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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

Immunohistochemical staining for indicated proteins expression in rat brains was analyzed: Saline infusion control rats (I); rats with SAM (II). The positive stained area was evaluated from three randomly selected observation fields of each brain sections. Quantitative of immunohistochemical proteins COMT, CATD and CALB2 by average integrated optical density (AIOD). Positive stained area was evaluated from three randomly selected observation fields of each brain section. Data were expressed as mean ± SD (n = 6/group). * p < 0.05, compared with control group. Magnification ×400.
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ijms-16-11873-f004: Immunohistochemical staining for indicated proteins expression in rat brains was analyzed: Saline infusion control rats (I); rats with SAM (II). The positive stained area was evaluated from three randomly selected observation fields of each brain sections. Quantitative of immunohistochemical proteins COMT, CATD and CALB2 by average integrated optical density (AIOD). Positive stained area was evaluated from three randomly selected observation fields of each brain section. Data were expressed as mean ± SD (n = 6/group). * p < 0.05, compared with control group. Magnification ×400.

Mentions: Spots 1, 5 and 14 were subsequently identified using 2D proteomic analysis, including Catechol O-methyltransferase (COMT), Cathepsin D (CATD), as well as Calretinin (CALB2), which may mediate the oxidative injury effects in the neuronal cells of rats with ischemic stroke (Table 1). In order to verify the effects of intravenous SAM administration on the relationships between neurotoxicity, inflammation and oxidative stress in the neuron cells were examined by using 2-DE proteomic analyses. These proteins were selected for further examination using immunohistochemistry assays in vivo after ischemic injury and OGD treatment in N2a cells in vitro. As shown in Figure 4, rats with ischemic impairment demonstrated reduced expression of COMT and CATD in the brain tissue samples. In particular, ischemic brain damage resulted in significant expression of CALB2. These results are consistent with the proteomic results, indicating that a proteomic differential display model is applicable when assessing SAM-induced brain infarction.


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)

Immunohistochemical staining for indicated proteins expression in rat brains was analyzed: Saline infusion control rats (I); rats with SAM (II). The positive stained area was evaluated from three randomly selected observation fields of each brain sections. Quantitative of immunohistochemical proteins COMT, CATD and CALB2 by average integrated optical density (AIOD). Positive stained area was evaluated from three randomly selected observation fields of each brain section. Data were expressed as mean ± SD (n = 6/group). * p < 0.05, compared with control group. Magnification ×400.
© Copyright Policy
Related In: Results  -  Collection

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

ijms-16-11873-f004: Immunohistochemical staining for indicated proteins expression in rat brains was analyzed: Saline infusion control rats (I); rats with SAM (II). The positive stained area was evaluated from three randomly selected observation fields of each brain sections. Quantitative of immunohistochemical proteins COMT, CATD and CALB2 by average integrated optical density (AIOD). Positive stained area was evaluated from three randomly selected observation fields of each brain section. Data were expressed as mean ± SD (n = 6/group). * p < 0.05, compared with control group. Magnification ×400.
Mentions: Spots 1, 5 and 14 were subsequently identified using 2D proteomic analysis, including Catechol O-methyltransferase (COMT), Cathepsin D (CATD), as well as Calretinin (CALB2), which may mediate the oxidative injury effects in the neuronal cells of rats with ischemic stroke (Table 1). In order to verify the effects of intravenous SAM administration on the relationships between neurotoxicity, inflammation and oxidative stress in the neuron cells were examined by using 2-DE proteomic analyses. These proteins were selected for further examination using immunohistochemistry assays in vivo after ischemic injury and OGD treatment in N2a cells in vitro. As shown in Figure 4, rats with ischemic impairment demonstrated reduced expression of COMT and CATD in the brain tissue samples. In particular, ischemic brain damage resulted in significant expression of CALB2. These results are consistent with the proteomic results, indicating that a proteomic differential display model is applicable when assessing SAM-induced brain infarction.

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