Expression and histogenesis of matrix metalloproteinase-9 and transforming growth factor-beta 1 in acute cerebral ischemia model of rats

AIM:To observe the expression and tissue localization of matrix metalloproteinase 9 (MMP-9) and transforming growth factor beta 1 (TGF-β1) in the rat acute cerebral ischemia model. METHODS:Male Wistar rats were used to establish acute cerebral ischemia model by a suturing method. The rats were divided into normal control group, sham group and ischemia 6 h, 12 h, 1 d, 2 d, 6 d and 14 d groups. The rat cerebral cortex and hippocampus of the brain were collected at different time points.The mRNA and protein levels of MMP-9 and TGF-β1 in the brain tissues were detected by real-time PCR and in situhistochemistry staining, respectively. The levels of MMP-9 and TGF-β1 in the plasma were also measured by ELISA. RESULTS:The results of real-time PCR showed that the mRNA levels of MMP-9 began to increase 6 h after acute ischemia and reached to a peak 2 d after acute ischemia. Similarly, the mRNA level of TGF-β1began to rise 12 h after acute ischemia and reached to the highest level 6 d after acute ischemia. Compared with the sham rats, the mRNA levels of MMP-9 and TGF-β1 in the rat brains that collected at ischemic time of 12 h, 1 d, 2 d, 6 d and 14 d were significantly increased. Moreover, results of in situhistochemical staining showed that the expression of MMP-9 was detected at cerebral cortex and hippocampus 1 d after acute cerebral ischemia.Further studies showed that MMP-9 dyeing of the rat cerebral cortex was most obvious 2 d after the acute cerebral ischemia. Similarly, the rat cortex and hippocampus began to express TGF-β1 2 d after acute ischemia and TGF-β1 staining at rat cerebral cortex was most obvious 6 d after the acute cerebral ischemia. In addition, ELISA showed that the increase in MMP-9 and TGF-β1 was detected in the plasma 12 h after ischemia. Compared with the sham rats, the level of these 2 factors significantly upregulated since 1 d after ischemia. CONCLUSION: The brain tissue itself contributes to the upregulation of MMP-9 and TGF-β1 post acute cerebral ischemia, which shed light on the related research in the field.     

Effects of Astragalus injection on neuronal apoptosis and expression of JNK3 in rat hippocampus after cerebral ischemia reperfusion

AIM:To investigate the effects of Astragalus injection on neuronal apoptosis and expression of c-Jun N-terminal kinase 3(JNK3) in the rat hippocampus after cerebral ischemia reperfusion. METHODS:The rat model of cerebral ischemia reperfusion was set up by a four-vessel occlusion method. The SD rats were randomly divided into 4 groups:sham operation group, cerebral ischemia reperfusion group(model group), cerebral ischemia reperfusion+Astragalus injection group(Astragalus injection group) and cerebral ischemia reperfusion+vehicle group(vehicle group). The rats in model group, Astragalus injection group and vehicle group after transient global cerebral ischemia(30 min) were then divided into 7 subgroups according to the reperfusion time of 0 h, 0.5 h, 2 h, 6 h, 24 h, 72 h and 120 h. The apoptosis of the neuron in the hippocampus was measured by the method of TUNEL staining. The expression of JNK3 at mRNA and protein levels was determined by real-time PCR and Western blotting,respectively. RESULTS:Compared with sham operation group, the number of apoptotic neurons increased in model group(P<0.05). Compared with model group, the number of apoptotic neurons decreased obviously in Astragalus injection group(P<0.05). Compared with sham operation group, the expression of JNK3 at mRNA and protein levels in the hippocampus increased obviously in model group at all time points except 120 h(P<0.05). Compared with model group, the expression of JNK3 at mRNA and protein levels in the hippocampus decreased obviously in Astragalus injection group at all time points except 120 h(P<0.05). CONCLUSION:Astragalus injection decreases neuronal apoptosis in rat hippocampus after cerebral ischemia reperfusion by inhibiting the expression of JNK3 at mRNA and protein levels.     

Effects of cerebral ischemia and postconditioning on signaling molecules PERK and GRP78 in hippocampus tissue of tree shrew during endoplasmic reticulum stress

AIM: To investigate the effects of cerebral ischemia and postconditioning on protein kinase R-like endoplasmic reticulum kinase (PERK) and glucose-regulated protein 78 (GRP78) in the hippocampus tissue of tree shrew during endoplasmic reticulum stress and the mechanism of post-conditioning protecting the brain from damage. METHODS: The focal cerebral ischemic model was duplicated by photochemical reaction in tree shrew and the postconditioning was induced by alternatively occluding and opening the carotid artery of ischemic side for 3 cycles (5 min each cycle) at 3.5 h after ischemia. The damage and ultrastructural changes of the hippocampal neurons were observed by HE staining. The expression of PERK and GRP78 at mRNA and protein levels in the hippocampal tissue at different time points after cerebral ischemia and postconditioning was determined by RT-PCR, immunohistochemistry and Western blot. RESULTS: The injuries of hippocampal neurons were aggravated with prolonged cerebral ischemia, which was most severe at 24 h after ischemia while the postconditioning alleviated these damages correspondingly. The expression of PERK at mRNA and protein levels was upregulated at 4 h, 24 h and 72 h after ischemia (P<0.05), while postconditioning downregulated the expressions of PERK at ischemia and postconditioning 4 h (IP4 h) gruop and IP24 h group (P<0.05). The expression of GRP78 at mRNA and protein levels was not changed at 4 h, 24 h and 72 h after ischemia, while postconditioning upregulated the expressions of GRP78 at IP24 h group (P<0.05). CONCLUSION: The focal thrombotic cerebral ischemia activates the endoplasmic reticulum stress in ischemic hippocampus of tree shrews, leading to the changes in mRNA and protein expression of PERK in the PERK/eIF2α signal transduction pathway. The postconditioning treatment alleviates endoplasmic reticulum stress and neuronal damages by downregulating PERK and upregulating GRP78, thereby protecting the brain from injury.     

Fluorocitrate inhibits regulatory effect of ischemic postconditioning on HIF-1α/iNOS signaling pathway after cerebral ischemia in tree shrews with brain injury

AIM: To investigate the regulatory effect of HIF-1α/iNOS signaling pathway on the neuroprotection of ischemic postconditioning (PC) in tree shrews, and to explore the mechanisms of deteriorated cerebral injury after inhibiting astrocyte (AS) metabolism. METHODS: Thrombotic cerebral ischemia was induced by photochemical reaction in tree shrews. Fluorocitrate (FC) was used to inhibit AS metabolism and the ischemic PC was established at 4 h after ischemia followed by clipped ipsilateral common carotid artery on the ischemia side for 3 times, 5 min/time. A total of 67 male tree shrews were randomly divided into 7 groups:control (n=9), ischemia (4 h and 24 h, n=9 for each group), ischemia with PC (4 h and 24 h, n=9 for each group), and FC pretreatment (4 h and 24 h, n=11 for each group). The cerebral infarction size was detected by TTC staining, and the histological changes of hippocampal neurons were observed under light microscope. The regional cerebral blood flow (rCBF) in ischemic cortex was monitored by laser Doppler brain flowmetry. The protein expression of iNOS in hippocampus was detected both by immunohistochemistry and Western blot. The production of NO detected by spectrophotometer. The level of HIF-1α in hippocampus analyzed by ELISA. RESULTS: The cerebral infarct volume was increased with prolonged duration of ischemia, and the changes of ischemia at 24 h were significant (P<0.05). The cortical rCBF was progressively decreased, and it was decreased at 4 h and 24 h after ischemia (P<0.05). The expression of HIF-1α and iNOS in hippocampus was enhanced, and the production of NO was increased significantly (P<0.05). Ischemic PC restored the cortical rCBF (P<0.05), reduced cerebral infarction volume (P<0.05), down-regulated iNOS expression and reduced NO production in the hippocampus (P<0.05). However, the cortical rCBF in FC pretreatment group was significantly lower than that in ischemic group (P<0.05), the neuronal damage was aggravated, and the infarction volume was increased after pretreatment with FC (P<0.05). CONCLUSION: Ischemic PC may reduce cerebral injury by regulating the expression of HIF-1α and iNOS. Inhibition of AS function may attenuate the protective effect mediated by ischemic PC and aggravate brain injury.     

Effects of adrenal gland on the expression of bax and bcl-2 in hippocampus after cerebral ischemia

AIM: To observe the effects of adrenal gland on the hippocampus responses to cerebral ischemia. METHODS: 36 Wistar rats were randomly divided into three groups: sham-operated control group (sham), unilateral adrenalectomy were performed in ADX and GC group, and GC group were injected with 5 mg/per rat of dexamethasone before cerebral ischemia. Fourteen days after the first operation, all animals were performed occlusion of bilateral carotid artery for 15 min, and then reperfusion. 3 rats of each group were sacrificed at 1 h, 4 h, 8 h and 24 h after reperfusion and hippocampus were dissected. The total RNA was rapidly extracted from hippocampus tissue. The expressions of c-fos, bcl-2 and bax gene were quantified with the method of semiquantitive RT-PCR. RESULTS: The expressions of c-fos and bax in three groups showed no statistical differences (P>0.05). The expression of bcl-2 in sham group was significantly higher than that in GC and ADX groups (P<0.05). However, no differences of bcl-2 expression between GC and ADX group (P>0.05) was observed. The ratio of bax to bcl-2 in sham group was significantly lower than that in GC and ADX groups (P<0.05), no significant differences of the ratio displayed between ADX and GC group. CONCLUSION: The expression of c-fos and bax in hippocampus after cerebral ischemia is not affected by adrenal gland. The excision of unilateral adrenal gland downregulates bcl-2 expression and raises the ratio of bax to bcl-2 in rat hippocampus after cerebral ischemia. Dexamethasone treatment does not alter the expression of bcl-2 in ADX and GC groups. The results indicate that the adrenal gland can counteract cell apoptosis in hippocampus tissue induced by cerebral ischemia. Adrenal steroids are not sufficient to enable the compensatory increase in bcl-2 expression in steroid-deficient animal, some other mechanism may exist.     

Effects of astragalan on neurotransmitters and expression of c-fos mRNA in hippocampus after ischemic brain injury in rats

AIM: To explore the effects of astragalan (AG) on the neurotransmitters,acetylcholine (ACh), norepinephrine (NE) and 5-hydroxytryptamine (5-HT), and the expression of c-fos mRNA in hippocampus after ischemic brain injury in rats. METHODS: Male Wistar rats (180~220 g) were randomly divided into 10 groups (n=10): sham-operated group (SOG), 3 model groups (MG 1 d, 3 d, 7 d) and 3 low- or high-dose AG treatment groups (L/H-AGTG 1 d, 3 d, 7 d), respectively. The middle cerebral artery of the rats in MG group and AGTG group were blocked by operation to induced brain injury. The cerebral blood vessels of the animals were blocked on day 1, day 2 and day 7, respectively, after the L/H-AGTG were treated with AG (5 mg/kg and 15 mg/kg, ip). The content of ACh,5-HT and NE was determined using their respective ELISA kits, and the expression of c-fos mRNA in the hippocampus homogenate was semiquantitative analyzed by RT-PCR after neurologic impairment (NIP) was scored. RESULTS: AG attenuated the injury in hippocampus by cerebral ischemia in a dose-dependent manner. The content of ACh, 5-HT and NE in L-AGTG 7 d,H-AGTG 3 d and 7 d groups was significantly higher than that in MG group, but was lower in SOG group (P<0.05 or P<0.01). The mRNA expression of c-fos in SOG group was lower than that in MG group (P<0.05 or P<0.01), indicating that reinforcement expression of c-fos mRNA by cerebral ischemia and the expression of downstream genes may be beneficial for protecting the neurons. The mRNA expression of c-fos in H-AGTG 3 d/7 d groups was higher than that in MG group (P<0.05). CONCLUSION: AG attenuates the damage of neurons and improves the functions of hippocampus under the condition of cerebral ischemia/reperfusion by increasing the content of ACh, NA and 5-HT, and the mRNA expression of c-fos in hippocampus.     

Postconditioning modulates TLR4 expression in hippocampus of tree shrews with thrombotic cerebral ischemia

AIM: To observe the effects of thrombotic cerebral ischemia and postconditioning on the expression of toll-like receptor 4 (TLR4) in hippocampus of tree shrews.METHODS: The model of thrombotic focal cerebral ischemia was established by photochemical reaction.Four hours after the onset of photochemical reaction, ischemic postconditioning was induced by 3 repeated cycles of carotid artery occlusion for 5 min and reperfusion for 5 min. The histological changes of hippocampus (by HE staining), TLR4 protein level (by Western blotting) and TLR4 mRNA expression (by semiquantitative RT-PCR) were observed.RESULTS: The extensive neuronal degeneration in hippocampus was observed from 4 h to 72 h and peaked at 24 h after cerebral ischemia, but was significantly attenuated after postconditioning. Cerebral ischemia caused a progressive increase in the expression of TLR4 protein at 4 h and 24 h (P<0.05), and decreased at 72 h (P<0.05). In contrast to ischemia groups, postconditioning decreased the expression of TLR4 protein at 4 h and 24 h (P<0.05), but an increase in the expression of TLR4 at 72 h (P<0.05) was observed. Simultaneously, the level of TLR4 mRNA in hippocampus showed the tendency of approximate variation in accordance with the protein expression.CONCLUSION: The expression of TLR4 increases by cerebral ischemia. The protection mechanisms of postconditioning may be associated with modulating TLR4 expression.     

Hypothermia postconditioning protects hippocampal CA1 neurons from focal cerebral ischemia in tree shrew

AIM: To observe the changes in vascular endothelial growth factor (VEGF) expression and the cell numbers of cellular necrosis in hippocampus CA1 area after cerebral ischemia and hypothermia postconditioning (HPC). METHODS: The focal thrombotic cerebral ischemia was induced by photochemical reaction in tree shrews. 6 h after ischemia, HPC was executed by a focal homoeothermic equipment, which reduced the brain temperature and maintained at 31-32 ℃ for 1 h. VEGF expression in hippocampus CA1 area was detected by immunohistochemistry. The numbers of death cells were counted and the ultrastructure was observed under the electron microscope. RESULTS: Compared to control group, VEGF expressions increased in neuron of hippocampus CA1 area at 24 h, and decreased at 72 h in HPC group (P<0.01). Meanwhile the numbers of necrotic cells reduced at 24 h, and increased at 72 h. In accordance with this, ultrastructure of mitochondrion and endoplasmic reticulum became deterioration at 72 h. CONCLUSION: During the early stage, VEGF expression maybe directly protects neurons from cerebral ischemia. HPC has a remarkable significance to neuroprotective function in this time, but it may aggravate neuron injury in the last stage of cerebral ischemia. The HPC may prolong the treatment time-windows in the acute phase of cerebral ischemia.     

Preliminary study on pyroptosis involved in cerebral ischemia-reperfusion injury

AIM:To investigate the changes of pyroptosis in hippocampus and cortex at different time points after cerebral ischemia-reperfusion, and to explore its mechanism from NLRP3-mediated classical pyroptosis pathway, and to analyze the role of pyroptosis in different parts of cerebral injury. METHODS:SD rats were randomly divided into sham operation group (sham group) and model group (MCAO/R group). The rats in model group was further divided into cerebral ischemia-reperfusion 6 h group (MCAO/R 6 h group), 12 h group (MCAO/R 12h group)and 24 h group (MCAO/R 24 h group). The rat model was established on rats by middle cerebral artery occlusion and reperfusion (MCAO/R) induced by modified right-side thread method. Neurologic function score, 2, 3, 5-triphenyltetrazolium chloride (TTC) staining and morphological observation were used to evaluate the degree of nervous cell injury. TUNEL and caspase-1 immunofluorescence double staining were used to detect pyroptosis. The protein expression of NLRP3, cleaved caspase-1, pro-caspase-1 and interleukin-1β (IL-1β) was determined by Western blot. RESULTS:Neurological damage occurred at different times after cerebral ischemia-reperfusion. TTC staining showed that the volume of cerebral infarction gradually increased with the prolongation of reperfusion time (P<0.05). The hippocampal CA1 area and cortical area showed typical morphological features such as loose tissue structure, interstitial edema, disordered arrangement of nerve cells, deepening of nucleus staining, nuclear fragmentation and decreased cell number. Immunofluorescence double staining showed that there was a phenomenon of pyroptosis at different time after cerebral ischemia-reperfusion. The pyroptosis of hippocampal CA1 and cortical area was most obvious at 12 h and 24 h after reperfusion (P<0.05). Western blot analysis showed that the expression of NLRP3, cleaved caspase-1, pro-caspase-1 and IL-1β in NLRP3-mediated classic pyroptosis pathway was regulated in different degrees after cerebral ischemia-reperfusion. The protein expression of NLRP3 in hippocampus was significantly increased at 12 h and 24 h after reperfusion (P<0.05), and the protein expression of NLRP3 in cortex was significantly increased at 6 h after reperfusion (P<0.05). The protein expression of pro-caspase-1 in hippocampus was significantly increased at each time points of reperfusion (P<0.05), and the protein expression of pro-caspase-1 in the cortex was significantly increased at 24 h after reperfusion (P<0.05). The protein expression of cleaved caspase-1 in the hippocampus was significantly increased at 12 h after reperfusion (P<0.05), and increased in the cortex at 24 h after reperfusion (P<0.05). The protein expression of IL-1β in the hippocampus was significantly increased at 24 h after reperfusion (P<0.05), and increased in the cortex at 6 h after reperfusion (P<0.05). CONCLUSION:Pyroptosis is involved in neuronal injury after cerebral ischemia-reperfusion. The classic pyroptosis pathway plays an important regulatory role in hippocampus and cortex, especially in hippocampus, suggesting that hippocampus is the main part of secondary nerve impairment induced by pyroptosis and inflammation after cerebral ischemia-reperfusion.     

Effect of naoluo xintong on expression of Fas,Fas L in hippocampus CA1 area and Fas mRNA in the cortex of frontal or parietal lobe after local cerebral ischemia/reperfusion in MCAO rats

AIM: To investigate the effets of naoluo xintong on the expression of Fas, FasL protein in hippocampus CA1 area and Fas mRNA in the cortex of frontal or parietal lobe after local cerebral ischemia/reperfusion in MCAO rats. METHODS: The local cerebral ischemia /reperfusion model was established by intraluminal thread occlusion of the middle cerebral arteries (MCAO), the middle cerebral arteries of rats were occluded for 2 hours and reperfused for 1, 3 and 7 days. The animals were divided into pseudo surgery group(sham group), model group, Yiqi group, Huoxue group and naoluo xintong group. Using the techniques of immuno-histochemical staining and in situ hybridization, the expression of Fas and FasL was observed in hippocampus CA1 area, the expression of Fas mRNA was also observed in the cortex of frontal and parietal lobe. RESULTS: A value of Fas and FasL protein expression or A value and positive unit of Fas mRNA expression in control group were higher than those in sham in hippocampus CA1 area, the cortex of frontal or parietal lobe after local cerebral ischemia/reperfusion in MCAO rats (P<0.01). A value and/or positive unit of their expression in naoluo xintong group were lower than those in control group (P<0.05 or P<0.01). A value and/or positive unit of their expression in Yiqi and Huoxue groups were higher than those in naoluo xintong group for 3 and/or 7 days (P<0.05 or P<0.01). CONCLUSION: naoluo xintong could resist neuron apoptosis, alleviate pathologic injury after local cerebral ischemia/reperfusion in MCAO rats by inhibiting the expression of Fas, FasL protein and Fas mRNA.     

Protective effect of allitridi on hippocampus of rats with cerebral ischemia-reperfusion and P53 expression

AIM: To observe the protective effect of allitridi on hippocampal neuron of rats with cerebral ischemia-reperfusion (I/R) injury and to investigate its effects on P53 expression in hippocampus.METHODS: The global cerebral ischemia-reperfusion models were established by 4-vessel occlusion. Allitridi at doses of 10, 20 or 30 mg/kg was injected through rat’s tail vein, half dose at 30 min before brain ischemia and another half dose at 10 min after reperfusion were injected, respectively. The hippocampus of rat was removed 24 h after reperfusion. Toluidine blue staining was applied to estimate morphologic changes. Flow cytometry was used to evaluate neuronal apoptosis rate of hippocampus. Immunohistochemistry was used to observe the expression of P53 protein.RESULTS: Compared with sham group, survival neuronal density in I/R group was significantly depressed. The rate of neuronal apoptosis and the expression of P53 protein were significantly increased. Allitridi significantly increased the number of survival neurons in hippocampus compared to I/R group. Meanwhile, allitridi remarkably inhibited the rate of neuronal apoptosis and the expression of P53 protein.CONCLUSION: Allitridi has protective role against brain ischemia reperfusion injury. The mechanism may be involved in blocking P53 protein expression in hippocampus of rats with ischemia-reperfusion.     

Effects of flavonoids isolated from Scutellaria stem and leaf on expression of NMDAR and VEGF in chronic cerebral ischemia rats

AIM: To study the effects of flavonoids isolated from Scutellaria stem and leaf (SSF) on the expression of N-methyl-D-aspartate receptor (NMDAR) and vascular endothelial growth factor (VEGF) in chronic cerebral ischemia rats. METHODS: The model of chronic cerebral ischemia was established by bilateral carotid artery occlusion for 2 months in female SD rats. The effects of SSF on mRNA expression of NMDAR in hippocampus and VEGF in cerebral cortex were evaluated by the method of RT-PCR. RESULTS: Compared with the sham group, the expression of NMDAR1, NMDAR2A and NMDAR2B in hippocampus and VEGF in cerebral cortex were significantly increased (P<0.01). However, the cerebral ischemia rats daily and orally administered with SSF at doses of 17.5 mg·kg^-1·d^-1, 35 mg·kg^-1·d^-1 and 70 mg·kg^-1·d^-1 for 38 days appeared that the mRNA expression of NMDAR1, NMDAR2A and NMDAR2B in hippocampus was obviously reduced (P<0.05), and the mRNA content of VEGF in the cortex (P<0.05) was increased. CONCLUSION: SSF decreases the expression of NMDAR in hippocampus, increases the expression of VEGF in cerebral cortex of cerebral ischemia rats, suggesting that the neuroprotective effect of SSF may be exerted by influencing the production of NMDAR and VEGF in the brain.     

Expression of CRF and PKC proteins in hippocampus of rats with cerebral ischemia and reperfusion

AIM: To observe the expression of CRF and PKC in rats with cerebral ischemia.METHODS: Using immunohistochemistry technique we measured the expression quantitatively of CRF and PKC proteins in the hippocampus in rats induced by MCAO at 2 h,6 h and 24 h after reperfusion,contrast to CRF antagonist.RESULTS: (1) CRF: there were lots of positive and deeper dyeing neurons in hippocampus in model group and normal saline group rats,while there were a few positive and lighter dyeing neurons in sham group and CRF antagonist group.The positive expression areas of CRF protein in hippocampus in model group and normal saline group were significantly bigger than those in sham group and CRF-antagonist group(P<0.01),respectively.(2) PKC：there were a great number of denser positive granules in hippocampus in model group and normal saline group rats,while there were a few of scattered positive granules in sham group and CRF antagonist group.The positive expression areas of CRF protein in hippocampus in model group and normal saline group were significantly bigger than that in sham group and CRF-antagonist group (P<0.01),respectively.CONCLUSION: The high expression of CRF and PKC induced by cerebral ischemia may be one important factors that resulted in the delayed neuronal death in hippocampus.The CRF protein activated PKC expression,indicating an important pathology mechanism of nerve tissue damage induced by CRF.     

Acute cerebral ischemia activates EphB2/ephrin-B1/NMDA receptor signaling pathway to promote hippocampal neurogenesis in mice

AIM To investigate the effect of acute cerebral ischemia on hippocampal neurogenesis in mice and its possible mechanism involving EphB2/ephrin-B1/NMDA receptor signaling pathway. METHODS C57BL/6 mice (n=52) were randomly divided into sham group and acute cerebral ischemia group (model group). The model of acute cerebral ischemia in mice was established by bilateral common carotid artery occlusion. The pathological changes of the hippocampal CA1 region in mice were observed by HE staining. The learning and memory functions of the mice were assessed by Morris water maze. The BrdU positive cells and doublecortin (DCX) protein expression were observed by immunofluorescence staining for detecting hippocampal neurogenesis. The mRNA and protein expression levels of EphB2, ephrin-B1, reelin, microtubule-associated protein-2 (MAP-2) and NMDA receptor subunits NR2A and NR2B in the hippocampus were determined by RT-qPCR and Western blot. RESULTS The neuronal damage in the hippocampal CA1 region was significant (P<0.01), and the learning and memory functions were significantly decreased in the cerebral ischemia mice(P<0.01), suggesting that the cerebral ischemia model was successfully established. The BrdU positive cells and DCX protein expression were increased significantly (P<0.01), indicating that neurogenesis occurred in the hippocampus after cerebral ischemia. At the same time, the mRNA and protein expression levels of EphB2, ephrin-B1, reelin, MAP-2, NR2A and NR2B in the hippocampus were also significantly up-regulated (P<0.05). CONCLUSION Acute cerebral ischemia promotes the proliferation of hippocampal neural stem cells and endogenous neurogenesis, which may be related to the activation of EphB2/ephrin-B1/NMDA receptor signaling pathway.     

Changes of cortisol content in plasma and hippocampus after focal cerebral ischemia-reperfusion in rats

AIM: To investigate the relationship between glucocorticoid (Gc) and injury of hippocampus neurons and the effect of Gc on dementia episode after cerebral ischemia-reperfusion. METHODS: The rat model of middle cerebral artery occlusion (MACO) was established. Cortisol contents in hippocampus and plasma of the model rats were examined by means of the radioimmunoassay at 2 h, 6 h, 12 h, 24 h after reperfusion. RESULTS: The levels of cortisol content in model group were significantly higher than those in sham group and normal group both in hippocampus and plasma. The highest cortisol content was observed at 6 hours after reperfusion. HE staining showed that the impairment of hippocampus neurons was aggravated progressively with reperfusion interval elongating. CONCLUSION: The increased cortisol in hippocampus and plasma, after 2 h cerebral ischemia and 24 h reperfusion, could aggravate the injury of hippocampus neurons and lead to dementia post stroke.     

Effects of hyperglycemia and cerebral ischemia on VEGF expression in different subfield of cerebral cortex in tree shrews

AIM: To observe the changes of VEGF expression in different subfield of brain in tree shrews during hyperglycemia and focal cerebral ischemia, in order to explore the relationship between cerebral ischemia, hyperglycemia and VEGF. METHODS: High blood glucose in tree shrews was induced by intraperitoneal injection of streptozotoctin. Focal cortical thrombotic cerebral ischemia was induced by photochemical method in tree shrews. At 4 h, 24 h and 72 h after cerebral ischemia, the histopathological changes and hippocampal neuronal density were examined. VEGF expressions in the ischemic core, penumbra and contralateral cerebral cortex were detected by immunohistochemistry technique at different times after cerebral ischemia. RESULTS: The results of histopathological study showed that there was infarction zone in the exposured cerebral cortex at 4 h after photochemical reaction, and the damage was most severe at 24 h, subsequently accompanied with the glia multiplication and rehab reaction at 72 h. The animals in hyperglycemic ischemic group suffered from greater neurological lesion than the normoglycemic stroke animals, especially at 24 h (P<0.01) and 72 h (P<0.05) after cerebral ischemia. Immunohistochemical analyses of VEGF expression revealed that it started to increase at 4 h after brain ischemia in the penumbra, reached a peak at 24 h, and weakened at 72 h. The stimulated VEGF production was also observed in hyperglycemic only group. When hyperglycemia and brain ischemia were combined, the VEGF expression was higher than that in hyperglycemic only group (P<0.05). Compared to normoglycemic ischemic group, no additivity of the effects of hyperglycemia combined with brain ischemia was observed. CONCLUSION: (1) The model of experimental hyperglycemia and cerebral ischemia is replicated successfully by applying the method combined in vivo injection of streptozotocin in the lower primate tree shrew with thrombotic focal cerebral ischemia. (2) This study shows that hyperglycemia aggravates the focal cerebral ischemia damage. (3) Cerebral ischemia and hyperglycemia both can independently up-regulate VEGF expression, but there is no additional increase in VEGF expression when hyperglycemia combined with brain ischemia is applied.     

Effect of edaravone on expression of Aβ and AQP4 and activity of MMP2 and MMP9 in acute cerebral ischemia/reperfusion rats

AIM: To investigate the effect of edaravone on acute cerebral ischemia/reperfusion rats. METHODS: SD rats were randomly divided into sham operation group (saline), model group (modeling given saline), low dose group (edaravone at 6 mg/kg) and high dose group (edaravone at 10 mg/kg). The rat model was established by Zea Longa suture method. The nerve function scores were evaluated after operation, and the infarct volume was measured by TTC assay. The mRNA expression of aquaporin 4 (AQP4) and amyloid β-protein (Aβ) in the brain tissue was detected by RT-qPCR. The protein levels of APQ4 and Aβ were determined by Western blot. The activity of matrix metalloproteinase 2 (MMP2) and matrix metalloproteinase 9(MMP9) was detected by gelatin zymography. RESULTS: Compared with model group, edaravone administration markedly alleviated neurological deficits, histological damages and brain edema. The mRNA and protein levels of AQP4 and Aβ, and the activity of MMP2 and MMP9 were downregulated (P<0.05). Furthermore, the improvements in high dose group were significantly more effective than those in low dose group. CONCLUSION: Edaravone significantly reduces neurological deficits and brain edema in the rats with acute ischemic stroke, and the mechanisms may be related to the downregulation of AQP4 and Aβ, and the activation of MMP2 and MMP9.     

Effects of Le Er Mai on apoptosis of hippocampus neuronal cells in anaphase of cerebral ischemic reperfusion injury in rats

AIM: To investigate the effects and mechanism of Le Er Mai (LEM) on the apoptosis of hippocampus neuronal cells in the anaphase of cerebral ischemic reperfusion injury in rats.METHODS: A rat model of middle cerebral artery occlusion reperfusion (MCAO) was produced with the intraluminal filament. During reperfusion for 30 d after 2 h of ischemia, the TUNEL staining methods were used to detect apoptosis of hippocampus neuronal cells, and immunohistochemical technique were employed to examine the protein expression of Fas, Bax, caspase-3 and caspase-9 in the hippocampial. The gene expressions of fas, bax, caspase-3 and caspase-9 in hippocampial were examined by RT-PCR. RESULTS: After 2 h ischemia and 30 d reperfusion, compared with sham-operated group, TUNEL-positive staining cells and expression levels of Fas, Bax as well as caspase-3 and caspase-9 obviously increased, and the mRNA expressions of fas, bax, caspase-3 and caspase-9 in hippocampial markedly up-regulated in model group. Compared with model group, LEM at dose of 2.00 g/kg or 0.87 g/kg, and flunarizinum significantly reduced apoptosis and decreased the protein expressions of Fas, Bax, caspase-3 and caspase-9 in hippocampial, and down-regulated the mRNA expressions of fas, bax, caspase-3 and caspase-9 (P<0.05), those action of LEM in 0.87 g/kg dosage group was lower than those in 2.00 g/kg dosage group.CONCLUSION: LEM obviously lower the injury of hippocampial in the anaphase of cerebral ischemia reperfusion through inhibiting the apoptosis of hippocampus neuronal cells. The mechanism of LEM may be related to regulate the expression of signal transduction pathway correlated gene of apoptosis in neuronal cells.