh in the anterior hypertrophic chondrocytes of the fluoride-induced group showed a downward trend,and the difference between the 100 ppmF~-group and the 150 ppm F~-group was statistically significant(P<0.05).Conclusion:1.High fluoride reduced the body weight,body length,length of the tibia and length of the femur,and increased blood fluorine and bone fluoride.2.The content of HS in the growth plate of fluorosis rats was abnormally increased.3.The expression of Ihh protein in the growth plate of fluorosis rats decreased,and the expression of PTHrP protein increased.Fluorine inhibits the normal proliferation and differentiation of chondrocytes by inhibiting the Ihh/PTHrP negative feedback loop.4.The expression of FGFR3 protein in the growth plate of fluorosis rats was increased,and the downstream STAT1 pathway was activated to inhibit the proliferation of chondrocytes. |
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