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Global proteomic analysis of brain tissues in transient ischemia brain damage in rats.

Chen JH, Kuo HC, Lee KF, Tsai TH - Int J Mol Sci (2015)

Bottom Line: It mediated the effects of SAM administration on the apoptotic and ER stress pathways.Our results demonstrate that the ischemic injury of neuronal cells increased cell cytoxicity and apoptosis, which were accompanied by sustained activation of the IRE1-alpha/TRAF2, JNK1/2, and p38 MAPK pathways.Based on these results, we also provide the molecular evidence supporting the ischemia-reperfusion-related neuronal injury.

View Article: PubMed Central - PubMed

Affiliation: Institute of Traditional Medicine, School of Medicine, National Yang-Ming University, Taipei 112, Taiwan. chenjiannhwa@yahoo.com.tw.

ABSTRACT
Ischemia-reperfusion injury resulting from arterial occlusion or hypotension in patients leads to tissue hypoxia with glucose deprivation, which causes endoplasmic reticulum (ER) stress and neuronal death. A proteomic approach was used to identify the differentially expressed proteins in the brain of rats following a global ischemic stroke. The mechanisms involved the action in apoptotic and ER stress pathways. Rats were treated with ischemia-reperfusion brain injuries by the bilateral occlusion of the common carotid artery. The cortical neuron proteins from the stroke animal model (SAM) and the control rats were separated using two-dimensional gel electrophoresis (2-DE) to purify and identify the protein profiles. Our results demonstrated that the SAM rats experienced brain cell death in the ischemic core. Fifteen proteins were expressed differentially between the SAM rats and control rats, which were assayed and validated in vivo and in vitro. Interestingly, the set of differentially expressed, down-regulated proteins included catechol O-methyltransferase (COMT) and cathepsin D (CATD), which are implicated in oxidative stress, inflammatory response and apoptosis. After an ischemic stroke, one protein spot, namely the calretinin (CALB2) protein, showed increased expression. It mediated the effects of SAM administration on the apoptotic and ER stress pathways. Our results demonstrate that the ischemic injury of neuronal cells increased cell cytoxicity and apoptosis, which were accompanied by sustained activation of the IRE1-alpha/TRAF2, JNK1/2, and p38 MAPK pathways. Proteomic analysis suggested that the differential expression of CALB2 during a global ischemic stroke could be involved in the mechanisms of ER stress-induced neuronal cell apoptosis, which occurred via IRE1-alpha/TRAF2 complex formation, with activation of JNK1/2 and p38 MAPK. Based on these results, we also provide the molecular evidence supporting the ischemia-reperfusion-related neuronal injury.

No MeSH data available.


Related in: MedlinePlus

A close view of selected protein spots representing differentially expressed SAM-related proteins between in control and SAM-operated rats. The proteins with expression compared with SAM group are shown. (A) Nine proteins were consistently down-regulated; (B) Six identified proteins showed a greater-than-threefold change in density. The red arrows correspond to that in Table 1. Six pairs of brain protein extracts were evaluated, and the results shown are derived from one pair of experiments; fifteen reproducible blots were performed in total.
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ijms-16-11873-f003: A close view of selected protein spots representing differentially expressed SAM-related proteins between in control and SAM-operated rats. The proteins with expression compared with SAM group are shown. (A) Nine proteins were consistently down-regulated; (B) Six identified proteins showed a greater-than-threefold change in density. The red arrows correspond to that in Table 1. Six pairs of brain protein extracts were evaluated, and the results shown are derived from one pair of experiments; fifteen reproducible blots were performed in total.

Mentions: The proteomic profile of the cortex region was used to examine the events of SAM-induced brain infarction and stress responses. In this study, the protein of the cell was extracted from the control group and SAM-treated samples (n = 6 for each group). Protein samples were prepared, and more than 400 protein spots were viewed. Using silver-staining and 2D-PAGE analysis, several differences were observed with transient global ischemia in the six pairs of rats. Protein expression gel profiles (six pairs from both rat groups) were compared using ImageMaster software (Canon A640, Tokyo, Japan). The focus was then on identifying proteins found to be statistically significant at more than threefold after ischemic injury. When proteins were resolved by SDS-PAGE electrophoresis on a 12% gel, protein spots appeared to be differentially expressed more than three times greater in the SAM group than in the control group (Figure 2). In SAM rats, nine proteins were consistently down-regulated (Figure 3A), while six identified proteins showed a greater-than-threefold change in density (Figure 3B). All spots were subjected to peptide fingerprint identification using MALDI-TOF/TOF. Fifteen proteins were successfully identified by MALDI-TOF/TOF Mass Spectrometry (Table 1). Zoomed views of representative gel regions depicted several differentially expressed proteins related to oxidative stress, inflammatory response and apoptosis-related proteins. Statistical analyses were conducted after quantification of the differentially expressed proteins.


Global proteomic analysis of brain tissues in transient ischemia brain damage in rats.

Chen JH, Kuo HC, Lee KF, Tsai TH - Int J Mol Sci (2015)

A close view of selected protein spots representing differentially expressed SAM-related proteins between in control and SAM-operated rats. The proteins with expression compared with SAM group are shown. (A) Nine proteins were consistently down-regulated; (B) Six identified proteins showed a greater-than-threefold change in density. The red arrows correspond to that in Table 1. Six pairs of brain protein extracts were evaluated, and the results shown are derived from one pair of experiments; fifteen reproducible blots were performed in total.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4490420&req=5

ijms-16-11873-f003: A close view of selected protein spots representing differentially expressed SAM-related proteins between in control and SAM-operated rats. The proteins with expression compared with SAM group are shown. (A) Nine proteins were consistently down-regulated; (B) Six identified proteins showed a greater-than-threefold change in density. The red arrows correspond to that in Table 1. Six pairs of brain protein extracts were evaluated, and the results shown are derived from one pair of experiments; fifteen reproducible blots were performed in total.
Mentions: The proteomic profile of the cortex region was used to examine the events of SAM-induced brain infarction and stress responses. In this study, the protein of the cell was extracted from the control group and SAM-treated samples (n = 6 for each group). Protein samples were prepared, and more than 400 protein spots were viewed. Using silver-staining and 2D-PAGE analysis, several differences were observed with transient global ischemia in the six pairs of rats. Protein expression gel profiles (six pairs from both rat groups) were compared using ImageMaster software (Canon A640, Tokyo, Japan). The focus was then on identifying proteins found to be statistically significant at more than threefold after ischemic injury. When proteins were resolved by SDS-PAGE electrophoresis on a 12% gel, protein spots appeared to be differentially expressed more than three times greater in the SAM group than in the control group (Figure 2). In SAM rats, nine proteins were consistently down-regulated (Figure 3A), while six identified proteins showed a greater-than-threefold change in density (Figure 3B). All spots were subjected to peptide fingerprint identification using MALDI-TOF/TOF. Fifteen proteins were successfully identified by MALDI-TOF/TOF Mass Spectrometry (Table 1). Zoomed views of representative gel regions depicted several differentially expressed proteins related to oxidative stress, inflammatory response and apoptosis-related proteins. Statistical analyses were conducted after quantification of the differentially expressed proteins.

Bottom Line: It mediated the effects of SAM administration on the apoptotic and ER stress pathways.Our results demonstrate that the ischemic injury of neuronal cells increased cell cytoxicity and apoptosis, which were accompanied by sustained activation of the IRE1-alpha/TRAF2, JNK1/2, and p38 MAPK pathways.Based on these results, we also provide the molecular evidence supporting the ischemia-reperfusion-related neuronal injury.

View Article: PubMed Central - PubMed

Affiliation: Institute of Traditional Medicine, School of Medicine, National Yang-Ming University, Taipei 112, Taiwan. chenjiannhwa@yahoo.com.tw.

ABSTRACT
Ischemia-reperfusion injury resulting from arterial occlusion or hypotension in patients leads to tissue hypoxia with glucose deprivation, which causes endoplasmic reticulum (ER) stress and neuronal death. A proteomic approach was used to identify the differentially expressed proteins in the brain of rats following a global ischemic stroke. The mechanisms involved the action in apoptotic and ER stress pathways. Rats were treated with ischemia-reperfusion brain injuries by the bilateral occlusion of the common carotid artery. The cortical neuron proteins from the stroke animal model (SAM) and the control rats were separated using two-dimensional gel electrophoresis (2-DE) to purify and identify the protein profiles. Our results demonstrated that the SAM rats experienced brain cell death in the ischemic core. Fifteen proteins were expressed differentially between the SAM rats and control rats, which were assayed and validated in vivo and in vitro. Interestingly, the set of differentially expressed, down-regulated proteins included catechol O-methyltransferase (COMT) and cathepsin D (CATD), which are implicated in oxidative stress, inflammatory response and apoptosis. After an ischemic stroke, one protein spot, namely the calretinin (CALB2) protein, showed increased expression. It mediated the effects of SAM administration on the apoptotic and ER stress pathways. Our results demonstrate that the ischemic injury of neuronal cells increased cell cytoxicity and apoptosis, which were accompanied by sustained activation of the IRE1-alpha/TRAF2, JNK1/2, and p38 MAPK pathways. Proteomic analysis suggested that the differential expression of CALB2 during a global ischemic stroke could be involved in the mechanisms of ER stress-induced neuronal cell apoptosis, which occurred via IRE1-alpha/TRAF2 complex formation, with activation of JNK1/2 and p38 MAPK. Based on these results, we also provide the molecular evidence supporting the ischemia-reperfusion-related neuronal injury.

No MeSH data available.


Related in: MedlinePlus