| Lead is difficult to degrade in its natural state.As industry continues to grow,lead mining continues to rise and lead accumulates in the environment,lead poisoning has become an increasingly serious public safety issue.The animal reproductive system is very sensitive to environmental heavy metals and continued elevated levels of lead in the environment can seriously affect the health of the mammalian reproductive system.Lead poisoning causes damage to the body mainly through oxidative stress.Melatonin,as a potent antioxidant,can effectively reduce oxidative damage to oocytes when added to oocyte maturation cultures in vitro,and little research has been done on melatonin in the treatment of oocyte lead poisoning.Therefore,it is of great interest to investigate the effect of melatonin on the in vitro maturation of oocytes and early embryonic development in lead-poisoned mice.In this study,6-8 week old ICR female rats were used as study subjects and were divided into melatonin-treated,lead poisoning control and saline control groups according to their treatments.In vitro maturation,in vitro fertilisation and early embryonic development tests were performed on the oocytes of each group.The results showed that after 16-18 h of in vitro maturation,the first polar body expulsion rate of lead-poisoned oocytes was significantly lower compared to control oocytes(p<0.01),and in vitro treatment with 10-7 mol/L melatonin significantly increased the polar body expulsion rate of lead-poisoned oocytes(p<0.01).To further verify the effect of melatonin on the function of lead-poisoned oocytes,this study was followed by in vitro fertilization and early embryonic development tests on mouse oocytes.The results showed that the oocyte cleavage rate and blastocyst rate were significantly lower in the oocytes after lead poisoning than in the control group(p<0.05),and the oocyte cleavage rate was significantly higher and the blastocyst rate increased significantly after treatment with 10-7 mol/L melatonin in the culture medium compared to the lead poisoning group.Combining the above tests,10-7 mol/L was screened as the optimum in vitro melatonin addition concentration for subsequent oxidative stress related parameter determination.By examining ROS levels,GSH content,mitochondrial distribution,ATP content,spindle morphology and the occurrence of early apoptosis in oocytes,it was found that lead poisoning significantly increased ROS levels,abnormal mitochondrial proportions and early apoptosis rates in oocytes,and significantly decreased GSH content,ATP content and normal spindle proportions,causing oxidative stress in oocytes.Treatment with melatonin at 10-7 mol/L significantly reduced these negative effects of lead poisoning on oocytes.Subsequently,the experiment measured the expression of Nrf2-Keapl-related pathway genes in oocytes by RT-PCR.The results confirmed that oxidative stress caused by lead poisoning activated the Nrf2-Keap1 pathway,increased the expression of the pathway genes Nrf2 and Keapl genes and decreased the expression of the antioxidant enzymes CAT and SOD-1.Melatonin treatment at 10-7 mol/L up-regulated the expression of Nrf2,CAT and SOD-1 and down-regulated the expression of Keapl in lead-poisoned oocytes,reduced the oxidative stress of lead poisoning on oocytes,improved the quality of oocytes after lead poisoning and promoted normal oocyte maturation in vitro,in vitro fertilization and early embryonic development.In conclusion,melatonin at 10-7 mol/L can reduce the level of oxidative stress in oocytes of lead-poisoned mice,improve mitochondrial function,ensure normal meiosis,enhance in vitro maturation of oocytes,in vitro fertilization and early embryonic development,and provide a corresponding reference and reference for the improvement and repair of reproductive function in lead-poisoned animals. |
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