| Background:With the development of industry and the increase of human activities,increasingly cadmium(Cd)pollution has gradually become a dangerous factor threatening human health.Epidemiological studies showed that people exposed to Cd mainly by smoking and inhaling polluted air,which could result in the accumulation of Cd in the respiratory system especially in the lungs and lead to lung injury and dysfunction.It was reported that Cd exposure could result in lung injury by activating oxidative stress,inflammatory reaction and apoptosis pathway in lung tissues and cells.Ferroptosis is a type of programmed cell death which is iron-dependent and lead to lipid peroxidation.Ferroptosis had been proved to participate in the pathological process of many diseases and was associated to the biological toxicity of several environmental pollutants.However,whether ferroptosis was an important mechanism in the process of lung injury caused by Cd exposure remained to be unknown.Melatonin was reported as a good antioxidant and could alleviate brain injury caused by cadmium,but the protective effects and mechanism of melatonin on cadmium-induced lung epithelial cells injury were still unclear.Objective:This study explored the lung toxicity mechanism of heavy metal Cd and investigated the pharmacological activity of melatonin in inhibiting ferroptosis,so as to provide new ideas and medication guidance for prevention and treatment of lung injury caused by Cd exposure.Methods:BEAS-2B cells were used experimental subjects,and cell staining models were established.(1)In this study,human bronchial epithelial cells line(BEAS-2B)was used as the experimental object to establish Cd exposure model on cells.(1)BEAS-2B cells were treated with culture medium containing different concentrations of Cd(0μM,5μM,10μM,20μM),then the cells were observed by light microscope;CCK-8 was used to test the cell viability;The level of oxidative stress was tested by measuring the content of ROS and MDA and the activity of SOD;The iron level were investigated by iron content measurement;the protein level of GPX4 was measured by Western blot;The m RNA levels of TFR and FPN1 were tested by RT-PCR experiment;The morphology of mitochondria was observed by electron microscope.(2)According to the results of the above part,20 μM of cadmium was chosen as the optimal treatment concentration for further experiments,the cells were pretreated 0μM Fer-1and 5μM Fer-1 respectively,and then cells were treated by culture medium containing0μM Cd and 20μM Cd.After exposure,the above experimental methods were used to test the cell morphology,cell viability,oxidative stress level,GPX4 protein level and mitochondrial ultrastructure under microscope.(3)The cells were treated with culture medium containing 0μM Cd+0μM melatonin,20μM Cd,20μM melatonin and 20μM Cd+20μM melatonin respectively.After exposure,the microscopic morphology,cell viability,oxidative stress level,iron content,GPX4 protein level,TFR and FPN1 m RNA level and mitochondrial ultrastructure of the cells were measured by the above methods.Results:(1)Cd exposure reduced the activity of BEAS-2B cells,induced the increase of oxidative stress level and iron content of cells,decreased the level of GPX4 protein,increased the level of TFR m RNA and decreased the level of FPN1 m RNA,and led to the characteristic changes of ferroptosis in mitochondria of cells in a dose-dependent manner.(2)Fer-1 effectively inhibited the decrease of cell viability,the increase of oxidative stress level,the decrease of GPX4 protein level and the characteristic changes of mitochondrial ferroptosis in BEAS-2B cells induced by Cd exposure.(3)Melatonin effectively inhibited the increase of oxidative stress level,the decrease of cell viability,GPX4 protein level,FPN1 m RNA level and the characteristic changes of mitochondrial ferroptosis of BEAS-2B cells caused by Cd exposure.Conclusion: Cd exposure led to the injury of BEAS-2B cells by inducing ferroptosis,which could be alleviated by melatonin. |
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