| Objective: To observe the expression of silent information regulator(SIRT)in chronic fluorosis rat brain tissue in and fluorine treated SH-SY5 Y cells,and to explore its protective effect on experimental fluorosis neurological injury.Methods: 60 SD rats(equal number in both genders)were selected with the weight(100±20)g.After a week of adaptive feeding,they were randomly divided into three groups(10 in each with equal number of both genders),the time of exposing to fluorine was three and six months.In the control group,the rats were fed with tap water(less than 0.5 mg/l fluoride);the rats in the low and high fluoride groups were fed with drinking water containing 5.0 mg/l and 50.0 mg/l fluoride,respectively.All rats were fed with the same standard food containing no more than 0.6 mg/kg fluoride.SH-SY5 Y cells were cultured in vitro in six groups: Control group(Control),exposing to fluorine group(NaF),SIRT1 agonist group(RSV),exposing to fluorine + agonist group(NaF+ RSV),the SIRT1 inhibitor group(Suramin),exposing to fluorine + Suramin inhibitor group(NaF +Suramin).The effect of RSV and Suramin on the activity of SH-SY5 Y cells detected by CCK-8.After 3 and 6 months of establishing models,dental fluorosis of rats was examined,determination of fluoride in rat urine,serum and brain tissue by Fluoride selective electrode method.Morris water maze was used to detect the ability of learning and memory of rats.Hematoxylin-Eosin(HE)staining was used to observe the morphology of the brain.The protein and mRNA expressions of SIRT1 cultured in vitro for 48 hours and SIRT1,SIRT3,SIRT4,SIRT5 of rats brain tissues after 3 and 6 months of modeling,were detected by Western blotting and real-time PCR.After 3 and 6 months of modeling,the content of MDA,SOD and GSH-PX activity in rat serum,as well as MDA content and SOD activity of cells cultured in vitro were detected by kit.The apoptosis rate of SH-SY5 Y cells was detected by flow cytometry.Results: Rat experiment: No dental fluorosis was observed in the control group for 3 and 6 month,while dental fluorosis was observed in the low-fluorine group and the high-fluorine group,especially in the high-fluorine group.The urine,serum and brain tissue fluorine content of rats in the low-fluorine group and the high-fluorine group treated for 3 months and 6 months was significantly higher than that of control group,and high-fluorine group was significantly lower than that of low-fluorine group(P < 0.05).The escape latency time of rats in the high fluoride group with 3 months was higher than that in the control group(P<0.05),the number of times crossing the platform and stay time in platform quadrant were lower than that of the control group(P<0.05);while the rats in the low fluoride group showed no significant changes compared with the controls.The escape latency time of rats in the low fluoride and the high fluoride groups with 6 months were all higher than that of control group(P<0.05);the number of crossing the platform and stay time in platform quadrant were significantly lower than that of the control group.HE staining showed no significant difference in brain tissue morphology among every group of three and six months of the experimental period.The expression of protein and mRNA of SIRT1,SIRT3,SIRT4,SIRT5 in the hippocampus and cortex of rats fed with high fluoride for 3 months were significantly lower than that of control group(P<0.05),and whereas no significant changes from the rats in the low fluoride group;expression of protein and mRNA of SIRT1,SIRT3,SIRT4,SIRT5 in the hippocampus and cortex of rats fed with high fluoride were lower than those of the control group,while high fluoride group showed more obvious changes than low fluoride group(P < 0.05).The content of MDA in the serum of the rats in the fluorine group at 3 and 6 months was obviously higher than that in the control group,the activity of SOD and GSH-PX was lower than that in the control group,and in the high-fluorine group showed obvious changes than low-fluorine group(P < 0.05).Cell experiment showed:SH-SY5 Y cells were cultured in vitro,the result of CCK-8 showed that the activity of SH-SY5 Y cells was treated with 100 ?mol/L RSV and 400 ?g/mL Suramin for 24 hours was signficantly lower than those in the control group(P < 0.05).The expression of protein and mRNA of SIRT1 in the fluoride group,the SIRT1 inhibitor group,the fluoride and the SIRT1 inhibitor group were significantly lower than that of the control group(P < 0.05);the expression of protein and mRNA of SIRT1 in the SIRT1 agonist group were significantly higher than those in the control group,the fluoride and the SIRT1 agonist group(P < 0.05).The content of MDA in the fluoride group,the SIRT1 inhibitor group,the fluoride and the SIRT1 inhibitor group were significantly higher than those in the control group(P < 0.05);the content of MDA in the SIRT1 agonist group were significantly lower than those in the control group,the fluoride and the SIRT1 agonist group(P < 0.05).The activity of SOD in the fluoride group,the SIRT1 inhibitor group,the fluoride and the SIRT1 inhibitor group were significantly lower than those in the control group(P < 0.05);the activity of SOD in the SIRT1 agonist group were significantly higher than those in the control group,the fluoride and the SIRT1 agonist group(P < 0.05).The apoptosis rate of the fluoride group,the SIRT1 inhibitor group,the fluoride and the SIRT1 inhibitor group were significantly higher than those of the control group(P < 0.05);and the apoptosis rate of the SIRT1 agonist group was significantly lower than those of the fluoride and the SIRT1 agonist group(P < 0.05).Conclusion: The decrease of the expression of protein and mRNA of SIRT1,SIRT3,SIRT4,SIRT5 in rats brain tissues of excessive fluoride accumulation,it may be related to the decrease of the ability of learning and memory in rats with chronic fluorosis and the increase of oxidative stress levels in rats with chronic fluorosis.The decrease of the expression of protein and mRNA of SIRT1 of SH-SY5 Y resulted from nerve cells cultured in vitro treated by fluorine,and the protection of the nerve system was weakened.Using the SIRT1 agonist(RSV)can improve the nerve injury of nerve cells.SIRT may have neuroprotective effects against fluorotocicity. |
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