| Objective :In order to search the differences of the substantia nigra protein between the mouse model group of Parkinson's disease(PD) which were injected (1mg/kg/d) rotenone and control mouse grou which were injected (1mg/kg/d) physiology saline by proteomics technology。We find biomarkers to diagnose PD,the target of treatment and disease-specific proteins of PD. In order to identify these different proteins with MAILD-TOF-MS. As a result,we confirm the disease-specific proteins (DSPs) of PD by the data searched.Methods The test group adopted the mouse(C57BL/6) model of chronic PD which was accepted bake hypodermic injections of retenone and the control group which was accepted the same way injections of physiology saline. Observed if there would appear parkinsonism symptoms on the test group and control group after injected at different periods of time. Behavioral methods and immunohistochemistry were used to conclude the success or failure of the mouse model. Extracted protein from the test group and control group mice's substantia nigra. Pured protein with TCA/acetone. Used these protein to run immobilized IPG-IEF, and then SDS-PAGE. The protein spots were stained through silver in gels. Scanned gels and analyzed the images. Searched for the differential expression protein spots by comparing TG and CG of gel images. Identified some of the great differential expression protein spots with MS (mass spectrometry) after digested with enzyme.Results The results of behavior observation: compared with the control group, open field,the pole climbing time and swim test of the test group was obviously lasting(p<0.05).The test group which received with rotenone appeared markedly hypoactive parkinsonism behavior and the control group didn't appeared. Through the comparison of2-DE mappings(pH3-10 IPG DryStrip)of the mouse substantia nigra between the test group and control group. There were 711±17 protein spots on the substantia nigra 2-DE images of test group in the range of pH3-10, and 674±12 on the controls,(through the analysis of protein spots of two groups with PDQuest8.0 analysis software), 11 protein spots were hyp-expression and 7 protein spots were low-expression on the substantia nigra 2-DE mappings of PD by comparing the mappings of two groups,two protein spots were new. Through the MS identifition, the low-expression proteins spot in substantia nigra were Ubiquitin-like protein 3 precursor. The new protein spots were Coiled-coil domain-containing protein85A(CC85A), Centriolin(CP110). The hyp-expression protein spots includedα-enolase and the two new protein spots. Conclusion Rotenone chronic parkinsonism mouse model could better represent natural couse of Parkinson's disease than others.α-synuclein could express in substantia nigra through immunohistochemistry. The ideal substantia nigra 2-DE mappings could be obtained with pH3-10 IPG DryStrip, both IPG-IEF and vertical flat SDS-PAGE.We accepted different protein expressions on the substantia nigra 2-DE mappings in PD and controls, some protein spots were hyp-expression and others low-expression. We would further study and learn the function of protein spots in the pathogenesy and progression of PD by identifying the function messages of them.Though proteomics technology selected biomarkers to diagnose PD,the target of treatment and disease-specific proteins of PD. |
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