Protective Effects Of Mangiferin In Subchronic Developmental Lead-exposed Rats And The Study On Its Signal Transduction Mechanism

Mangiferin(MGN) is an effective antioxidant, and it was reported to chelate with metals and relieve toxicity caused by heavy metals such as mercury, cadmium and arsenic. These properties make it possible to relieve the toxicity caused by lead. Lead is a ubiquitous environmental and industrial pollutant. Exposure to excessive amounts of lead is especially harmful to the central nervous systems of infants and young children, and oxidative stress has been reported as a major mechanism of lead-induced toxicity.Recent studies have shown that Nrf2-ARE pathway has neuroprotective effect. In the central nervous system, Nrf2plays an important role by protecting neurons against oxidative stress. Nrf2promotes the transcription of a series of genes such as Glutathione-S-transferase (GST), glutamate cysteine ligase(GCLC and GCLM), NAD(P)H: quinone oxidoreductase1(NQO1), heme oxygenase-1(HO-1).In our study, molecular biological technique were used to evaluate①the influence of MGN on the structure and function of nervous system; the influence of MGN on lead burden.②The influence of MGN on lipid peroxidation and the Nrf2downstream enzymes in the cerebral cortex and serum.③the role of Nrf2-ARE pathway in the protective effect of MGN in lead-exposed rats. Major findings are as follows:Objective:To evaluate the influence of MGN on the structure, function of nervous system and the influence on the lead burden.Methods:96weaned Wistar rats were divided into six groups (n=16in each group, half male and half female):five groups exposed to500ppm of lead acetate in the drinking water and one group as blank control. After8weeks, MGN (50,100,200mg/kg body weight) and DMSA were orally administrated to intoxicated groups for four supplemental weeks; the other intoxicated group was left as lead-exposed model group. After the four weeks of administration, all rats received Morris water maze training and test. Rat brain sections were detected by HE staining and transmission electron microscopy(TEM). Blood, femur, brain, liver and kidne lead were determined by ICP-MS.Results:During the course of the Morris water maze experiment, compared with blank control group, the lead-exposed group showed no significant difference. While compared with the lead-exposed group, the MGN-treated group(200mg/kg) showed significant differences in spatial probe test(p<0.05). Under TEM, in the region CA1of lead-exposed group, the changes include neuropile vacuole, abnormal dense bodies in cytoplasm and lysosome in the peripheral vessels, pyknotic compact of gliocyte, etc. While in the MGN-treated groups, the anomaly was mild. The cells in the CA1area in hippocampus were almost normal. Compared with the lead-exposed group, MGN-treated group lowered the lead concentration in blood, bone and brain significantly (p<0.05), the effect of MGN in bone and brain showed no significant difference between DMSA group and MGN-treated group(200mg/kg).Conclusion:In our study, compared with blank control group, the lead-exposed group showed no deficit in spatial learning. It may be due to the neural compensation. MGN can improve the spatial learning in the MGN-treated group, compared with the lead-exposed group. MGN may ameliorate histopathological lesion in hippocampus. MGN can chelate with lead in vivo, that maybe due to its chelating property.Objective:To study if MGN intervene the lipid peroxidation and the Nrf2downstream enzymes.Methods:Commercial kit were used to detect H2O2, MDA, and the Nrf2downstream antioxidant enzymes, phase II detoxification enzymes and GSH related enzymes and GSH/GSSG content.Results:Lead can significantly improve the level of lipid peroxides, reduce the antioxidant enzyme activity. MGN-treated groups could significantly reduce the level of lipid peroxidation, improve the activity of antioxidant enzymes, phase II detoxification enzymes and GSH related enzymes, wherein the200mg/kg mangiferin has the most obvious effect.Conclusion:The results of the above studies showed that MGN-treated groups can improve the activity of antioxidant enzymes, phase II detoxification enzymes and GSH related enzymes, inhibit the oxidative stress induced by lead, and thus resist the lead-induced damage. It indicated that Nrf2-ARE maybe involved in the antioxidant mechanism of MGN.Objective:To study the role of Nrf2-ARE pathway in protective effects of MGN in lead-exposed rats.Methods:Real-time quantitative polymerase chain reaction(RT-qPCR), Western-blot and immunohistochemistry detection were used to detect the expression of Nrf2, GCLC, GCLM, HO-1mRNA and protein expression; Western-blot was used to detect the expression of nuclear Nrf2protein and total Nrf2.Results:RT-qPCR showed that Nrf2mRNA levels in lead-exposed group and MGN-treated groups increased. y-GCS, HO-1mRNA and protein expression were inhibited in lead-exposed group, while in MGN-treated group, they were improved significantly and were in a dose-dependent manner. In immunohistochemistry results, no Nrf2positive cells in blank control group were observed. A few in lead-exposed group were observed, while lots of Nrf2positive cells in MGN-treated group were observed. y-GCS positive cells in lead-exposed group were less than that in blank control group, the amount in MGN-treated groups increased significantly.Conclusion:Nrf2can be activated by lead, while MGN can further activate it, the regulation is not at the level of gene transcription, but may be in the post-transcriptional level.Nrf2may be the critical transcription factor when MGN intervene the expression of antioxidant genes. y-GCS、HO-1levels were raised after the activation of Nrf2, it indicated that Nrf2-ARE pathway take part in protective effects of MGN in lead-exposed rats. In summary, we can draw the following conclusions:MGN can relieve the oxidative stress caused by lead, it has neuroprotective effects to lead-exposed rats. The effects were achieved by the activation of the Nrf2downstream genes such as antioxidant enzymes, phase II detoxification enzymes and GSH related enzymes.The innovation of this study:Antioxidant mangiferin was used in the lead (Pb) study; Nrf2pathway was used to explain the neuroprotective effect of MGN.