Comparative Proteomics Profiling Of Pallium In Focal Cerebral Ischemic Rat And The Effect Of Acupuncture

[Objective] Proteome, which was named after genome, is a major theme in future life science research. Rapid development in protein research is expected after completion of the human genome sequence. However, proteins are not merely the downstream expression of genes but are regulated in cells by a balance in their generation, modification and degradation under systematic stimuli of the influence of homeostasis. An insight into protein mechanisms involved in disease is critical to accelerate the development of specific diagnostic and prognostic markers, neuropsychiatric disease markers, and the corresponding therapeutic tools. Conventional and functional proteomics have significant potential to expand our understanding of cerebral ischemia but have not yet been used. The purpose of the research was to screen alteration in protein level in cerabral cortex of MCAO rats and "Xing Nao Kai Qiao" acupuncture - treated rats.[Mathods] The rats were divided in five groups: Normal group, sham operated group, MCAO model group, acupuncture group and control group. Each group was divided into two inferior groups: 6h group and 24h group. Ischemia was induced with a 00.26 or 0.28 nylon monofilament as Longa described previously. Selecting Nei Guan, Ren Zhong points on acupuncture group and Qu Chi, Zu Sanli points on control group, electric needle parameter as: frequency:2HZ, electric current: 3mA, time: 10min. Each rat was decapitated rapidly in defined times. The paiiium proteis were analyzed by 2-DE using an immobilized pH gradient (IPG) for the first isoelectric focusing electrophresis, and software of Image master 2D Elite v3.01 as the program for data analysis. The proteins of interest were digested in-gel and identified using MALDI-TOF mass spectrometry.[Results] Part I Improving of 2-DE technique of the rat pallium proteins.In order to produce the best gels, many factors must be considered, including sample reparation methods, the isoelectric focusing range, the type of protein staining, the detection capabilities of the hardware employed for imaging as well as the way in which the sample is applied to an IPG strip and the amount of protein loaded. We improved the2-DE technique of rat pallium proteins and made the foundation of the experiment.Part II Preliminary proteomic study of rat pallium.Pallium proteins of three Wistar rats were separated by 2-DE, identified using MALDI-TOF-MS and 84 prominent proteins with various functional characteristics were identified. These proteins included housekeeping, signaling, cytoskeletal, intermediary metabolism, antioxidant proteins on the one and neuron, synaptosomal specific proteins on the other hand. The database of rat pallium proteins was initially established.ParIII Comparative proteomic profiling in MCAO rat pallium.Comparative proteome analysis of MCAO rat pallium was performed using 2-DE combining with MALDI-TOF-MS to identify proteins altered expression level. In MCAO 6h group, the levels of 11 proteins were significantly increased, 15 proteins were significantly decreased and 2 proteins were detected only in MCAO 6h group, 1 protein only in normal control group. In MCAO24h group 13 proteins were significantly increased, 16 proteins were significantly decreased and 2 proteins were detected only in MCAO 24h group.Part IV Comparative proteome analysis of cerebral cortex from "XingNaoKaiQiao" acupuncture treated rats.In "XNKQ"6h group, 17 increased proteins were identified as Superoxide dismutase [Cu-Zn], Antioxidant protein 2, Myelin basic protein S, Phosphoglycerate mutase l,et al and 7 decreased proteins were identified as Voltage-dependent anion-selective channel protein 1, Ubiquitin carboxyl-terminal hydrolase isozyme LI, 14-3-3 protein tau, Cytoch-rome c oxidase polypeptide Vb, mitochondrial [Precursor], Peptidyl-prolyl cis-trans isomerase A, Alpha enolase, Serine-threonine kinase receptor-associated protein.In "XNKQ"24h group, 10 increased proteins were identified as Superoxide dismutase [Cu-Zn], Antioxidant protein 2, Phosphoglycerate mutase 1, Glyceraldehyde-3-phosphate dehydrogenase, Stress-70 protein [mitochondrial precursor], Tissue-type plasminogen...