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Objective To evaluate the influence of fluoride on mitochondrial membrane potential of neuroblastoma SH-SY5Y cells,and on the expression levels of mitochondrial proteins mitofusion 1 (Mfn1) and fission 1 (Fis1).Methods A stable and feasible culture method of SH-SY5Y cells in vitro was established with different concentration of sodium fluoride [0.0 (control),0.4,2.0 and 4.0 mmol/L],and various periods exposure of 6,12,24,48 h;the mitochondrial membrane potential of SH-SY5Y cells was detected by mitochondrial membrane potential assay kit (JC-1);and the expression levels of Mfn1 and Fis1 proteins were detected by Western blotting.Results Compared with the control group (1.63 ± 0.18,1.13 ± 0.15,1.30 ± 0.02) for various periods exposure (6,12,48 h),the red/green fluorescence ratios of the mitochondrial membrane potential of SH-SY5Y cells exposed to 2.0 and 4.0 mmol/L of sodium fluoride were decreased significantly (1.01 ± 0.10,0.80 ± 0.04;0.75 ± 0.13,0.62 ± 0.10;0.82 ± 0.01,0.56 ± 0.04,P ＜ 0.05);compared with the control group (0.93 ± 0.03,1.05 ± 0.07,1.17 ± 0.04) for various periods exposure,the expression levels of mitochondrial Mfn1 protein were decreased significantly in 0.4,2.0,4.0 mmol/L sodium fluoride groups (6,12,48 h:0.75 ± 0.02,0.65 ± 0.05,0.57 ± 0.06;0.83 ± 0.06,0.79 ± 0.06,0.69 ±0.06;0.98 ± 0.05,0.73 ± 0.07,0.62 ± 0.09,P ＜ 0.05).Compared with the control group (0.90 ± 0.05) for exposure time 12 h,the expression levels of Fis1 protein were increased significantly in 2.0,4.0 mmol/L sodium fluoride groups (1.14 ± 0.06,1.23 ± 0.06,P ＜ 0.05).Conclusions The mitochondrial membrane potential and the expression levels of mitofusion 1 and fission 1 of SH-SY5Y cells treated with fluoride are abnormal,which might be associated with the theory of nerve cell damage from high oxidative stress.
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Background and purpose:Ethanol has been reported to stimulate progression of breast cancer, yet the underlying mechanism is not fully understood. This study aimed to investigate effects of ethyl alcohol (EtOH) on the calcium-activated neutral protease (CANP)-cyclin E/focal adhesion kinase (FAK) signaling and cell migration in breast cancer cells, as well as the role of epidermal growth factor receptor (EGFR) in the EtOH-stimulated effects, in order to assess the signaling mechanism(s) underlying how EtOH enhances cancer progression.Methods:Human breast cancer cell line MCF-7 was employed as a model system, with MCF-10A mammary epithelial cells as control. In vitro wound healing assay was carried out to evaluate EtOH-induced cell migration. The effects of EtOH or epidermal growth factor on the proteolysis of cyclin E/FAK were detected by Western blot. EGFR inhibitor (EGFR-I) and a speciifc inhibitor for CANP, Calpeptin, were applied to pretreat cultured cells to explore their inlfuences on the cell migration and cyclin E/FAK proteolysis triggered by EtOH.Results:Treatment of model cells with EtOH (0.3%) stimulated significant proteolysis of cyclin E/FAK in a dose-/time-dependent manner and increased migration (+47.30%,P<0.05) in MCF-7 breast cancer cells, but had no signiifcant effect on migration in MCF-10A cells. Pretreatment with Calpeptin (10 μmol/L) signiifcantly reduced EtOH (0.3%)- or EGFR (10 ng/mL)-induced cyclin E/FAK truncation. EGFR-I (3 μmol/L) pro-foundly reduced EtOH-indcued CANP dependent proteolysis of CANP1 and cyclin E/FAK as well as cell migration (-53.00%,P<0.01).Conclusion:EtOH signiifcantly stimulates activation of CANP via EGFR pathway, resulting in proteolysis of cyclin E/FAK and migration in MCF-7 breast cancer cells, suggesting EGFR-CANP signaling to be a potential target for suppression of metastasis in breast cancer.
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Objective To observe the effects of fluorosis on the levels of endogenous cystathionine beta-synthase (CBS) and hydrogen sulfide (H2S) in rats.Methods According to body weight,forty-eight Sprague-Dawley rats (body weight 105-180 g) were divided into three groups by a random number table(16 rats in each group,half male).Fluorine contents of the feed in control group,low-fluoride group and high-fluoride group were 9.80,15.40 and 23.80 mg/kg.After 6 months of fluorine exposure,the fluorine contents of urine and bone were determined by the method of fluorine ion-selective electrode ; H2S levels in serum and brain and the activity of CBS in brain were detected by methylene blue; and protein expression of CBS was detected by Western blotting.Results Compared with control group,dental fluorosis was found in rats of low-fluoride and high-fluoride groups.The differences of fluorine contents of urine and femur were statistically significant between groups(F =65.16,67.93,all P ＜ 0.05).The urinary and femoral fluorine in low-fluoride groups [(5.25 ± 0.45)mg/L,(1 196.54 ± 72.78)mg/kg] and high-fluoride groups[(13.17 ± 0.98)mg/L,(2 656.61 ± 170.12)mg/kg] were higher than those of control groups [(3.64 ± 0.20)mg/L,(870.71 ± 71.51)mg/kg,all P ＜ 0.05],and the increases were in a dose-dependent fashion(all P ＜ 0.01).The differences of H2S contents in serum and brain were statistically significant(F =4.83,1 456.13,all P ＜ 0.05).The H2S content in serum was higher in high-fluoride group [(17.64 ± 2.38) μ mol/L] than that of the control group [(10.29 ± 0.74) μ mol/L,P ＜ 0.01].The H2S contents in brain were higher in the low-fluoride [(364.74 ± 2.06)μmol/L] and high-fluoride groups [(513.43 ± 4.18) μmol/L] than those of the control group[(314.94 ± 0.72)μmol/L,all P ＜ 0.01],and the increase was in a dose-dependent fashion (P ＜ 0.01).The difference of CBS activity was statistically significant between groups (F =760.63,P ＜ 0.01).The CBS activities were lower in low-fluoride [(438.90 ± 2.83) mmol· kg-1· min-1] and high-fluoride groups [(529.83 ± 2.37)mmol· kg-1· min-1] than those of the control group [(596.33 ± 2.75) mmol · kg-1· min-1,all P ＜ 0.01],whereas the protein expression of CBS in brain in high-fluoride group (1.49 ± 0.08) was higher than that of the control group (1.19 ± 0.06,P ＜ 0.05).Conclusion Chronic fluorosis can affect the levels of endogenous CBS and H2,S,and the increases are in a dose-dependent fashion in addition to CBS activity.
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Aim To investigate the effects of endogenous and exogenous hydrogen sulfide(H_2S)on the proliferation and apoptosis of human fetal lung fibroblasts.Methods Human fetal lung fibroblasts were cultured under 2% O2～93% N_2～5% CO_2 for 24 h to produce hypoxia.Cells were divided into 6 groups:(1)Hypoxia group(N_2);(2)N_2+600 μmol·L~(-1) NaHS group;(3)N_2+1 200 μmol·L~(-1) NaHS group;(4)N_2+6 400 μmol·L~(-1) NaHS group;(5)N_2+400 μmol·L~(-1) cysteine(Cys)group;(6)N_2+200 μmol·L~(-1) S-adenosyl-L-methionine(SAM)group.After they were cultured for 24 h, MTT assay was used to evaluate the cell proliferation, and flow cytometry was used to detect cell apoptosis.Results Compared with N_2 group, 600 and 1 200 μmol·L ~(-1) NaHS(H_2S donor)significantly reduced proliferation induced by hypoxia of human fetal lung fibroblasts(P <0.01)without effects on apoptotic rates of cells(P >0.05)and 6 400 μmol·L~(-1) NaHS increased apoptosis of human fetal lung fibroblasts during hypoxia significantly(P <0.05), although no effects were found on proliferation of cells(P >0.05).In addition, Cys, substrate of cystathionine β-synthetase(H2S synthase, CBS) or SAM(activator of CBS)did not affect proliferation of human fetal lung fibroblasts induced by hypoxia(P >0.05), whereas apoptotic rates were increased significantly compared with that of N_2 group(P <0.05).Conclusions Endogenous and exogenous hydrogen sulfide can inhibit proliferation induced by hypoxia and promote apoptosis of human fetal lung fibroblasts, suggesting endogenous hydrogen sulfide may play a protective role through lung fibroblasts by inhibiting the pulmonary vascular structural remodeling caused by hypoxia.
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ObjectiveThe heroin-dependent animal model of rats was used to investigate the effects ofheroin on regulation of pain perception in the dorsal and ventral hippocampus of the heroin-dependent rats. Meth odsHeroin was injected subcutaneously twice a lay for 9 days according to the principle of daily increasing dose in the Sprague-Dawley rats. From the 10th day,rats were given heroin at dose of 27 mg · kg-1 once a day until the14th day, then the unit discharges of the dorsal and ventral hippocampus of rats were observed respectively afternoxious electric stimulation of the rat-tail by the extracellular single-unit recording with glass microelectrodes. ResultsWhen given noxious stimulation, most of the neurons in the dorsal hippocampus in the heroin-dependent ratswere unaffected(59.09％ ) ,whereas in the control rats ,the ratio of the neurons of the dorsal hippocampus affectedby noxious stimulation was about 66.67％, respectively(P ＜ 0.05 ). However,in the ventral hippocampus, the ratioof the neurons activated,inhibitory or unaffected was 20. 69％ ,41.38％ and 37.93％ from the control and was40.74％ ,33. 33％ and 25.93％ from the heroin group respectively with no significant difference between two groups (P ＞ 0.05 ) . ConclusionHeroin changed the regulation of pain perception in the hippocampus,primarily the dorsal hippocampus of rats.
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Objective To explore the prevalence of mild cognitive impairment (MCI) among elderly veterans. Methods 2 674 veterans ( aged 60 years and over) from 26 military sanatorium in Shijiazhuang city were studied. The Mini-Mental State Examination, Global Deterioration Scale, Activity of Daily Living, Hachinski Ischemic Scale and Hamilton Depression Scale were served as screening tools. Results The prevalence of total MCI was 8 08% in elderly people. The standardized prevalence of MCI was 6 87% in male and 10 38% in female (P
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0.05),the content ratio of glutamate to gamma-aminobutyric acid in the hippocampus of the dependent rats was lower than that in the control group(P