Objective:To investigate the regulatory effects of transcutaneous auricular vagus nerve stimulation (taVNS) on functional connectivity (FC) of thalamic subregions in patients with premenstrual syndrome (PMS).Methods:This study was a cross-sectional investigation. Clinical, laboratory, and imaging data were retrospectively collected from 56 PMS patients (PMS group) and 66 healthy controls (control group) recruited from various universities and hospitals in Nanning between November 2021 and June 2024. Resting-state functional MRI (fMRI) data and fMRI data during taVNS immediate stimulation (2 Hz, 25 Hz) were acquired from subjects during their late luteal phase. Using thalamic subregions (anterior thalamic nucleus, lateral nucleus, ventral nucleus, medial nucleus, central nucleus, posterior nucleus) as seeds, two-sample t-tests or paired t-tests were employed to analyze alterations in thalamic subregion FC in PMS patients and the regulatory effects of taVNS on these changes. Independent samples t-test were used to compare the differences in clinical and laboratory indicators between the PMS group and the control group. The relationship between taVNS regulation of thalamic subregion FC in PMS patients and thalamic internal functional connectivity were analyzed using mediation effect analysis. Results:Compared to the control group, patients in the PMS group showed increased scores on the Daily Record of Severity of Problems, Pittsburgh Sleep Quality Index, Self-Rating Anxiety Scale, Self-Rating Depression Scale, Hamilton Anxiety Rating Scale 17, and Hamilton Depression Rating Scale 14 during the late luteal phase ( P<0.05). At baseline, PMS patients exhibited higher FC between the left thalamic lateral nucleus and the left insula, and lower FC between the left medial nucleus, posterior nucleus, and ventral nucleus of the thalamus and the right middle frontal gyrus (MFG) compared to the control group (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). During 2 Hz taVNS immediate stimulation in PMS group, FC between the left thalamic medial nucleus, posterior nucleus, ventral nucleus and the right MFG, as well as the FC between the left thalamic ventral nucleu and the left MFG increased compared to baseline levels; meanwhile, FC between the left thalamic posterior nucleus, ventral nucleus and the left insula decreased compared to baseline levels (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). During 25 Hz taVNS immediate stimulation, the FC between the left thalamic ventral nucleus and the right MFG decreased compared to the baseline level (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). Mediation effect analysis showed that the FC between the left thalamic posterior nucleus and the left lateral nucleus mediated part of the association between the FC of the left lateral thalamic nucleus-left insula and the FC of the left ventral thalamic nucleus-left putamen/insula; there were significant direct effects between the FC of the left lateral thalamic nucleus-the left posterior nucleus and FC of the left lateral thalamic nucleus-the left insula, as well as between the FC of the left ventral thalamic nucleus-the left MFG and FC of the left ventral thalamic nucleus-the right MFG. Conclusions:taVNS can modulate abnormal FC of the left thalamic subregions in PMS patients, restoring it toward normalization. The regulatory effects of 2 Hz stimulation are more pronounced than those of 25 Hz stimulation. This modulation primarily operates through two pathways: the left thalamic lateral nucleus-left insula-left thalamic ventral nucleus pathway and the left MFG-left thalamic ventral nucleus-right MFG.

Objective To modify the dosage form of Guoganglong Sugar-Coated Tablets,and to investigate the chemical equivalence and pharmacodynamic equivalence between the new and old dosage forms of Guoganglong Tablets.Methods The preparation technology was screened by single-factor exploration with the indexes of hardness and dispersion uniformity,the quality standard was established by thin layer chromatography identification,high performance liquid chromatography(HPLC)content determination and charactertistics chromatograms,the constituents absorbed into blood were analyzed by ultra-high performance liquid chromatography quadrupole time of flight mass spectrometry(UHPLC-Q-TOF-MS),and the pharmacodynamics was evaluated by anti-inflammatory experiment and analgesic experiment in mice model with inflammatory pain.Results The molding process of Guoganglong Dispersible Tablets was determined.A thin layer chromatographic identification method was established.A method for detecting the content of Guoganglong Tablets and the charactertistic chromatograms was established by HPLC,and a total of 15 common peaks were identified,with the similarity of the new-old dosage forms being greater than 0.9.A total of 12 constituents absorbed into blood were identified.Guoganglong Tablets could alleviate swelling degree of toes in mice with inflammatory pain,reduce the levels of interleukin 6(IL-6)in serum,improve spleen index,and increase the thermal pain threshold and acetic acid twisting frequency in mice.Conclusion The prescription process of Guoganglong Dispersible Tablets is stable,the quality standard is feasible,the chemical composition and pharmacological actions between the new and old dosage forms are basically the same,and in terms of certain indexes,Dispersible Tablets of Guoganglongare superior to Sugar-Coated Tablets.

Objective:To investigate the regulatory effects of transcutaneous auricular vagus nerve stimulation (taVNS) on functional connectivity (FC) of thalamic subregions in patients with premenstrual syndrome (PMS).Methods:This study was a cross-sectional investigation. Clinical, laboratory, and imaging data were retrospectively collected from 56 PMS patients (PMS group) and 66 healthy controls (control group) recruited from various universities and hospitals in Nanning between November 2021 and June 2024. Resting-state functional MRI (fMRI) data and fMRI data during taVNS immediate stimulation (2 Hz, 25 Hz) were acquired from subjects during their late luteal phase. Using thalamic subregions (anterior thalamic nucleus, lateral nucleus, ventral nucleus, medial nucleus, central nucleus, posterior nucleus) as seeds, two-sample t-tests or paired t-tests were employed to analyze alterations in thalamic subregion FC in PMS patients and the regulatory effects of taVNS on these changes. Independent samples t-test were used to compare the differences in clinical and laboratory indicators between the PMS group and the control group. The relationship between taVNS regulation of thalamic subregion FC in PMS patients and thalamic internal functional connectivity were analyzed using mediation effect analysis. Results:Compared to the control group, patients in the PMS group showed increased scores on the Daily Record of Severity of Problems, Pittsburgh Sleep Quality Index, Self-Rating Anxiety Scale, Self-Rating Depression Scale, Hamilton Anxiety Rating Scale 17, and Hamilton Depression Rating Scale 14 during the late luteal phase ( P<0.05). At baseline, PMS patients exhibited higher FC between the left thalamic lateral nucleus and the left insula, and lower FC between the left medial nucleus, posterior nucleus, and ventral nucleus of the thalamus and the right middle frontal gyrus (MFG) compared to the control group (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). During 2 Hz taVNS immediate stimulation in PMS group, FC between the left thalamic medial nucleus, posterior nucleus, ventral nucleus and the right MFG, as well as the FC between the left thalamic ventral nucleu and the left MFG increased compared to baseline levels; meanwhile, FC between the left thalamic posterior nucleus, ventral nucleus and the left insula decreased compared to baseline levels (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). During 25 Hz taVNS immediate stimulation, the FC between the left thalamic ventral nucleus and the right MFG decreased compared to the baseline level (GRF corrected, voxel-level P<0.001, cluster-level P<0.05). Mediation effect analysis showed that the FC between the left thalamic posterior nucleus and the left lateral nucleus mediated part of the association between the FC of the left lateral thalamic nucleus-left insula and the FC of the left ventral thalamic nucleus-left putamen/insula; there were significant direct effects between the FC of the left lateral thalamic nucleus-the left posterior nucleus and FC of the left lateral thalamic nucleus-the left insula, as well as between the FC of the left ventral thalamic nucleus-the left MFG and FC of the left ventral thalamic nucleus-the right MFG. Conclusions:taVNS can modulate abnormal FC of the left thalamic subregions in PMS patients, restoring it toward normalization. The regulatory effects of 2 Hz stimulation are more pronounced than those of 25 Hz stimulation. This modulation primarily operates through two pathways: the left thalamic lateral nucleus-left insula-left thalamic ventral nucleus pathway and the left MFG-left thalamic ventral nucleus-right MFG.

Objective To analyze the prognosis of children with severe mycoplasma pneumonia (MPP) and its correlation with serum SAA, PCT and SF levels, so as to provide a basis for evaluating the prognosis of children with MPP. Methods A total of 273 children with MPP admitted to our hospital from January 2020 to December 2020 were divided into mild MPP children (n=187) and severe MPP children (n=86) according to the severity of their disease. According to the prognosis, children with severe MPP were divided into survival group (n=65) and death group (n=21). Serum SAA, PCT and SF levels were determined. Pearson correlation analysis was used to analyze the correlation between serum SAA, PCT and SF levels and APACHE ⅱ score. ROC curve was used to analyze the predictive value of serum SAA, PCT and SF levels for poor prognosis of children with severe MPP. Results The levels of serum SAA, PCT and SF and APACHE II score in children with severe MPP were significantly higher than those in children with mild MPP (P<0.05). Serum SAA, PCT and SF levels and APACHE II score in death group were significantly higher than those in survival group (P<0.05). Pearson correlation analysis showed that APACHE II score was positively correlated with serum SAA, PCT and SF levels (r =0.474,0.519,0.446,P<0.05). The AUC, sensitivity and specificity of combined ROC curve analysis to predict the prognosis of severe MPP were 0.871, 85.9% and 93.6% respectively, which were higher than those of SAA, PCT and SF alone. Conclusion SAA, PCT and SF are closely related to the prognosis of severe MPP, and can be used as potential markers to predict poor prognosis of severe MPP children.

Objective To explore the value of KRAS mutation predicting prognosis of patients with colorectal liver-only metastasis after hepatectomy.Methods The retrospective case-control study was conducted.The clinicopathological data of 79 patients with colorectal liver-only metastasis who underwent hepatectomy in the Sun Yat-sen University Cancer Center between October 2010 and October 2016 were collected.KRAS mutation in colorectal cancer tissue was detected by fluorescent quantitative polymerase chain reaction (PCR) and laser flight mass spectrometer.Observation indicators:(1) KRAS mutation;(2) relationship between KRAS mutation and clinicopathological factors of patients with colorectal liver-only metastasis;(3) follow-up and survival situations.Follow-up using outpatient examination and telephone interview was performed to detect recurrence-free survival and overall survival up to June 30,2017.The relationship between KRAS mutation and clinicopathological factors of patients with colorectal liver-only metastasis was analyzed by the chi-square test or Fisher exact probability.The survival curve and time were respectively drawn and calculated by the Kaplan-Meier method,and COX regression model was used for survival analysis.Results (1) KRAS mutation:79 patients received KRAS gene detection of surgical tumor tissues,including 54 in wide-type mutation and 25 in mutant-type mutation.Of 25 patients in mutant-type mutation,mutation at codon 12 of KRAS exon 2 was in 21 patients,and GGT＞GAT (G12D),GGT＞GTT (G12V),GGT＞TGT (G12C),GGT＞GCT (G12A) and GGT＞CGT (G12R) of mutation types were respectively detected in 13,4,2,1 and 1 patients;mutation at codon 13 of KRAS exon 2 was in 3 patients,with a mutation type of GGC＞GAC (G13D);mutation at codon 61 of KRAS exon 3 was in 1 patient,with a mutation type of CAA＞CAT (Q61H).(2) Relationship between KRAS mutation and clinicopathological factors of patients with colorectal liver-only metastasis:primary tumor located in right and left hemicolon were detected in 11,14 patients with mutant-type mutation and 7,47 patients with wide-type mutation,respectively,with a statistically significant difference (x2=9.357,P＜0.05).(3) Follow-up and survival situations:79 patients were followed up for 2.0-71.0 months,with a median time of 29.0 months.Median recurrence-free survival time and median overall survival time were respectively 11.3 months,43.5 months in patients with mutant-type mutation and 9.9 months,44.3 months in patients with wide-type mutation,respectively,with no statistically significant difference in recurrence-free and overall survivals [hazard ratio (HR)=1.255,1.108,95％ confidence interval (CI):0.741-2.126,0.521-2.355,P＞0.05].Further analysis:of patients with low clinical risk score (CRS) of Memorial Sloan Caitlin Cancer Center (MSKCC),median recurrence-free survival time was 11.3 months in 17 patients with mutant-type mutation and 23.5 months in 26 patients with wide-type mutation,with a statistically significant difference in recurrence-free survival of patients (HR=2.082,95％CI:1.006-4.307,P＜0.05).The median overall survival time was 44.6 months in 17 patients with mutant-type mutation and 49.0 months in 26 patients with wide-type mutation,with no statistically significant difference in overall survival of patients (HR =1.165,95％CI:0.413-3.282,P＞0.05).Of patients with high CRS of MSKCC,median recurrence-free survival time and median overall survival time were respectively 5.6 months,28.7 months in 7 patients with mutant-type mutation and 4.5 months,36.7 months in 24 patients with wide-type mutation,with no statistically significant difference in recurrence-free and overall survivals (HR=0.402,1.197,95％CI:0.284-1.656,0.371-3.866,P＞0.05).Conclusions KRAS mutation is often detected in patients with right colon cancer.Recurrence-free survival time is obviously reduced in patients with KRAS mutation and low CRS of MSKCC.

Objective To study the expression of silent information regulator (SIRT) in brains of rats with chronic fluorosis and reveal the correlation between SIRT1 and the ability of learning and memory of rats.Methods Sixty SD rats were selected and their body weight was (100 ± 20) g,according to the body mass of the rats,random number table method was used to divide rats into control group,low and high fluoride groups,experimental period was 3 and 6 months (ten rats in each experimental period,half males and half females).In control group,the rats were fed with drinking water containing no more than 0.5 mg/L fluoride;the rats in low and high fluoride groups were fed drinking water containing 5.0 and 50.0 mg/L fluoride,respectively.All of rats were fed the same standard food containing no more than 0.6 mg/kg fluoride.Three degree method was used to check the formation of dental fluorosis.Rat urinary fluoride was determined via the fluoride electrode method;Morris water maze method was used to detect the ability of learning and memory of rats (the escape latency time,the number of crossing the platform and stay time in platform quadrant);the protein and mRNA expression levels of SIRT1 were detected by Western blotting and Real-time PCR,respectively.Results In the experimental period of 3 and 6 months,no dental fluorosis was observed of rats in control group,but there were different degrees of dental fluorosis in low and high fluoride groups,especially in high fluoride group.The urinary fluorine contents [(1.60 ± 0.09),(1.91 ± 0.16) mg/L;(1.94 ± 0.19),(2.31 ± 0.18) mg/L] of rats fed with low and high fluoride for 3 or 6 months were significantly higher than those in control group [(1.08 ± 0.15),(1.09 ± 0.17) mg/L,P ＜ 0.05].The escape latency time [(18.36 ± 2.80) s] of rats in the high fluoride group at the end of 3 months was higher than that of control group [(6.68 ± 3.01) s,P ＜ 0.05],the number of stay time in platform quadrant [(12.91 ± 3.25) s] was lower than that of control group [(19.97 ± 3.30) s,P ＜ 0.05].The escape latency time [(15.46 ± 4.56),(28.16 ± 4.00) s] of rats in low and high fluoride groups at the end of 6 months were all higher than that of control group [(6.62 ± 2.31) s,P ＜ 0.05];the number of crossing the platform and stay time in platform quadrant [(2.25 ± 1.71) times,(12.73 ± 3.55) s;(1.40 ± 1.15) times,(9.26 ± 1.72) s] of these rats were significantly lower than those of the control group [(4.00 ± 1.58) times,(19.53 ± 4.36) s,P ＜ 0.05].The expression levels of protein [(73.84 ± 9.68)％,(73.23 ± 4.51)％;(53.30 ± 17.63)％,(54.69 ± 18.71)％] and mRNA [(70.33 ± 4.89)％,(66.27 ± 3.38)％;(37.72 ± 4.89)％,(44.15 ± 1.74)％] of SIRT1 in the hippocampus and cortex of rats fed with high fluoride for 3 or 6 months were significantly lower than those in control group [(100.00 ± 13.51)％,(100.00 ± 13.60)％;(100.00 ± 15.37)％,(100.00 ± 12.19)％;(100.00 ± 2.65)％,(100.00 ± 4.34)％;(100.00 ± 3.40)％,(100.00 ± 4.52)％,P ＜ 0.05].Whereas,the decreased expression levels of protein [(77.65 ± 14.51)％,(71.51 ± 8.27)％] and mRNA [(57.78 ± 1.96)％,(63.76 ± 2.16)％] of SIRT1 in the hippocampus and cortex of rats in the low fluoride group were only observed at the end of 6 month of experiment (P ＜ 0.05).The expression of protein of SIRT1 in the hippocampus and cortex of rats in 3 or 6 months was negatively correlated with the escape latency time of rats (r=-0.598 5,-0.493 2;-0.782 6,-0.777 3,P＜ 0.05),and it was positively correlated with the number of crossing the platform (r =0.547 7,0.523 3;0.720 5,0.715 4,P ＜ 0.05).Conclusion The decrease of the ability of learning and memory in rats with chronic fluorosis may be related to the decreased expression of SIRT1 influenced by chronic fluorosis.

Objective To detect the expression of peroxisome proliferator-activated receptor γ (PPARγ) in the brains of rats with chronic fluorosis and elucidate the relationship between PPARγand oxidative stress in chronic fluorosis.Methods According to body weight (100-120 g),sixty healthy SD rats were divided into control group (less than 0.5 mg/L fluoride in drinking water),low fluoride group (5.0 mg/L fluoride in drinking water,prepared by NaF),and high fluoride group (50.0 mg/L fluoride in drinking water) via the random number table method,20 rats in each group (half male and half female).The experiment periods were 3 and 6 months,respectively.Then 24-hour urine samples of rats were collected from each group,all rats were put to death and brain tissues were taken.The fluoride contents in urine and brain tissue were measured with fluoride-ion selective electrode;the levels of PPARγ protein and mRNA in the cortex and hippocampus were determined by Western blotting and Real-time fluorescence quantitative PCR,respectively;and the activities of superoxide dismutase (SOD) and level of malondialdehyde (MDA) in serum were detected by xanthine oxidase method and thiobarbituric acid method;the correlation between PPARγ protein expression and oxidative stress was analyzed.Results After 3 and 6 months of treatment,the contents of fluoride in urine and brain in low fluoride group [(1.57 ± 0.18) mg/L,(3.43 ± 0.70) μg/g;(1.79 ± 0.17) mg/L,(7.40 ± 1.21) μg/g] were higher than those of control group [(1.11 ± 0.17) mg/L,(2.39 ± 0.50) μg/g;(1.02 ± 0.15) mg/L,(2.87 ± 0.82) μg/g,P ＜ 0.05],and the values in high fluoride group [(1.91 ± 0.23) mg/L,(6.70 ± 0.87) μg/g;(2.44 ± 0.51) mg/L,(12.10 ± 1.30) μg/g] were significantly higher than those in low fluoride group (P ＜ 0.05).In high fluoride group after 3 months of treatment,the expression of PPARγprotein [(79.00 ± 3.46)％,(80.35 ± 2.50)％] and mRNA [(79.11 ± 11.18)％,(82.10 ± 9.94)％] in hippocampus and cortex of rat brains were significantly lower than those of low fluoride group [(104.01 ± 5.77)％,(101.17 ± 6.35)％;(112.88 ± 22.15)％,(101.14 ± 8.60)％,P＜ 0.05];the expression of PPARγprotein [(64.32 ± 10.43)％,(60.20 ± 10.92)％] and mRNA [(41.03 ± 9.93)％,(52.25 ± 11.48)％] in the same brain regions of the rats after 6 months of treatment in high fluoride group were significantly lower than those of control group [(99.99 ± 11.19)％,(100.00 ± 11.30)％;(100.00 ± 10.00)％,(100.00 ± 9.00)％] and low fluoride group [(73.88 ± 3.36)％,(81.50 ± 14.90)％;(76.02 ± 8.65)％,(73.36 ± 7.43)％,P ＜ 0.05].The activities of SOD in serum in low and high fluoride groups after 6 month treatment [(37.94 ± 1.92),(35.54 ± 2.53) U/ml] were significantly lower than that of control group [(41.24 ± 0.66) U/ml,P ＜ 0.05],and the value in high fluoride group was lower than that in low fluoride group (P ＜ 0.05);serum MDA contents in high fluoride group after 3 and 6 month treatment [(8.29 ± 1.49),(11.63 ± 1.04) nmol/mg pr] were higher than those in low fluoride group [(6.39 ± 0.69),(7.50 ± 1.64) nmol/mg pr] and control group [(5.02 ± 0.71),(5.87 ± 1.03) nmol/mg pr,P ＜ 0.05].The correlation analysis results showed the levels of PPARγprotein in hippocampus and cortex of rats were negatively correlated with fluoride contents in brain tissues (3 month:r=-0.769,-0.793;6 month:r =-0.832,-0.870;P ＜ 0.05),positively correlated with SOD activities (3 month:r =0.550,0.826;6 month:r =0.822,0.896;P ＜ 0.05) and negatively correlated with MDA contents (3 month:r =-0.703,-0.609,6 month:r =-0.792,-0.657;P ＜ 0.05) in serum.Conclusions Declined expression of PPARγat protein and mRNA levels has been detected in brains of rats with chronic fluorosis,which might be related to the increase of oxidative stress.PPARγ may be involved in the occurrence of chronic fluorosis.

Objective: To investigate the effect of lovastatin on oxidative stress and apoptosis in neurons induced by β-amyloid peptide (Aβ). Methods: Primary culture of rat hippocampal neuron was treated with Aβ oligomers alone or combined with lovastatin. The levels of OH^-, H₂O₂, O₂·^-, malondialdehyde, superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) activities were measured by biochemical methods and protein expression of caspase-3 and bcl-2 was detected by Western blot. Results: As compared with the control group, treatment of 0.5 μmol/L Aβ oligomers for 48 h led to significant increase of OH^-, H₂O₂, O₂·^- and malondialdehyde content, inhibition of SOD and GSH-PX activities, enhanced caspase-3 expression and decreased bcl-2 expression. Interestingly, these neurotoxic modifications on the levels of OH^-, H₂O₂, O₂·^- and malondialdehyde content, SOD and GSH-PX activities, and the protein expression of cleaved caspase-3 and bcl-2 were significantly attenuated when the cells were pretreated with 0.1 μmol/L lovastatin for 24 h before exposure of Aβ oligomers. Conclusion Lovastatin may play an important role in antagonizing the neurotoxicity of Aβ through a mechanism likely related to the inhibition of oxidative stress.

Objective To explore the influences of protein kinase Cβ (PKC3) inhibitor LY333531 on oxidative injury and apoptosis of SH-SY5Y cells induced by fluorosis.Methods The SH-SY5Y cell model of fluorosis was established,and the experiment was divided into three groups:control group [0.0 mmol/L sodium fluoride (NaF) and 0.0 μmol/L LY333531],the fluoride group (0.5 mmol/L NaF and 0.0 μmol/L LY333531),and the PKCβ inhibitor group (0.5 mmol/L NaF and 0.2 μmol/L LY333531),n =3.Flow cytometry was used to detect the changes of reactive oxygen species (ROS) and apoptosis rate,fluorescent probe technique was used to detect mitochondrial membrane potential after each group for 48 h.Results Compared with the control group [(3.32 ± 0.29) × 103,0.60 ± 0.09,(7.58 ± 1.20)％],the level of ROS [(5.99 ± 0.32) × 103] was increased,mitochondrial membrane potential (0.28 ± 0.06) was decreased,and the apoptosis rate [(18.00 ± 2.32)％] was increased in the fluoride group (all P ＜ 0.05);compared with the fluoride group,the level of ROS [(5.12 ± 0.25) × 103] was decreased,mitochondrial membrane potential (0.42 ± 0.03) was increased,and the apoptosis rate [(11.79 ± 0.70)％] was decreased in the PKCβ inhibitor group (all P ＜ 0.05).Conclusions Excess fluoride could cause oxidative damage and apoptosis in cells.PKC3 inhibitor LY333531 has a protective effect in oxidative damage and apoptosis by fluorosis.

Objective To investigate the influence of chronic fluorosis on protein kinase Cβ (PKCβ)/p66shc signal pathway in the brain of rats,and reveal the molecular mechanism of brain damage.Methods According to body weight by the random number table method thirty SD rats were divided into three groups of 10 each (half females and half males),the normal control group [less than 0.5 mg/L of fluorine (prepared with NaF) in drinking water],low fluoride exposure group (10.0 mg/L fluorine),and high fluoride exposure group (50.0 mg/L fluoride).The experiment period was 6 months.The protein level of PKCβ,p66shc,phospho-p66shc and preserved ammonia acyl isomerase (Pin1) in rat brain was detected by Western blotting.The level of neuron nuclear antigen (NeuN),p66shc and phospho-p66sh in brain of rats was detected by immunohistochemistry.Results By Western blotting,the levels of PKCβ,Pin1 and phospho-p66shc protein in brain tissue in high fluoride exposure group [(193.00 ± 57.53)％,(228.21 ± 71.14)％,(201.54 ±:50.86)％] were higher than those of the normal control groups [(100.00 ± 21.24)％,(100.00 ± 40.55)％,(100.00 ± 13.35)％,all P ＜ 0.05].By immunohistochemistry,the numbers of NeuN staining in brain tissue of the rats in both high and low fluoride exposure groups [(49.50 ± 12.57)％,(65.66 ±14.58)％] were lower than that of the control group [(100.00 ± 18.32)％,all P ＜ 0.01].The level of phospho-p66shc protein in brain tissue in high fluoride exposure group [(242.66 ± 93.01)％] was higher than those of the low fluoride exposure and the normal control groups [(152.53 ± 60.65)％,(100.00 ± 25.63)％,all P ＜ 0.01].Conclusion Chronic fluorosis has increased the expressions of PKCβ,Pin1 and phospho-p66shc at protein level in brain of rats,which may be related to the molecular mechanism of brain damage resulted from chronic fluorosis.