Background Neurotransmitter secretion disorder induced by chronic manganese poisoning has always been one of the important causes of body injury, but the mechanism of neurotransmitter secretion disorder caused by manganese is not clear at present. Objective To investigate the effects of presynaptic membrane intracellular protein 13-1 (Munc13-1) and synapse fusion protein binding protein 18-1 (Munc18-1) on dopamine secretion dysfunction induced by manganese chloride (MnCl₂) in human neuroblastoma (SH-SY5Y) cells. Methods A SH-SY5Y cell model induced by MnCl₂ was established. Cell viability was measured by MTT assay. Four experimental groups were set up: control group and low-, medium-, and high-dose manganese groups (0, 100, 200, and 400 μmol·L⁻¹ MnCl₂). They were treated with corresponding doses of MnCl₂ for 24 h. The secretion of dopamine was measured by enzyme-linked immunosorbent assay. The mRNA expression of Syntaxin-1 was detected by real-time quantitaive PCR. Total cell proteins were extracted, and the protein expression levels of Munc13-1, Munc18-1, and Syntaxin-1 were detected by Western blotting. The correlations of MnCl₂ exposure and dopamine secretion with the protein expressions of Munc13-1 and Munc18-1 were also analyzed by Pearson correlation. Results Compared with the control group, the cell viability rate decreased gradually with the increase of manganese exposure concentration, and the difference between the medium- and the high-dose manganese groups was statistically significant (P<0.05). The concentration of dopamine in cell culture medium of all manganese exposure groups decreased with the increase of manganese concentration, and compared with the control group and the low-dose manganese group, the medium- and the high-dose manganese groups were statistically significant (P<0.05). The expression of Syntaxin-1 at mRNA or protein level did not change significantly among groups (P>0.05). Compared with the control group, the protein expression of Munc13-1 decreased and that of Munc18-1 increased with the increase of manganese concentration (P<0.05). Compared with the low-dose manganese group, the changes of Munc13-1 protein in the high-dose manganese group and Munc18-1 protein in the medium- and high-dose manganese groups had statistical significance (P<0.05). Compared with the medium-dose manganese group, the protein changes of Munc18-1 in the high-dose manganese group were statistically significant (P<0.05). The correlation analysis showed that MnCl₂ dose was negatively correlated with Munc13-1 protein expression (r=−0.898, P<0.05), and positively correlated with Munc18-1 protein expression (r=0.678, P<0.05). Dopamine secretion was positively correlated with Munc13-1 protein expression (r=0.932, P<0.05), and negatively correlated with Munc18-1 protein expression (r=−0.817, P<0.05). Conclusion The inhibition of dopamine secretion in SH-SY5Y cells induced by manganese exposure is related to up-regulation of Munc18-1 and down-regulation of Munc13-1 expression levels, which may be one of the reasons for nerve injury caused by manganese.

Objective To observe the effects of curcumin on hepatic oxidant stress in water arsenic-exposed rats and to study its mechanism,which can offer references for curcumin used in antioxidant therapy of arsenic poisoning.Methods Thirty-two SD rats were divided into 4 groups according to body weight by random number table,half male and half female.Including control group (lavaged 135 days with deionized water),arsenic poisoning group (lavaged 45 days with deionized water after lavaging 90 days with 10 mg/kg sodium arsenite),pure curcumin group (lavaged 135 days with 1 000 mg/kg curcumin solution) and curcumin treatment group (lavaged 45 days with 1 000 mg/kg curcumin solution after lavaging 90 days with 10 mg/kg sodium arsenite),8 rats in each group.The arsenic contents of urine (urine creatinine corrected) and liver were detected by hydride generation inductively coupled plasma optical emission spectrometer (HG-ICP-OES);the activity of Cu/Zn-superoxide dismutase (SOD1) and catalase (CAT),the contents of malondialdehyde (MDA) in serum and liver homogenate by colorimetric method;the protein expression of liver antioxidant enzyme (SOD 1 and CAT) was assayed by Western blotting.Results The arsenic contents of urine and liver in arsenic poisoning group [(5.83 ± 0.29)μg/g Cr,(15.76 ± 1.65)μg/g] and the arsenic contents of urine in curcumin treatment group [(1.07 ± 0.14)μg/g Cr] were obviously higher than those of control group [(0.40 ± 0.14)μg/g Cr,(4.56 ± 1.05)μg/g,all P ＜ 0.05];compared to arsenic poisoning group,the arsenic contents of urine and liver in curcumin treatment group [(1.07 ± 0.14)μg/g Cr,(5.42 ± 1.76)μg/g] were obviously lower (all P ＜ 0.05).The contents of serum and liver SOD1,CAT and MDA in control group respectively were (102.46 ± 5.03),(29.33 ± 8.13)U/ml,(3.11 ± 0.49)μ mol/L and (204.05 ± 18.33),(126.26 ± 13.19)U/mg prot,(1.62 ± 0.42) μmol/g prot.Compared to the control,the activity of serum and liver SOD1 and CAT in arsenic poisoning group [(60.97 ± 7.94),(13.56 ± 5.14)U/ml and (133.66 ± 11.51),(74.01 ± 13.30)U/mg prot] were lower,the contents of MDA [(7.26 ± 0.54)μmol/L and (2.61 ± 0.52)μmol/g prot] were higher (all P ＜ 0.05).Compared to arsenic poisoning group,the activity of serum and liver SOD1 and CAT in curcumin treatment group [(87.39 ± 9.38),(20.45 ± 6.49) U/ml and (178.27 ± 9.32),(93.70 ± 20.35)U/mg prot] were higher,the contents of MDA [(4.34 ± 0.79)μmol/L and (1.92 ± 0.18)μmol/g prot] were lower (all P ＜ 0.05).The protein expressions of SOD1 and CAT in control group respectively were 0.64 ± 0.32 and 0.72 ± 0.31.Compared to the control group,the protein expressions of SOD1 and CAT in pure curcumin group (1.03 ± 0.23,1.02 ± 0.20) were significantly higher (all P ＜ 0.05) and in arsenic poisoning group (0.34 ± 0.12,0.39 ± 0.11) were lower (all P ＜ 0.05);Compared with the arsenic poisoning group,the protein expressions of SOD1 and CAT in curcumin treatment group (0.58 ± 0.09,0.68 ± 0.29) were significantly higher (all P ＜ 0.05).The arsenic content of urine in rats were positively related with arsenic content of liver and the content of MDA [correlation coefficient (r) =0.952,0.732,all P ＜ 0.05],but negativity related with the activity of SOD1 and CAT in liver (r =-0.874,-0.679,all P ＜ 0.05);the activity of SOD1 and CAT and the content of MDA in serum and liver were positively related (r =0.796,0.484,0.607,all P ＜ 0.05),the activity and protein expression of SOD1 and CAT in liver were positively related (r =0.748,0.424,all P ＜ 0.05).Conclusion The curcumin may improve the activity of hepatic antioxidant enzyme in water arsenic-exposed rats and effectively decrease lipid poroxidation damage caused by arsenic via promoting the excretion of arsenic and the protein expression of hepatic antioxidant enzyme.

Objective:To investigate the effects of different doses of sodium arsenic (NaAsO 2) on mRNA transcription levels of nucleotide excision repair (NER) related genes in normal hepatocytes (L-02 cells) and the modification levels of histone H3 ninth lysine dimethylization (H3K9me2) in the promoter region. Methods:L-02 cells were treated with 0 (the control group) , 5, 10 and 20 μmol/L NaAsO 2 for 24 h ( n = 3). Single cell gel electrophoresis (SCGE) was used to detect DNA damage [Olive tail distance (OTM) and Tail DNA percentage (Tail DNA%)] in L-02 cells. The mRNA expression levels of Xeroderma pigmentosum (XP) gene A (XPA), XP gene D (XPD) and XP gene F (XPF) were detected by real-time fluorescence quantitative PCR. The modification levels of H3K9me2 in XPA, XPD and XPF gene promoter regions (CHIP1 and CHIP2) were detected by quantitative chromatin immunoprecipitation. Results:OTM and Tail DNA% were positively correlated with arsenic doses (in the control and 5, 10 and 20 μmol/L arsenic exposure groups, the values were 0.35 ± 0.09, 0.56 ± 0.18, 3.18 ± 0.31, 4.52 ± 0.55, 0.72 ± 0.05, 1.34 ± 0.26, 3.93 ± 0.43, 5.47 ± 0.65, respectively, r = 0.927, 0.948, P < 0.05). Compared with the control group, the mRNA expression levels of XPA, XPD and XPF in L-02 cells of 10 and 20 μmol/L arsenic exposure groups were significantly lower ( P < 0.05). Compared with the control group, the enrichment levels of H3K9me2 in XPA, XPD and XPF gene promoter regions (CHIP1 and CHIP2) in L-02 cells of 20 μmol/L arsenic exposure group were significantly higher ( P < 0.05). Conclusion:Arsenic may inhibit the transcription of NER related genes by increasing the enrichment level of H3K9me2 in the promoter regions (CHIP1 and CHIP2) of NER related genes, thereby reduce the DNA damage repair ability of L-02 cells, resulting in the aggravation of DNA damage.

Objective:To establish the animal model of subchronic manganism, and to explore the effect of manganese on neurofunction of rats and the protective effect of curcumin on neurotoxicity of manganism rats.Methods:From July to December 2019, 80 SPF male SD rats were divided into 8 groups according to body weight by random number table method, which were blank control group, low, middle and high dose manganese exposure group, low, middle and high dose curcumin antagonistic group and curcumin group, with 10 rats in each group. The low, middle and high dose manganese groups were given intraperitoneal injection of 5 mg/kg, 10 mg/kg and 15 mg/kg MnCl 2·4H 2O respectively. The low, middle and high dose curcumin antagonistic groups were given 100 mg/kg, 200 mg/kg and 400 mg/kg curcumin orally along with 15 mg/kg MnCl 2·4H 2O intraperitoneal injection. Curcumin group was given 400 mg/kg curcumin orally. The rats were exposed to 5 days a week, once a day for 16 weeks. After exposure, neurobehavioral tests (balance beam test, Morris water maze, passive avoidance test) were carried out in each group. Hippocampus tissues were taken for pathological examination and oxidative stress indexes were detected. Results:The balance beam test results showed that, compared with the blank control group, the scores of balance beam of the rats in the middle and high dose manganese exposure groups increased ( P<0.05) . Compared with the high dose manganese exposure group, the balance beam scores of the low, middle and high dose curcumin antagonistic groups were decreased ( P<0.05) .The results of Morris water maze showed that, compared with the blank control group, the escape latency of middle and high dose manganese exposure groups was prolonged from the third day ( P<0.05) , and the average number of crossing the platform area of each manganese exposure group was decreased ( P<0.05) .Compared with the high dose manganese exposure group, the escape latency of the middle and high dose curcumin antagonistic groups was shortened ( P<0.05) , and the average number of crossing the original platform was increased ( P<0.05) . The results of passive avoidance test show that, compared with the blank control group, the number of errors were increased in middle and high dose manganese exposure groups ( P<0.05) . Compared with the high dose manganese exposure group, the number of errors in the passive avoidance test in the middle and high dose curcumin antagonistic groups were decreased ( P<0.05) . Pathological examination showed that the rats treated with manganses had different degrees of degeneration and necrosis of nerve cells, and the structure of nerve cells was blurred and the number of nerve cells decreased. The above phenomena were improved after curcumin antagonism. The results of oxidative stress index showed that, compared with blank control group, the activity of superoxide dismutase (SOD) decreased and the content of malondialdehyde (MDA) increased in the hippocampus of rats exposed to middle and high dose of manganese ( P<0.05) . Compared with the high dose manganese exposure group, the SOD activity increased and the MDA content decreased in the middle and high dose antagonist group ( P<0.05) . Conclusion:Subchronic manganese exposure can reduce the balance function, learning and memory ability of rats, and damage the hippocampal nerve cells in oxidative stress state. Curcumin can improve the balance function and learning and memory ability of rats with manganese poisoning, improve the hippocampal nerve damage caused by manganese exposure, and has a certain protective effect on manganese induced neurotoxicity.

Objective:To establish the animal model of subchronic manganism, and to explore the effect of manganese on neurofunction of rats and the protective effect of curcumin on neurotoxicity of manganism rats.Methods:From July to December 2019, 80 SPF male SD rats were divided into 8 groups according to body weight by random number table method, which were blank control group, low, middle and high dose manganese exposure group, low, middle and high dose curcumin antagonistic group and curcumin group, with 10 rats in each group. The low, middle and high dose manganese groups were given intraperitoneal injection of 5 mg/kg, 10 mg/kg and 15 mg/kg MnCl 2·4H 2O respectively. The low, middle and high dose curcumin antagonistic groups were given 100 mg/kg, 200 mg/kg and 400 mg/kg curcumin orally along with 15 mg/kg MnCl 2·4H 2O intraperitoneal injection. Curcumin group was given 400 mg/kg curcumin orally. The rats were exposed to 5 days a week, once a day for 16 weeks. After exposure, neurobehavioral tests (balance beam test, Morris water maze, passive avoidance test) were carried out in each group. Hippocampus tissues were taken for pathological examination and oxidative stress indexes were detected. Results:The balance beam test results showed that, compared with the blank control group, the scores of balance beam of the rats in the middle and high dose manganese exposure groups increased ( P<0.05) . Compared with the high dose manganese exposure group, the balance beam scores of the low, middle and high dose curcumin antagonistic groups were decreased ( P<0.05) .The results of Morris water maze showed that, compared with the blank control group, the escape latency of middle and high dose manganese exposure groups was prolonged from the third day ( P<0.05) , and the average number of crossing the platform area of each manganese exposure group was decreased ( P<0.05) .Compared with the high dose manganese exposure group, the escape latency of the middle and high dose curcumin antagonistic groups was shortened ( P<0.05) , and the average number of crossing the original platform was increased ( P<0.05) . The results of passive avoidance test show that, compared with the blank control group, the number of errors were increased in middle and high dose manganese exposure groups ( P<0.05) . Compared with the high dose manganese exposure group, the number of errors in the passive avoidance test in the middle and high dose curcumin antagonistic groups were decreased ( P<0.05) . Pathological examination showed that the rats treated with manganses had different degrees of degeneration and necrosis of nerve cells, and the structure of nerve cells was blurred and the number of nerve cells decreased. The above phenomena were improved after curcumin antagonism. The results of oxidative stress index showed that, compared with blank control group, the activity of superoxide dismutase (SOD) decreased and the content of malondialdehyde (MDA) increased in the hippocampus of rats exposed to middle and high dose of manganese ( P<0.05) . Compared with the high dose manganese exposure group, the SOD activity increased and the MDA content decreased in the middle and high dose antagonist group ( P<0.05) . Conclusion:Subchronic manganese exposure can reduce the balance function, learning and memory ability of rats, and damage the hippocampal nerve cells in oxidative stress state. Curcumin can improve the balance function and learning and memory ability of rats with manganese poisoning, improve the hippocampal nerve damage caused by manganese exposure, and has a certain protective effect on manganese induced neurotoxicity.