Proteomics Analysis Of Hippocampus And Striatum In Rats With Hyperglycemia And Parkinson's Disease

Objective: This study is to investigate the differential protein expression patterns in hippocampus and striatum in rat models of Parkinson's disease(PD)and hyperglycemia(DM)-PD,and to elucidate the effects of hyperglycemia on PD pathogenesis.Methods: 1)SD rats were randomly divided into normal group(n=5),physiological saline control group(n=10),PD group(n=20),hyperglycemia group(n=20),and DM-PD(n=35);Rat models of diabetes were induced by the high-fat and high-sugar diet combined with single intraperitoneal injection of streptozotocin.Then unilateral lesion with 6-OHDA was used to establish the DM-PD rat model.Modeling time was 10 weeks.Then ethology analysis(exceptionally dependent movement,posture asymmetry,anterior limb askinesia,revolving test)was conducted on 5 groups.Tyrosine proteinase in rat hippocampus striatum was detected using immunohistochemistry(IHC)method.2)Differential proteins in the hippocampus and striatum of PD and DM-PD rats were analyzed by quantitative proteomics,with the iTRAQ technique combined with two-dimensional liquid chromatography-tandem mass spectrometry(2D LC-MS/MS).And then the Gene Ontology(GO)annotation,Cluster of Orthologous Groups of proteins(COG)and Pathway orthology have been used to analysis the bioinformation of significant differentially expressed proteins and pathways involved.3)3-5 differentially expressed proteins elected from the Differential proteins by iTRAQ was detected with Western blot analysis.Results: 1)We established DW-PD rat model successfully.There was significant differences in the parameters of exceptionally dependent movement,posture asymmetry,anterior limb askinesia,revolving test between the PD group,DM-PD group with normal group,physiological saline control group,diabetic group(P<0.01).There was significant differences in the parameters of exceptionally dependent movement,anterior limb askinesia between PD group and DM-PD group.With the extension of time,asymmetry of spontaneous behavior score and posture asymmetry increase gradually in the PD group and DM-PD group.High blood sugar was not influenced evolving ability of PD rats obviously.Four weeks after building the rat model,the nubmer of the tyrosine hydroxylase positive neurons cell on the damage side of PD group,and DM-PD group was decreased significantly compared with normal group,physiological saline control group and hyperglycemia group.In group comparison,the number of tyrosine hydroxylase reaction positive cells decreased in DM-PD group,but it was was no statistically significant difference.Model success rate of DM-PD group was 51.4%.2)Totally 4988 proteins were identified.There were 463 differentially expressed proteins between the PD and DM-PD groups,including 281 up-regulated and 182 down-regulated proteins.The GO analysis demonstrated that the cellular component of differentially expressed proteins were mostly in organelle,cytoplasm,intracellular organelle part and membrane.The main molecular functions of the differentially expressed proteins were transporter activity,enzyme binding,receptor binding.The major biological process of these proteins were including single-organism cellular process,multi-cellular organism process,single-multicellular organism process,cellular component organization or biogenesis,cellular component organization,signaling,single organism signaling.Moreover,the pathway of Dopaminergic synapse,Complement and coagulation,Gap junction,PPAR signaling pathway,MAPK signaling pathway,Wnt signaling pathway,Vascular smooth muscle contraction,Glutamatergic synapse as well as Parkinson's disease pathway were involved with these significant differentially expressed proteins.3)Proteins interaction network indicated that proteins of alpha B-crystallin,beta-crystallin A3,haptoglobin,FGG,and complexin-1 worked as the intersections.Similar results were obtained for the Western blot analysis.Conclusion: 1)Hyperglycemia can aggravate the PD rats' motor in-coordination,with the extension of time,the influence of hyperglycemia on the behavior of PD rats will be gradually increased,and high blood glucose has no significant effect on the rotation ability of PD rats.2)With the combination of iTRAQ and 2D LC-MS/MS,the main difference protein in DM-PD rat hippocampus striatum were screened effectively.there was screened 463 different proteins,including 281 up-regulated proteins and 182 down regulated proteins.The differential protein was analyzed by GO and Pathway analysis.The differentially expressed proteins may provide different perspectives and new insights for future research on the relationship between hyperglycemia and PD.3)We screened 5 differentially expressed proteins by Western Blot,which provided new evidence for the correlation between the high blood glucose and PD.4)The group of significant differentially expressed proteins(Alpha B Crystallin,beta Crystallin A3,Haptoglobin,FGG,Complexin-1)identified and verified from this study are supposed to be used as preliminary proteomic biomarkers of DM-PD.It provide clues to study on the related mechanism of high glucose with PD.