Study On The ATF-6/CHOP Pathway Of Endoplasmic Reticulum In Hippocampus Of Rats With SD Exposed To Fluoride

ObjectiveThe present study serves to discuss ERS in fluorosis to the nervous system damage and assess the affect of fluorosis in vivo experiments, which could provide a new way for demonstrating the forming mechanism of nervous system lesion caused by fluorosis. Methods 1. Animals and treatmentsSixty healthy male Sprague-Dawley(SD) rats 4~5 weeks old were divided into 4 groups randomly. Each group had 15 rats. Their weights were from 80 g to 100 g. The 4 experimental groups were control group, NaF group, NaF+NAC group, and NAC group. The control group was treated with 0.9% sodium chloride or deionized water. The NaF group and NAC group were respectively treated with 25mg/kg Bw·d NaF solution and 200mg/kgBw·d NAC solution, and the NaF+NAC group was treated with 25mg/kgBw·d NaF+200mg/kgBw·d NAC. Each group was administrated with gastric perfusion for 7 weeks, and every week the rats were continuously administered six days, with no-treatment in the last day. 2. The effects of fluoride-induced nervous system damageAfter fluoride exposure for 7 weeks, the rats were weighed, anesthetized and killed finally. Then the rat brains and hippocampus were taken out immediately, which were weighted for calculating their organic coefficients. Fluoride ion selective electrode method was used to determine the content of fluorine and apoptosis was detected by the TUNEL method in the hippocampus. 3. Measurement of the expression of ER stress markers in hippocampusThe expression of ATF-6 protein in hippocampus was assayed by immunohistochemistry. Western blot was used to detect the marker protein expression of GRP78 and CHOP. Results1. Analyzing the weight of rats, hippocampuses and their organic coefficients, compared with the control group, we found that the weight of rats and hippocampuses decreased significantly in NaF group(P<0.05). Compared with the control group, the content of fluorine in the hippocampus in NaF group significantly increased(P<0.05).2. TUNEL method was used for the detection of cell apoptosis, compared with the control group, the level of cell apoptosis elevated significantly with NaF treated group(P<0.05). Whereas, the rate of cell apoptosis in the NaF +NAC group markedly decreased, compared with the NaF group(P<0.05).3. Immunohistochemical method was used to test the ERS protein—ATF-6. The result shown that fluoride exposure raised the expression of ATF-6 significantly, compared with the control group(P<0.05).4. Western blot was used to detect the expression of ER stress markers in hippocampus: GRP78 and CHOP. The expression of GRP78 and CHOP increased markedly, compared with the NaF group(P<0.05).The antioxidant NAC suppressed the expression of GRP78 and CHOP(P<0.05) Conclusion1. ERS pathway might be one of the important mechanisms of fluoride-induced nerve damage in rats.2. Naf induced nerve damage possibly through activating ATF-6/CHOP pathway.