The Effect Of Zinc On The Expression Of C-fos And C-jun In Spermatogenic Cell In Rats Exposed To Fluorine

Skeletal fluorosis and dental fluorosis have been focused on the biological research of fluorine in the early days.The damage of fluorine to non-skeletal has been the focus in the research since 1980s,and more and more attention has been paid on the damage of fluorine on the reproductive system especially in recent years.Now,relevant research of fluorous damage on reproductive system have been focused on the damage testis,epididymis and the change of sperm quality,But there were few research on the antogonism of tluorine.Zinc which is in the bone and sperm play an important role on the development of reproductive system.Zinc is an essential component of CuZn-SOD,it can increase its enzymatic activity,induce the increase of MT,and accordingly play its anti-lipid peroxidative role.CuZn-SOD can inhibit the absorption of fluorine in intestinal tract and its accumulation in skeleton,and then lower the burden of fluorine in the body.There are lots of researches about the antagonism of fluorine on the reproductive system,but the mechanism is not certain and need to research deeply.This research is about the antagonism of zinc to the fluorous damage on reproductive system and its mechanism, using the four weeks male rats exposed to the fluorine and observe the impact of appropriate amount zinc to the c-fos,c-jun protein expression level.1.Materials and Methods1.1 dose grouping Fifty healthy,four weeks old,male rats weighted 60-80 grams were purchased from the animal experimental research center of Henan province. They were reared in room temperature(20-24℃),relative 30%-60%humidity,12 hours illumination,freely eating and drinking.After no abnormalities were observed, the fifty rats were allocated into five groups randomly,and ten rats were allocated in every group.The grouping and dose of narcotics were like that:intragastric administration by mouth,give demineralized water to the control group rats,25mg·kg^-1·d NaF to the fluorine treatment group,25mg·kg^-1·d NaF and 4mg·kg^-1·d ZnSO4 to the fluorine and low-dose zinc treatment group,25mg·kg^-1·d NaF and 20mg·kg^-1·d ZnSO4 to the flourine and middle-dose zinc treatment group,25mg·kg^-1·d NaF and 50mg·kg^-1·d ZnSO4 to the fluorine and high-dose zinc treatment group.Then observe itstoxic reaction.1.2 detection index After 6 weeks of narcotics used in every group rats,Weighing,anesthetize and put them to death,both testis and epididymis were dissected out and weighted for calculating their organic coefficient,and then proceed Sperm Count,determination of sperm activity ratio and the analysis of teratospermia.Fluorous content in testis was survey and evaluated by the method of fluorinion selective electrodes.The levels of serum testosterone(T) were examined by radio-immunity.The levels of LDH in testis were examined by colorimetric technique kit.The C-fos and c-jun protein expression level in germ cells were measured by immunohistochemistry.1.3 statistical analysis One-way analysis of variance,t-test and non-parametric tests were used according to the data type and characteristics of data distribution.There was significant difference when p was less than 0.05.2 Results2.1 Rats body weight,testis and epididymis weight and their organic coefficient:①Rats body weight:Significant difference was observed between control group and fluorine and high-dose zinc treatment group(P＜0.05);②testis weight and its organic coefficient: testis weight and its organic coefficient in fluorine and low-dose zinc treatment group and fluorine and high-dose zinc treatment group was significantly higher than in control group(P＜0.05);③epididymis weight and its organic coefficient:the epididymide organic coefficient in fluorine and middle-dose zinc treatment group was significantly higher than that in control group(P＜0.05).2.2 Rats Sperm Count,sperm activity ratio and malformation ratio2.2.1 Rats Sperm Count①Comparison of the sperm counts in fluorine and every-dose zinc treatment groups and fluorine treatment group:the sperm counts in all the fluorine and low-dose zinc treatment group,fluorine and middle-dose zinc treatment group and fluorine and high-dose zinc treatment group were significantly higher than that in fluorine group(P＜0.05);②Comparison of the sperm counts in the groups treated by fluorine and fluorine combined with zinc and control group:the sperm counts in all the fluorine and low-dose zinc treatment group,fluorine and middle-dose zinc treatment group,fluorine and high-dose zinc treatment group and fluorine group were significantly lower than that in control group(P＜0.05).2.2.2 Sperm activity ratio①Comparison of the sperm activity ratio in fluorine and every-dose zinc treatment groups and fluorine treatment group:the sperm activity ratio in all the three fluorine combined with zinc treatment groups were significantly higher than that in fluorine group(P＜0.05);②Comparison of the sperm activity ratio in the groups treated by fluorine and fluorine combined with zinc and control group:the sperm activity ratio in the fluorine and low-dose zinc treatment group, fluorine and high-dose zinc treatment group and fluorine group were significantly lower than that in control group(P＜0.05).However,no significant difference of the sperm activity ratio was found in the fluorine moderate-dose zinc treatment group and control group(P＞0.05).2.2.3 Sperm malformation ratio①Comparison of the sperm malformation ratio in fluorine and every-dose zinc treatment groups and fluorine treatment group:the sperm malformation ratio in all the three fluorine combined with zinc treatment groups were significantly lower than that in fluorine group(P＜0.05);②Comparison of the sperm malformation ratio in the groups treated by fluorine and fluorine combined with zinc and control group:the sperm malformation ratio in all the fluorine and low-dose zinc treatment group,fluorine and middle-dose zinc treatment group,fluorine and high-dose zinc treatment group and fluorine group were significantly higher than that in control group(P＜0.05).2.3 The contents of serum testosterone in rats①Compared with control group,it was significantly higher in groups treated with fluorine and zinc than that in control group(P＜0.05),but there was no significant difference between fluorine treatment group and control group(P＞0.05);②Compared with fluorine treatment group:it is significantly higher in all the groups treated by fluorine and zinc than that in fluorine group(P＜0.05);③Compared with fluorine and moderate-dose zinc treatment group: it is significantly lower in the fluorine and low-dose zinc group and fluorine and high-dose zinc group than that in fluorine group(P＜0.05).2.4 Testis fluoride content①it was significantly higher in groups treated with fluorine and fluorine combined zinc than that in control group(P＜0.05);②it is significantly lower in the group treated by fluorine and moderate-dose zinc than that in fluorine group(P＜0.05),it is significantly higher in the fluorine and high-dose zinc treatment group than that in fluorine group(P＜0.05),but no significant difference was found in the fluorine and low-dose zinc treatment group than that in fluorine group(P＞0.05);③Compared with fluorine and moderate-dose zinc treatment group: it is significantly higher in the fluorine and low-dose zinc group and fluorine and high-dose zinc group than that in fluorine group(P＜0.05).2.5 The activity of LDH in testis①it was significantly lower in all the fluorine treatment group and fluorine and every-dose zinc treatment group than that in control group(P＜0.05);②Compared with fluorine treatment group:it is significantly higher in the fluorine and moderate-dose zinc treatment group than that in fluorine group(P＜0.05);③Compared with fluorine and moderate-dose zinc treatment group: it is significantly lower in the fluorine and low-dose zinc group and fluorine and high-dose zinc group than that in fluorine group(P＜0.05p).2.6 The result of rats testis histopathology test testis spermatogenic epithelium morphosis is relatively integrated in control group and fluorine and moderate-dose zinc treatment group,amount of mature sperm can be observed.Only 2-3 stratum spermatogenic epithelium can be observed in fluorine treatment group and fluorine and moderate-dose zinc treatment group,lumina structure porosity,cell layer grow downwards,and the cell quantity decreased dramatically compared with control group, the same to sperm number.The cell layers decreased in fluorine and low-dose zinc treatment group compared with fluorine and moderate-dose zinc treatment group and control group,but it increased comparing with fluorine and moderate-dose zinc treatment group and fluorine group,primary spermatocyte and second spermatocyte increased dramatically in fluorine and low-dose zinc treatment group compared with control group,and interstitial cell also increased to a certain degree,dissociation mature sperm can be observed in lumens.2.7 The detection result of C-fos protein expression rate in testis spermatogenic cell①Compared with control group,the expression rate was significantly higher in groups treated with fluorine and fluorine combined zinc than that in control group(P＜0.05);②Compared with fluorine treatment group:it is significantly lower in the fluorine and moderate-dose zinc treatment group than that in fluorine group(P＜0.05),but no significant difference was found in fluorine and low-dose zinc treatment group and fluorine and high-dose zinc treatment group and fluorine treatment group(P＞0.05);③Significant difference was found in the fluorine and low-dose zinc group and fluorine and high-dose zinc group and fluorine and moderate-dose zinc group(P＜0.05).2.8 The detection result of c-jun protein expression rate in testis spermatogenic cell①Compared with control group,the expression rate was significantly higher in groups treated with fluorine and fluorine combined zinc than that in control group(P＜0.05);②Compared with fluorine treatment group:it is significantly lower in the fluorine and low-dose zinc treatment group and fluorine and moderate-dose zinc treatment group than that in fluorine group(P＜0.05),but no significant difference was found in fluorine and high-dose zinc treatment group and fluorine treatment group(P＞0.05);③Significant difference was found in the fluorine and low-dose zinc group and fluorine and high-dose zinc group and fluorine and moderate-dose zinc group(P＜0.05). 3 ConclusionMiddle-dose zinc(20 mg·kg^-1·d) can antagonize the reproductive toxicity of fluorine in male rats by lowering the fluorine burden,reinforcing LDH activity,and T and lowering the expression level of c-fos and c-jun protein expression in testis spermatogenic cell.