Effects Of Chronic Fluoride Exposure On M1/M2 Polarization Of Cortical Microglia In Mice

[Objective] Fluoride can not only cause bone phase damage,but also accumulate in the brain through the blood-brain barrier,resulting in the decline of learning and memory ability and the occurrence of inflammation in the central nervous system.Studies have shown that fluoride can activate microglia,but the effect of long-term fluoride exposure on morphological changes and M1/M2 polarization of mouse cortical microglia has not been reported.Therefore,this study mainly observed the effect of chronic fluoride exposure on the M1/M2 polarization of mouse cortical microglial cells,so as to provide a basis for exploring the mechanism of action of fluoride injury on the nervous system,and to provide a theoretical basis for the prevention and treatment of fluoride poisoning.[Method] Experimental animals: 48 healthy BCLB/C male mice,weighing about 25 g,provided by China Institute of Radiation Protection,were randomly divided into four groups: control group,low,medium and high fluoride group,with concentrations of 0,25,50 and 100 mg/L of Na F,respectively,12 mice per group.During the whole period of fluoride exposure,mice were allowed to drink and eat freely,and water and food intake of each group were recorded.After fluoride exposure for 150 days,8 mice were randomly selected from each group and sacrificed by the method of broken neck.Fluoride content in femur was determined by fluoride ion selective electrode method.Paraffin sections were made from the brain tissue of the remaining mice for HE and nissl staining,and the pathological changes were observed under a light microscope.Immunohistochemistry was used to analyze the morphological changes of microglia cells.The expressions of Iba-1,i NOS and Arg-1 were detected by western-blot and double immunofluorescence.RT-PCR was used to detect and analyze the expressions of cortical-related inflammatory factors,neuro-related factors and synaptic related factors.[Results] 1.The fluoride content in the femur of mice increased significantly with the increase of fluoride concentration(all p < 0.01).2.The cortex of the fluorinated group showed different degrees of morphological disorders compared with the control group,and nissl bodies and neurons were significantly reduced,and a large number of neurons showed swelling,evacuation,atrophy and necrosis.3.Immunohistochemical results showed that the Iba-1 immunoreactivity in the cortex of mice in the medium fluoride group and the high fluoride group was significantly increased(p < 0.01).The number of branched and intermediate microglia was significantly decreased(p < 0.01),and the number of ameboid microglia was significantly increased(p < 0.01).4.The results of immunofluorescence double staining showed that the i NOS expression was significantly increased(p < 0.01)and the Arg-1 expression was significantly decreased(p < 0.01)in the fluorinated group.At the same time,the results of western-blot and RT-PCR were also consistent with that,the expressions of Iba-1 and i NOS were on the rise,and the expressions of Arg-1 were on the decline,both of which were significant(p < 0.05).5.PCR results of inflammatory cortical factors in mice showed that the m RNA expression levels of i NOS,INF-?,TNF-? and TGF-? were significantly increased in the fluoride group(p < 0.05).The Arg-1 m RNA of the low-fluoride group was significantly lower than that of the control group(p < 0.05).The expression level of IL-4 m RNA in the high-fluoride group was significantly lower than that in the control group(p < 0.05).6.The results of PCR detection of cortical neurokines in mice showed that the expressions of IGF-1 and BDNF m RNA in the high-fluoride group were significantly lower than those in the control group(p < 0.05).7.The PCR results of cortical synaptic related factors in mice showed that the m RNA expression of Synaptophysin and PSD-95 in the high-fluoride group was significantly lower than that in the control group(p < 0.05).[Conclusion] Chronic fluoride exposure aggravates the transformation of mouse cortical microglia from branchlike to ameboid morphology and induces the polarization of microglias M1/M2.The results showed that the release of pro-inflammatory cytokines induced by fluoride increased,while the release of anti-inflammatory cytokines,neurokines and synaptic cytokines decreased,suggesting that the relationship between fluoride and neurotoxicity may be related to the neuroinflammation caused by inflammatory cytokines released after the overactivation of M1 microglia.