| Objective:(1)To study the changes of long-term potentiation(LTP) in hippocampus o f rats chronically exposed to aluminum via the pathway of PHF8-H3K9me2-BD NF.(2) To explore aluminum-induced variation of PHF8-H3K9me2-BDNF path way in SH-SY5 Y cell lines.Methods:1. In vivo study: 160 SD male rats were selected and divided into 4 batches(3,6,9,and 12 months) based on expected aluminum exposure duration and each batch contains 40 rats which were randomly divided into four groups by body weight including control group and low, medium and high dose aluminum(2,12. 2mg/kg respectively) exposed group. We measured LTP in hippocampus with electrophysiological grapier and the activity of histone demethylase(H3K9 specific)was tested with ELISA,and the gene expression of PHF8 was detected with RT-PCR,and expressions of PHF8, H3K9me2 and BDNF proteins in hippocampus were detected by Western-blot.2. In vitro study: the SH-SY5 Y cell were incubated in culture media containing Al(Cl)3 at final concentration of 0,1,2 and 4 m M for 48 hours. Cell viability was measured with CCK-8; PHF8 gene were detected with ELISA and RT-PCR respectively;the expressions of PHF8, H3K9me2, BDNF proteins were detected by Western-blot.Results:1. In vivo study:(1) There existed interactive effects among exposure time and dose(P<0.01).The differences of average of f EPSPs among groups exposed to aluminum(0,2,12,and 72mg/kg Al3+) were statistically significant(F=64.58,P<0.01;F=38.62,P<0.01;F=8.47,P<0.01;F=17.13,P<0.01)in each batch,and the average of f EPSPs decreased in a dose-dependent manner. Comparing between the controls and Al-exposed groups, the differences of average of f EPSPs among batches(3,6,9, and 12 months) were statistically significant(F=98.71,P<0.01;F=5.04,P<0.01;F=37.18,P<0.01) and the average of f EPSPs decreased in a time-dependent manner.(2) The interactive effects among exposure time and dose did not exist(P>0.05)The differences of activity of histone demethylase were statistically significant(P<0.01)between the control and Al-exposed groups. The activity of demethylase in 12,72 mg/kg Al3+ group decreased by 24.07% and 33.62%,respectively, compared with the control group. A significant difference existed among each batch and the activity of histone demethylase decreased in a time-dependent manner(P<0.01). The activity of demethylase in 9,12 months batch decreased by15.77%,15.23%, respectively, compared with the rats of the first batch(3 months).(3)The interactive effects among exposure time and dose in the change of expression of PHF8 gene did not exist(P>0.05),there was no statistically significant differences on the expression of m RNA of PHF8 among different batches and groups.(4) 1)There would exist interactive effects among exposure time and dose in the change of expression of PHF8(P<0.05).Comparing in each batch, the differences of PHF8 among aluminum-exposed groups(0,2,12,72mg/kg Al3+) were statistically significant(P<0.01)and the expression of PHF8 of 72mg/kg Al3+ group decreased by22.9%,28.02%,24.22%,27.14% compared with the control group in different batches. Further analysis shows that the statistically significant differences of PHF8 among batches(3, 6, 9, and 12months)(P<0.05)only existed in the group of 72mg/kg Al3+. 2) The interactive effects among exposure time and dose did not exist in the changes of the expression of both H3K9me2 and BDNF(P>0.05).We found that the difference of H3K9me2 and BDNF were statistically significant among the groups(both P<0.01),and the expression of H3K9me2 increased while the BDNF decreased with the dose increase. The H3K9me2 in 12 and 72 mg/kg Al3+ group increased by 13.0% and 27.55% compared to the control group, and increased by12.12% and 26.26% compared to the 2mg/kg Al3+group, respectively. However the BDNF in 12 and 72 mg/kg Al3+ group decreased by 20.88% and 30.1% compared to the control group, and decreased by 8.86% and 17.7% compared to the 2mg/kg Al3+group, respectively.Then we found that the difference of H3K9me2 and BDNF were statistically significant among the batches(P<0.05,P<0.01), and the expression of H3K9me2 increased while the BDNF decreased as the dose went up. The expression of H3K9me2 in the fourth batch(12months) increased by 10.34% compared to the first two batches(3 and 6 months), while BDNF in the last two batches(9 and 12months) decreased by 9.75% and 14.63% compared to the first batch(3 months).2. In vitro study:(1)The cell viability decreased following the increase of dose,and the differences of cell viability among the aluminum-exposed groups were significantly(P<0.01). Compared with the 0m M group, the cell viability of the 2 and4 m M decreased by 23% and 32% respectively.(2) The activity of histone demethylase deceased following the increase of dose, and the differences between each dose were statistically significant(P<0.05). Compared with 0, 1, and 2m M groups, the activity of 4m M group decreased by 24.87%, 24.47%, and 16.06%respectively.(3) No significant differences were found in the expression of PHF8 m RNA among different groups and batches(P>0.05).(4) The expression of PHF8 and BDNF decreased following the increase of dose, while the H3K9me2 increased and the differences of its expressions among aluminum-exposed groups were statistically significant(all P<0.05). Compared with the 0m M group, the expressions of PHF8 in 2 and 4m M group decreased by 12.3% and 12.9%, respectively and the expression of PHF8 in 4m M group showed a decrease of 12.9% compared with the1 m M group. The expression of H3K9me2 in 4m M group increased by 20.59% and14.71%, respectively, compared with 0,1m M group, and the expression of BDNF in4 m M group decreased by 20.68% and 13.75% respectively, compared with 0,1m M group.Conclusions:1.Chronic aluminum exposure by water drinking can suppress the induction and the maintaince of LTP in the hippocampus of rat, and the impairment would aggravate following the increase of dose and extending of treatment duration.2.Aluminum may induce the inhibition of the PHF8 activity and the reduction of the PHF8 and BDNF expressions,while H3K9me2 increases at the same time.3.Aluminum may affect the LTP in rat via the PHF8-H3K9me2-BDNF pathway.4.Aluminum may have an impact on histone methylation which then leads to changes of the protein involved in the learning and memory.This research is supported by project “Mechanism of regulating aluminum-induced neurotoxicity by histone lysine methylation modification”(NO.81372968) which funded by National Science Fundation of China. |
PDF Full Text Request