Effect Of PGDS-DP Signaling Pathway On Primary Cultured Rat Hippocampal Neuron Damage Induced By Aluminum Salt

Objective:(1) To observe the change of PGDS-DP signaling pathway in primary cultured rat hippocampal neuron treated with aluminum overload.(2) To study the effect of intervention for PGDS-DP signaling pathway on primary cultured rat hippocampal neuron treated with aluminum overload.Methods:(1) Change of PGDS-DP signaling pathway in primary cultured rat hippocampal neuron treated with aluminum overloadThe hippocampus was dissected out from fetal rat (embryonic 18d). After being cultured for 7d, the purity of neuron was identified by immunocytochemistry of neuron specific enolase and treated with aluminum maltolate (100μM) to establish the model of primary cultured rat hippocampal neuron damage (neuron in blank control group was treated with PBS and neuron in control group was treated with 300μM maltolate). After treatment with Al (malt)3 for 24 hours, the cell viability was measured by MTT and lactate dehydrogenase leakage rate of cell was detected by LDH. Furthermore, neuronal pathomorphology was observed by HE staining and content of PGD2 was detected by the method of ELISA. Additionally, the expressions of L-PGDS and DP receptors mRNA were measured by RT-PCR as well as the expressions of protein were measured by Western-Blotting.(2) Effect of intervention for PGDS-DP signaling pathway on primary cultured rat hippocampal neuron treated with aluminum overloadThe hippocampus was dissected out from fetal rat (embryonic 18d), After being cultured for 7d, the hippocampal neuron was treated with Al (malt)3 to establish the model of primary cultured rat hippocampal neuron damage and meanwhile treated with DPI agonists BW245C, DPI antagonists BWA868C, DP2 agonists DK-PGD2 and DP2 antagonists CAY 10471,respectively. After treatment for 24 hours, the cell viability was measured by MTT and lactate dehydrogenase leakage rate of cell was detected by LDH. Furthermore, neuronal pathomorphology was observed by HE staining. The level of Ca2+was detected by Fluo-3/AM.Results:(1) The purity of primary cultured rat hippocampal neurons was more than 95%. Compared with the blank control group, the number of hippocampal neurons was reduced and neurons became chromatic agglutination and karyopyknosis in aluminum overload group. Treatment of aluminum caused a significant decrease of cell viability and an increase of the LDH leakage rate and the PGD2 content in hippocampal neuron. Treatment of aluminum also induced a significant increase of mRNA and protein expressions of L-PGDS and DPI and a decrease of the DP2mRNA and protein expressions in hippocampal neuron.(2) Compared with that of the control group, the Ca2+ fluorescence intensity significantly increased in aluminum overload group.The administration of BW245C, a selective agonist of DPI, could significantly alleviate the primary cultured rat hippocampal neuron injury and blunt the increase of Ca2+fluorescence intensity caused by aluminum overload. While the BWA868, a selective antagonist of DP1, enhanced the primary cultured rat hippocampal neuron injury caused by aluminum overload. However, BWA868 had no significant effect on Ca2+ fluorescence intensity of aluminum overload hippocampal neuron. Treatment of DK-PGD2, a selective DP2 agonist, aggravated significantly the primary cultured rat hippocampal neuron injury caused by aluminum overload accompanied with the increase of Ca2+ fluorescence intensity of neuron. Treatment of CAY 10471, a selective DP2 antagonist, had opposite effects of DK-PGD2.Conclusions:(1) The expressions of L-PGDS and DP1 and the content of PGD2 were significantly increased in primary cultured rat hippocampal neuron caused by aluminum overload.While the expression of DP2 was significantly decreased.(2) The expression and activation of DPI can decrease the susceptibility of hippocampal neuron to injury caused by aluminum overload. However, the expression and activation of DP2 can increase the susceptibility of hippocampal neuron to injury caused by aluminum overload.(3) The PGDS-DP signaling pathway involves in the mechanism of damage to primary cultured rat hippocampal neuron induced by aluminum overload.