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Objective@#To investigate the role of lysosomes in manganese-induced toxicity in human neuroblastoma SK-N-SH cells.@*Methods@#SK-N-SH cells were treated with MnCl2 at doses of 0.062 5, 0.125, 0.25, 0.5, 1.0, 2.0 and 4.0 mmol/L for 24 h, and the cell viability was detected by MTT assay. Cells were treated with MnCl2 at doses of 0.125, 0.25, 0.5 and 1.0mmol/L for 24 h, and lysosomes labeled with lysotracker red were observed by laser confocal microscopy, the expression levels of LAMP1 and CTSD were detected by western blot, and CTSD activity was detected by Cathepsin D Activity Fluorometric Assay Kit.@*Results@#Compared with the control group, the survival rates of SK-N-SH cells were decreased significantly in the 0.5-4.0 mmol/L MnCl2 treatment groups (P<0.01) , the relative fluorescence intensities of 0.5 and 1.0 mmol/L MnCl2 treatment groups were increased (P<0.01) . Compared with the control group, the 0.125-0.5 mmol/L MnCl2 treatment groups had significant increase in the the expression of LAMP1 (P<0.01) . Compared with the control group, the expression of m-CTSD was significantly increased at the does of 0.125-0.25 mmol/L MnCl2, while it was decreased at the does of 1.0 mmol/L (P<0.01) . Otherwise, it wasn’t observed significant difference of the activity of CTSD between different MnCl2 treatment groups.@*Conclusion@#MnCl2 could cause cytotoxicity in SK-N-SH cells. Lysosomes may play a normal function at low doses of manganese, but they may be damaged at high doses of manganese. As an organelle that can degradate substrates in autophagy, lysosomes participate in the neurotoxic mechanism of manganese.
RESUMO
Objective@#To investigate the effect of manganese chloride (MnCl2) or 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) on the neurobehavioral and histopathology in C57BL/6 mice and provide evidence for the diagnosis, treatment and prevention of manganism.@*Methods@#Adult male C57BL/6 mice were treated with MnCl2 and MPTP respectively by intraperitoneal injection at the doses of 5, 10, 20mg Mn/kg and 30mg MPTP/kg. Controls were injected equivalent normal saline. All animals were administrated 5 times a week for 4 consecutive weeks and sacrificed after behavior tests on the fifth week. Balance ability, anxiety and depression level and cognitive function were tested respectively by vertical pole test, open field locomotion test and Morris swim task. The neuron pathological changes of striatum and substantia nigra were examined through HE-staining pathological section by using optical microscope.@*Results@#Compared with the control group, the high dose of MnCl2 reduced body weight obviously (P<0.01) . The results of vertical pole test showed that MnCl2 and MPTP lengthened the pole-climbing time and turnaround time. Open field locomotion test showed that movement distance, stand-up time and central field time were decreased after the exposure of MnCl2 or MPTP. In the Morris swim task, the escape latency time increased and the target quadrant activity time decreased significantly after the injection of MPTP as well as high-dose MnCl2, comparing with controls (P<0.05) . Moreover, the escape latency time of high dose MnCl2 prolonged prominently comparing with MPTP grou (P<0.05) . The results of histopathology showed that acidophilic changes elevated in MnCl2 and MPTP group, comparing with controls. Furthermore, in striatum the oxyphil cells number increased in MnCl2 high-dose group comparing with MPTP group (P<0.01) . On the contrary, there were more oxyphil cells in MPTP group comparing with MnCl2 groups in substantia nigra (P<0.01) .@*Conclusion@#Both manganese and MPTP can induce the impairment of dopaminergic neural system, but the symptons and injured location of manganism are inconsistent with PD models induced by MPTP.
