Proteomic Studies Of Cerebrum And Plasma During Focal Ischemia In Macaca Fascicularis Monkey

Stroke, as the third leading cause of mortality in the world, has been extensively studied experimentally and clinically. After years of research, scientists have come to the conclusion that the primate model is a better model for the pathological study of stroke than the rodent model. To understand the physiopathological cause of a disease, the study on cellular and molecular level is a prerequisite. Here, we designed a study by using high throughput shotgun proteomic method based on nanoLC-MS/MS to determine protein changes in brain tissue and plasma of cerebral ischemic macaca, followed by identification and analyses associated signal pathways.Non-human primate Macaca Fascicularis was used to construct focal cerebral ischemic model. After 4 h of middle cerebral artery occlusion(MCAO), tissue from ischemic site, penumbra site and normal site were collected and plasma were also collected after 4 h of MCAO and 0.5 h of reperfusion, respectively. Samples were fractionationed using strong cation exchange and identified by nano LC-MS/MS.1) Mitochondrial and nucleic proteome of ischemic and normal tissues were analyzed. Proteins associated with aerobic respiration and energy metabolism in the mitochondria were compared, as well as proteins in the nuclei associated with inflamation and apoptosis.2) Protome of normal, penumbra and ischemic tissue were analyzed. The pathways of metabolism, inflamation, apoptosis and transcription regulators were compared.3) Plasma after MCAO for 4 h and reperfusion for 0.5 h were analyzed. Brain-specific proteins were screened and compared to that of clinical samples.Results: 1) 19 and 21 proteins were identified in ischemic mitochondria and nuclei; 21 and 16 proteins in normal mitochondria and nuclei, respectively; 2) 178, 133 and 282 proteins were identified in the ischemic, penumbra and normal cytosol, respectively; 3) four brain-specific proteins were identified in the plasma after 4 h MCAO(LanC like protein 2, Protein kinase C binding protein NELL1 isoform 1, Tubulin beta 2A, Beta synuclein) and six after 0.5 h reperfusion(neurosecretory protein VGF fragment, G protein regulated inducer of neurite outgrowth 3, brain synembryn, cerebellar-degeneration-related antigen, myelin basic protein, tau-tubulin kinase).Conclusions: Primate stroke model was used to study the molecular mechanism of cerebral ischemia. It is preliminarily found that differences in metabolism, inflammation reaction and apoptosis among the ischemic, penumbra and normal tissues were significant. Several brian-specific proteins identified in the plasma could be taken as biomarkers for the stroke screening. General analytical methods were established and conclusions should be confirmed by further studies.