The Effects Of Lead Exposure On The AD-related Pathophysiological Changes And Its Mechanism At Different Life Stages

Alzheimer’s disease (AD) is a highly prevalent, progressive, and fatalneurodegenerative disease associated with aging. But so far, the pathogenesis of ADare still unclear, many hypotheses were established on the pathogenesis of AD, theaggregation and accumulation of Aβ is a common factor and key link to ADpathogenesis.Therefore it is the focus of the research today on the developmentprocess and its regulation mechanism of Aβ.Lead is widely recognised as a potent toxin to the central nervous system thatcan affect the developing brain by altering cognition, memory, and behaviouralcontrol. Epidemiological investigations have established that lead is an environmentalfactor that is suspected to contribute to neurodegenerative diseases, such as AD、Parkinson’s disease and Amyotrophic lateral sclerosis. Furthermore, studies in rodentshave demonstrated that lead exposure at brain developmental stages affected theexpression and regulation of the amyloidogenic product.Based on the above evidence, we hypothesised that lead exposure during specificlife stages may exert varied degrees of neurotoxic effects, including neurobehavioraldysfunction and neurodegenerative changes; lead exposure may effect Aβ metabolismto cause AD-like pathogenesis.Objectives1. To investigate the effect of early lead exposured on spatial learning andmemory capacity, the animal mode of lead exposured were established at early lifestages, the morris water maze tests was used to evaluate the changes of spatiallearning and memory capacity with aging. 2. To explore the relationship between lead exposured and AD,the levels of APP,BACE1, PS1expression were comparaed in mice after lead exposure.3. To explore the Potential mechanism of lead exposured, the study evaluated theeffect of early lead exposure on the global DNA methylation level. Furthermore, theDNA methylated levels of the promoter of APP gene, PS1gene were analysised.Materials and methods1. SubjectsA total of90adult C57BL/6mice were mated at a2:1ratio. After delivery, thelitters were normalised to6pups per litter. During the lactation period, the pups wererandomly separated into four groups: control, early exposure, late exposure, orlong-term exposure. These groups were not exposed to lead, exposed to lead frombirth to21d, exposed to lead from mouth6to mouth8, or exposed to lead from birthto21d and mouth6to mouth8of age, respectively. Lead exposure was induced byproviding Pb-contaminated drinking water at a concentration of0.2%.2. Methods2.1. The lead levels in their blood and hippocampus were measured via graphitefurnace atomic absorption spectrometry.2.2. Their spatial learning and memory capacity was evaluated via the Morriswater maze test at the different age2.3The mRNA level of APP, PS1, BACE1were tested by Real-time PCR in thehippocampus.Then, The protein expression of APP, PS1, BACE1in the hippocampuswas detected via Western blot.2.4The levels of Aβ1-40and Aβ1-42in hippocampus tissues of mice after leadexposure were detected by Elisa method.2.5The levels of global DNA methylation of lead exposed mice were detectedby HPLC method.2.6the levels of APP gene and PS1genepromoter methylation status wereanalysed with MS-PCR technique.3. Statistical AnalysisAll data were expressed as the means±SD. One-way ANOVA was performedusing SPSS21.0software to analyse the differences between the groups. P values less than0.05were considered to be significant.Results1. Effects of lead exposure on blood and the hippocampal lead content andspatial learning and memory1.1Lead content in the blood and the hippocampal tissueThe results revealed that the hippocampal and blood lead levels weresignificantly higher in all of the groups exposed to lead than the control group. Thelead levels in the blood and hippocampal were increased in the DP group and theDAP group at1months and at6months of age. Significant increases in the lead levelwere found in the blood and the hippocampus from all of the lead-exposed groupscompared to the control group at12months and at18months of age. The leadcontents in the blood and the hippocampus from the DAP group were significantlyhigher than those from the DP group and AP group.1.2Effect of lead exposure on spatial learning and memoryThe results showed that the lead-exposed groups exhibited a significantdifference in the escape latency compared to the control group beginning on thesecond day (P<0.05). Escapes latencies in both DP group and DAP group of micewere significantly longer than those of the control group from day3to day5.Compared with the AP group of mice, DP group and DAP group of mice requiredmore time to reach the platforms;The escape latencies of the mice in both the DPgroup and the DAP group were significantly longer than those of the control group at12months and at18months of age(P<0.05). The number of platform crossingswas significantly decreased in the lead-exposed groups. The lead-exposure mice hadless frequency of crossing the platform (P<0.05).2. Effects of lead exposure on the production of Aβ2.1The results of Real-time PCR showed that lead exposure significantlyincreased the APP, PS1, BACE1mRNA level in the hippocampus in lead-exposuredmice compared to controls. The expressions of APP, PS1, BACE1mRNA weresignificantly higher in DP and DAP groups than the C groups(P<0.05).2.2The results of Western blot showed that protein expression of APP, PS1, BACE1were increased in the hippocampus in lead-exposured mice compared tocontrols. Results showed that lead exposure affected APP protein level in12-monthmale mice. Furthermore, APP levels were also detected after lead exposure in18-month old male mice. lead exposure for a period of three months at different lifestages significantly increased APP and BACE1in the lead-exposured mice(P<0.05).2.3The results showed that the concentrations of Aβ1-40and Aβ1-42wereincreased in lead-exposured mice as compared to controls at12months and18months of age. The levels of Aβ1-40and Aβ1-42were higher in lead-exposured micethan the control mice. Moreover, the levels of Aβ1-40and Aβ1-42were significantlyhigher in DP groups of mice, the DAP groups of mice and the AP group of mice.Significant differences between DP and DAP groups were observed at12months and18months of age(P<0.05).3. Effects of lead exposure on the DNA methylation3.1The results showed that DNA methylation level were gradually reduced from1months to18months of age. The levels of methylation were lower in leadexposured mice than the control mice at different age(P<0.05)3.2The DNA methylation level of APP gene, PS1gene promoter were low inlead-exposured mice at different age. The DNA methylation level of APP genepromoter was statistically significant difference between the C group and the leadexposured groups (P<0.05). The DNA methylation level of PS1gene promoter inlead exposured groups were lower than the C groups, But there were no statisticallysignificant difference in lead exposured groups(P﹥0.05).Conclusion1. Pb ingestion resulted in a significant elevation in the lead content in the blood,as well as lead accumulation in the hippocampus. Lead excretion is very slowly.2. Lead exposure can damage the ability of spatial learning and memory in mice,lthe latent effects of lead exposure play a major role in the decreased capacity ofspatial learning and memory.3. Lead exposure can cause excessive expression of APP、BACE1and PS1inhippocampus of mouse. This study suggests that lead exposure at early life stages influences the production of Aβ, which are associated with an AD-like pathology.4. Lead exposure can decrease the levels of global DNA methylation.