Abstract
BackgroundSeveral proteins are known to be markedly expressed in the brain during cerebral ischemia; however, the changes in protein profiles within the ischemic brain after an ischemic insult have not been fully elucidated. We studied the changes in the ischemic brain proteome after focal cerebral ischemia, induced by middle cerebral artery occlusion (MCAO) in mice.MethodsLCMS-IT-TOF mass spectrometry was used to detect the changes in ischemic brain protein patterns after MCAO. We evaluated the protein expression detected in the ischemic area, by western blotting and immunohistochemistry.ResultsNine unique proteins were identified from the ischemic area at 10 h after ischemic insult. Among these proteins, we focused on Rab33b, a member of RAS oncogene family and we found that Rab33b was up-regulated in the ischemic striatum and the number of Rab33B-positive cells increased in a time-dependent manner. Rab33B colocalized with Iba-1 positive microglia in the ischemic area.ConclusionThese findings suggest that LCMS-IT-TOF is useful for identifying changes in proteins after cerebral ischemia and that Rab33B is partially related to the pathogenesis of transient cerebral ischemia in mice.
Highlights
Several proteins are known to be markedly expressed in the brain during cerebral ischemia; the changes in protein profiles within the ischemic brain after an ischemic insult have not been fully elucidated
From the NanoRPLC-MS/MS analysis, we identified nine unique proteins that were detected only in the ischemic area (Figure 2C)
The number of Rab33B-positive cells was increased in a timedependent manner from 1 h to 10 h following the 2 h ischemia-reperfusion treatment
Summary
Several proteins are known to be markedly expressed in the brain during cerebral ischemia; the changes in protein profiles within the ischemic brain after an ischemic insult have not been fully elucidated. Since stroke is one of the principal etiologies for neurological sequelae and/or death, it is very important to understand both its pathologic mechanisms and any effective treatments. The identification and characterization of the ischemic area are valuable and useful for understanding the pathogenesis and for establishing new therapeutic strategies to treat or prevent brain ischemia. Shimadzu’s new LCMS-IT-TOF is a novel hybrid mass spectrometer that is applicable to a wide range of bioanalytical research, including biomarker discovery, metabolite identification, and drug development, among others. The LCMS-IT-TOF allows more qualitative information about a sample to be collected in a single run. This enables researchers and scientists to: 1. The ion trap is used to focus ions prior to ejection into the TOF as well as to support MSn analysis with effective precursor ion selection capabilities (resolution > 1,000 at 1,000 m/z)
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