Studies Of The Treatment Of Epilepsy By Chloriti Lapis Based On Metallomics And Proteomics

This topic comes from the National Natural Science Foundation(81673566)------Mechanism studies of the treatment of epilepsy by Chloriti Lapis based on metallomics.Chloriti Lapis,which has clinical application in ancient medical book,commonly used in epileptic seizures,,irritability chest tightness and so on.Nowsdays we mainly focus on its composition analysis,and its mechanism of action on the disease is not clear.In the early stage of this group,it was proved that Chloriti Lapis had the function of epilepsy model rats,and the best mode and the dosage of administration were selected by means of biochemical index,morphological index and metabolomics technique.Based on the previous research,this paper mainly uses metallomics and proteomics technology,using the inductively coupled plasma mass spectrometry,liquid chromatography mass spectrometry and other modern analysis methods,to study and discuss the mechanism of the treatment of epilepsy infor Chloriti Lapis comprehensively,which provides reference for the development and utilization of the medicinal herbs and the target point of epilepsy treatment.The main research contents of the subject are as follows:Object:Using metallomics and proteomics techniques to study the effects of Chloriti Lapis on blood and hippocampus protein expression in normal rats,the effects of treatment of hippocampal-related proteins in rats with epilepsy induced by PTZ,and the effects on serum and hippocampal metal elements in rats.The content,at the overall animal level,clarifies the pharmacodynamic effect and possible mechanism of action of Chloriti Lapis in the treatment of epilepsy,and has certain laboratory guiding significance for the diagnosis and treatment of neurological diseases such as epilepsy.Methods and results:1.Effects of Chloriti Lapis on the identification and expression of differential proteins in blood and hippocampus of rats under non-pathological conditions:the identification and differential expression of protein in hippocampus and serum of rats under non-pathological conditions were compared by using non-marker(label free)quantitative proteomics technique.By comparing with the blank group,it was found that Chloriti Lapis had a certain effect on hippocampal protein in the nervous system of rats under non-pathological condition,and the data related to differential protein were obtained.This provides important information for further study of the effects of Chloriti Lapis on protein changes in rats epilepsy model in the later stage.2.Study on the identification and relative expression of differential proteins in the hippocampus of PTZ-ignited epileptic rats by Chloriti Lapis:by comparing the differential proteins between the blank group,the model group and the group after the treatment of Chloriti Lapis,the protein of Chloriti Lapis treatment in rats was determined comprehensively in detail.Based on the effect of the first experimental Chloriti Lapis on the identification and expression of differential proteins in the blood and hippocampus of rats under non-pathological conditions,the protein expression in hippocampus of rats was determined by quantitative proteomics of series quality labels(tandem mass tag,TMT)according to the characteristics of sample volume and quantitative protein analysis.And the qualitative and absolute quantitative analysis of specific proteins was carried out.The results showed that there were 28 proteins that were significantly up-regulated in the Mo/Co group,and one was significantly down-regulated.In the Me/Mo group,21 are up and 7 are down.3.Study on the qualitative and absolute quantitative expression of differential proteins in the hippocampus of PTZ-ignited epileptic rats by Chloriti Lapis:based on the second experiment,Study on the identification and relative expression of differential proteins in the hippocampus of PTZ-ignited epileptic rats by Chloriti Lapis,the early differential proteins were evaluated and tested,and the repeated proteins were selected for accurate location analysis.In this experiment,6 peptide segments of 4 target proteins of hippocampal tissue samples of rat species were quantitatively analyzed LC-PRM/MS,the target peptide segment had quantitative information in 6 samples,and the quantitative information was normalized by isotope heavy-labeled peptide segment,and the target peptide segment and target protein were analyzed relatively quantitatively.The results of differential multiples and TTEST test show that there are some differences in the expression of 4 kinds of target proteins under two different conditions,Mo and me.4.Study on some indexes in serum of PTZ-ignited epileptic rats:the model of rats with epileptiy was reproduced,and the index each groups of Bcl-2,caspase-3,Bax,p-JNK,p-p38,Tspan2,MbP and ERK were detected by ELISA.The content changes between the groups after the treatment of the model group and the Chloriti Lapis,and the correlation analysis was carried out,and the results showed that:the content of Bcl-2 in epilepsy Group was significantly lower than that in blank group,and the content of caspase-3 and Bax was significantly higher than that of Blank group,which indicated that there was excessive apoptosis of neurons in epileptic group rats.After the administration of Chloriti Lapis,the content of Bcl-2,Bax and caspase-3 got better,indicating that Chloriti Lapis administration may alleviate neuronal excessive apoptosis,but also participate in MAPK signaling pathway through ERK and p38 pathway to play a role.5.Effect on the metallic elements in rat Serum and hippocampus:the model of epilepsy in rats was reproduced,the content of metal elements in serum and hippocampus was analyzed by ICP-MS and other methods,and the correlation analysis between the elements was carried out.At the same time,the metal elements were correlation with the difference indexes in serum and hippocampal tissues.Compared with the model group,the 4 elements of K,Zn,Ni and Pb in hippocampus and serum,both blank group and drug delivery treatment group increased significantly(p<0.05),while Si and Sr those elements in hippocampus and serum,both blank group and drug delivery treatment group significantly decreased(p<0.05).Compared with the blank group,the Al,Si,V and As elements in the hippocampus model group increased significantly(p<0.01),while the Chloriti Lapis administration decreased significantly(p<0.05).In the blood,the As element decreased significantly(p<0.01)compared to the blank group,while the content of the drug was significantly increased after administration treatment(p<0.01).6.Study on the morphology of arsenic in the hippocampus with PTZ ignited epilepsy by Chloriti Lapis:the model of rats was reproduced to ignite epilepsy,and the arsenic morphology of metallic elements in serum and hippocampus was correlation analyzed with the difference index in hippocampal tissue by using HPLC-ICP-MS and other methods between blank group,model group and Chloriti Lapis administration.Conclusion:The morphology of strongly toxic arsenic in hippocampus may be transformed into less toxic DMA morphology remaining in the hippocampus,part of which is metabolized into the blood,resulting in an increase in the total amount of arsenic in the blood,which is combined with the caspase-3 index,inhibits caspase-3 activation,reduces apoptosis at the same time,directly participate in ERK pathway action and correlated p-p38 highly,to reduce its content,thereby reducing Tspan2 content,apoptosis,and epileptic seizures.