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 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.     

Analgesic effect of angiotensin angiotensin Ⅱ type 2 receptor antagonist EMA401 on neuropathic pain in rats and its mechanism

AIM: To explore whether angiotensin Ⅱ type 2 receptor antagonist EMA401 decreases neuropathic pain and the expression of growth-associated protein-43 (GAP-43), protein kinase C (PKC) and calmodulin (CaM) in dorsal root ganglia (DRG) during chronic constriction injury (CCI) in rats. METHODS: SD rats were used to establish CCI model and randomly divided into 4 groups. The rats in model group were given equal volume of normal saline by intragastric administration. The rats in low dose (LD) group were given 5 mg/kg EMA401 by intragastric administration. The rats in middle dose (MD) group were given 10 mg/kg EMA401 by intragastric administration. The rats high dose (HD) group were given 20 mg/kg EMA401 by intragastric administration. The rats in sham operation group received equal volume of normal saline by intragastric administration. Thermal withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) were measured before operation and 7 d, 14 d and 28 d after CCI. After behavioral test, DRG of lumbar spinal was obtained from each group, and was used to determine Ca²⁺ concentration by o-cresolphthalein complexone microplating method, and the expression of GAP-43, PKC and CaM at mRNA and protein levels by Western blotting and RT-PCR. RESULTS: Compared with model group, EMA401 significantly increased the TWL and MWT (P<0.05). Meanwhile, EMA401 significantly reduced Ca²⁺ concentration and the expression of GAP-43, PKC and CaM at mRNA and protein levels in the DRG (P<0.05). CONCLUSION: EMA401 may attenuate neuropathic pain of CCI by inhibiting Ca²⁺ concentration and the expression of GAP-43, PKC and CaM.     

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.     

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.     

Effect of concentrated decoction of Chinese herbal compound Buyanghuanwutang on cAMP-PKA-CREB signaling pathway in hippocampus of rats with vascular dementia

AIM: To evaluate the role of concentrated decoction of Chinese herbal compound Buyanghuanwutang (BYHWT) in cyclic adenosine monophosphate(cAMP)-protein kinase A(PKA)-cAMP response element-binding protein(CREB) signaling pathway in hippocampus of rats with vascular dementia (VD). METHODS: The rats were randomly divided into sham operation group (sham-operated rats treated with normal saline), VD model group (VD rats treated with normal saline), BYHWT treatment group (VD rats treated with BYHWT) and nimodipment treating group (VD rats treated with nimodipine). The rat model of VD was build by the method of four-vessel occlusion. The rats in all 4 groups were administered with the corresponding reagents for successive 30 days. The content of cAMP was measured by radioimmunoassay. The expression of PKA catalytic subunit (PKAc) was observed by Western blotting. The changes of DNA-binding activity of CREB in rat hippocampus were detected by electrophoretic mobility shift assay. RESULTS: The content of cAMP, the expression of PKAc and the DNA-bingding activity of CREB in the hippocampus of VD rats were lower than those in the hippocampus of sham-operated rats (P<0.01). The above indexes in both nimodipine treatment group and BYHWT treatment group were definitely higher than those in VD model group (P<0.01). CONCLUSION: BYHWT increases the content of cAMP, the expression of PKAc and the DNA-binding activity of CREB in VD rat hippocampus, thus strengthening the cAMP-PKA-CREB signaling pathway.     

Astragalus injection inhibits expression of Apaf-1 in rat hippocampus after cerebral ischemia and reperfusion

AIM: To investigate the effect of Astragalus injection on the expression of apoptotic protease-activating factor 1 (Apaf-1) in the hippocampus of global cerebral ische-mia-reperfusion rats. METHODS: Male SD rats were randomly divided into 4 groups with 30 each: sham operation group, cerebral ischemia-reperfusion group, cerebral ischemia-reperfusion+Astragalus injection group, and cerebral ischemia-reperfusion+vehicle group. The global cerebral ischemia-reperfusion model of the rats was established by 4-vessel occlusion. The rats in cerebral ischemia-reperfusion group, cerebral ischemia-reperfusion+Astragalus injection group and cerebral ischemia-reperfusion+vehicle group were further divided into 7 subsets, according to the reperfusion time of 0 h, 0.5 h, 2 h, 6 h, 24 h, 72 h and 120 h. After reperfusion, the brains were removed at the corresponding time points. The protein expression of Apaf-1 in hippocampal neurons was detected by immunohistochemistry and Western blotting. The mRNA expression of Apaf-1 was observed by RT-PCR. RESULTS: Compared with sham operation group, the expression of Apaf-1 at mRNA and protein levels at all time points except 0 h and 120 h increased obviously in cerebral ischemia-reperfusion group (P<0.05). Compared with cerebral ischemia-reperfusion group, the expression of Apaf-1 at mRNA and protein levels at all time points except 0 h and 120 h decreased obviously in cerebral ischemia-reperfusion+Astragalus injection group (P<0.05). However, those in cerebral ischemia-reperfusion+vehicle group had no obvious change (P>0.05). CONCLUSION: Astragalus injection inhibits the expression of Apaf-1 at mRNA and protein levels in hippocampus of global cerebral ischemia-reperfusion rats, thus inhibiting the apoptosis of hippocampal neurons.     

Pretreatment of hypertonic saline attenuates the hepatic ischemia reperfusion injury induced by neutrophils

AIM: To explore the effect of the pretreatment of hypertonic saline (HTS) in hepatic ischemia reperfusion (I/R) injury.METHODS: The rats were divided into sham group (sham group), ischemia reperfusion group (IR group) and pretreatment of hypertonic saline group (HTS group). Partial hepatic ischemia reperfusion model was used. The rats were sacrificed at the time of 1 h, 3 h, 6 h, 12 h and 24 h after reperfusion in each group, respectively. Blood samples were obtained to examine ALT. The expression of the CD11b/CD18 (Mac-1) on the neutrophils was analyzed by flow cytometry. RT-PCR and Western blotting were used to examine the expression of intercellular adhesion molecule-1 (ICAM-1) in livers and chromatometry was performed to detect the activity of myeloperoxidase (MPO) in livers. The morphology of hepatocytes and the structure of sinusoid were observed by histological examinations. RESULTS: ① HTS pretreatment decreased the level of ALT at the time points of 3 h, 6 h and 12 h after reperfusion (P<0.05). ② Mac-1 expression in HTS group was lower at 6 h and 12 h after reperfusion compared with IR group (P<0.05). ③ MPO activity in HTS group was lower at 6 h, 12 h and 24 h compared with IR group (P<0.05). ④ RT-PCR and Western blotting analysis indicated that the pretreatment of HTS inhibited the expression of ICAM-1 in livers after reperfusion. ⑤ Moderate hepatocyte swelling and few neutrophil infiltration were observed in HTS group.CONCLUSION: Pretreatment with HTS has the effect on hepatic ischemia reperfusion injury by inhibiting the expression of Mac-1 on circulating neutrophils and the expression of ICAM-1 in the liver.     

Effects of Xingnaojing injection on expression of ZO-1 in blood-brain barrier after global ischemia-reperfusion

AIM: To investigate the effect of Xingnaojing (XNJ) injection on the permeability of blood-brain barrier (BBB) and zonula occludens-1 (ZO-1) protein expression after global ischemia-reperfusion in rats. METHODS: Improved Pulsinelli four-vessel occlusion method was adopted to establish the global ischemia-reperfusion model in the rats. Male Wistar rats were randomly divided into sham group, model group, solvent group and XNJ group. The observations were conducted at the time points of 24 h, 48 h and 72 h after ischemia reperfusion. The water content of the brain tissues was determined by dry-wet weight method, while the Evans blue (EB) content of brain tissue was detected by spectrophotometry. The protein levels of ZO-1 in the cerebral cortex were analyzed by Western blot. RESULTS: The water contents in the brain tissues in model group, solvent group and XNJ group were significantly higher than those in sham group (P<0.05) 24 h after ischemia reperfusion. However, the brain water contents in model group and solvent group were significantly higher than those in XNJ group and sham group (P<0.05) 48 h and 72 h after ischemia reperfusion. The EB contents in the brain tissues in model group, solvent group and XNJ group were entirely higher than that in sham group 24 h after ischemia reperfusion (P<0.05). The EB contents in sham group and XNJ group were significantly lower than those in model group and solvent group 48 h and 72 h after ischemia reperfusion (P<0.05). The protein expression of ZO-1 in the rat cerebral cortex in model group, solvent group and XNJ group was significantly lower than that in sham group 24 h after ischemia-reperfusion (P<0.05). Similarly, 48 h and 72 h after ischemia reperfusion, ZO-1 protein level in the cortex in sham group and XNJ group was significantly higher than that in model group and solvent group (P<0.05). CONCLUSION: At 48 h and 72 h after global ischemia-reperfusion, Xingnaojing injection play a protective role in blood-brain barrier and this role may be associated with the increase in ZO-1 protein expression by Xingnaojing injection.     

Effects of edaravone on expression of caspase-3 and caspase-12 in juvenile rat hippocampus after status convulsion

AIM: To investigate the effect of edaravone (ED) on the expression of caspase-3 and caspase-12 in the juvenile rat hippocampus after status convulsion (SC).METHODS: Juvenile male Sprague-Dawley rats were randomly divided into normal saline (NS) control group, SC group and ED treatment group. The rats in each group were further divided into 5 subgroups according to different time points. The rats in SC group were kindled into epilepsy by lithium-pilocarpine chemical method. The protein expression of caspase-3 and caspase-12 was determined by immunohistochemistry methods. The mRNA expression of caspase-3 and caspase-12 was detected by RT-PCR. RESULTS: (1) The IA value of caspase-3 positive cells in 24~72 h SC group increased compared with NS group. With ED intervention, the IA value of caspase-3 positive cells decreased as compared with 48~72 h SC group. The results of RT-PCR showed that the mRNA expression of caspase-3 was similar to the changes of protein. (2) The results of immunohistochemistry showed that the IA value of caspase-12 positive cells in 12~72 h SC group increased compared with NS group. With ED intervention, the IA value of caspase-12 positive cells decreased as compared with 24~72 h SC group. The results of RT-PCR showed that the mRNA expression of caspase-12 was similar to the changes of protein. (3) In ED group, Ⅴ grade convulsion was lower than that in SC group, and the latent period of seizures in ED group was significantly longer than that in SC group. CONCLUSION: Edaravone inhibits the expression of caspase-3 and caspase-12 in pilocarpine-induced seizures in rat hippocampus, suggesting that edaravone has protective effect against the damage caused by status convulsion.     

Expression and significance of fgl2 in a model of rat coronary microthrombosis in situ

AIM: To explore the expression and significance of fibrinogen-like protein 2 prothrombinase/fibroleukin (fgl2) in a model of rat coronary microthrombosis. METHODS: Seventy-eight Sprague-Dawley male rats were randomly assigned to model group (n=36), sham operation group (n=36) for different time (1 h, 1 d, 1week, 2 weeks, 3 weeks, 4 weeks)(n=6, respectively), and normal control group (n=6). After closing aorta, sodium laurate was injected into aortic root of the animals in model group. The animals in sham operation group were injected with normal saline for control. Microthromb were detected with HE staining and modified martius-sarlet-blue stains (MMSB). von Willebrand factor (vWF) was measured with ELISA. fgl2 mRNA and protein expressions were detected by RT-PCR and immunohistochemistry staining, respectively. RESULTS: Microthromb were observed in rat coronary microvasculature. vWF was increased in plasma. fgl2 was induced in model group compared to sham operation group (P<0.05), no significant changes were observed in sham operation group and normal control group (P>0.05). CONCLUSION: The functional disorder of endothelial cells induces the expression of fgl2 in the coronary microvasculature. fgl2 prompts the thrombosis with platelet and fibrin.     

Preconditioning with 3-nitropropionic acid induces rat cerebral ischemic tolerance and increases expression of glucose transporter 1 and glucose transporter 3

AIM: To explore the mechanism of 3-nitropropionic acid (3-NPA) preconditioning that induces cerebral ischemic tolerance in rats by affecting the expression of brain-type glucose transporters (GLUT1 and GLUT3) at mRNA and protein levels in cerebral tissues.METHODS: The male SD rats were used in the experiments and divided randomly into sham operation group (sham group, n=4), control group of 3-NPA preconditioning (3-NPA group, n=4), cerebral ischemia group (M group, n=16) and 3-NPA preconditioning group (IPC group, n=16). M group and IPC group were further divided into 4 subgroups according to the different reperfusion time(4 h, 12 h, 24 h and 48 h). All rats were killed at the corresponding time points. The cerebral tissues in the ischemic side (left) and coronal intermediate 1/3 of cortex were collected. The protein levels and mRNA expression of GLUT1 and GLUT3 were determined by Western blotting and RT-PCR. RESULTS: Compared with M group, the ischemic reperfusion and 3-NPA preconditioning induced the upregulation of GLUT1 and GLUT3 at protein levels with significant differences (F=5.848, P<0.05 and F=6.295, P<0.05, respectively), especially after ischemia-reperfusion for 48 h. The mRNA expression of GLUT1 in IPC group began to increase at 4 h, peaked at 48 h after reperfusion, with significant difference as compared to M group at the corresponding reperfusion time points in each group or sham group. In contrast, the mRNA expression of GLUT3 in IPC group increased at 24 h, and was the highest at 48 h as compared to cerebral ischemia group at the corresponding reperfusion time points or sham group.CONCLUSION: 3-NPA preconditioning increases the expression of GLUT1 and GLUT3 at protein and mRNA levels to maintain the energy supply in brain tissues, indicating a cerebral protective mechanism.     

Effects of different duration of sleep deprivation on neurogranin,PKC and CaMKⅡ mRNA expression in hippocampus and forebrain of rats

AIM: To study the effects of different duration of sleep deprivation (SD) on neurogranin (Ng), protein kinase C (PKC) and Ca2+-calmodulin dependent protein kinase II (CaMK II) mRNA expressions in hippocampus and forebrain of rats. METHODS: Male Wistar rats were divided into three groups: 24 h, 48 h and 72 h experimental groups. Each group was divided into sleep deprivation group (SD group) and control group (C group). SD model was established by sleep deprivation box. One step methods by Trizol was used to extract hippocampus and forebrain neuronal total RNA. The changes of Ng, PKC and CaMK II mRNA expressions were detected by SYBRA green I RT-PCR. RESULTS: (1) The expression of Ng mRNA in hippocampus of SD group was significantly lower than that in C group after 24 h, 48 h and 72 h SD (P<0.05). The expressions of PKC and CaMK II mRNA in SD group were significantly lower than those in C group after 48 h and 72 h SD (P<0.05). (2) The expressions of Ng, PKC and CaMKⅡ mRNA in forebrain of SD group were lower than those in C group after 24 h and 48 h SD，and significantly lower than those in C group only after 72 h SD (P<0.05). CONCLUSION: The expressions of Ng, PKC and CaMKⅡ mRNA are down regulated by SD, which might be relevant to the impairment of cognitive function in rats.     

Effects of maternal limb ischemic preconditioning on structural and functional changes of mitochondria in fetal hippocampal neurons induced by intrauterine distress-reoxygenation in rats

AIM: To investigate the effects of maternal limb ischemic preconditioning (LIP) on the mitochondrial structures and functions of the hippocampal neurons induced by reoxygenation in the intrauterine distress fetal rats. METHODS: Pregnant rats (n=40) were randomly divided into 4 groups: sham (S) group, LIP group, fetal distress (FD) group and LIP+FD group. Intrauterine ischemia model was established through the experimental design. The ultrastructure of the mitochondria in CA1 area of the hippocampus was observed. The mitochondrial membrane potential and reactive oxygen species (ROS) were measured. The content of ATP and MDA in the hippocampus tissue was detected. The activity of Mn-SOD was observed. RESULTS: Compared with sham group, the ultrastructure of mitochondria in CA1 area of the hippocampus was damaged in FD group and LIP+FD group. The mitochondrial membrane potential, the content of ATP and the activity of Mn-SOD were decreased. However, the content of ROS and MDA was increased. Compared with FD group, the ultrastructure of mitochondria in CA1 area of the hippocampus was intact in LIP+FD group. Furthermore, the reduced mitochondrial membrane potential and ATP content were inhibited. The activity of Mn-SOD was increased, but the content of ROS and MDA was decreased in LIP+FD group. CONCLUSION: Limb ischemia preconditioning inhibits the damage the mitochondria of fetal hippocampal neurons induced by reoxygenation in the intrauterine distress fetal rats.     

Effects of bFGF on neuronal apoptosis and fractalkine expression in cerebral ischemia/reperfusion rats

AIM: To study the effects of basic fibroblast growth factor (bFGF) on neuronal apoptosis and fractalkine expression in ischemic penumbra after cerebral ischemia/reperfusion in rats.METHODS: Thirty-six rats were randomly divided into 3 groups: sham operation group, ischemia/reperfusion group and bFGF group. The model of middle cerebral artery occlusion was established by the method of intraluminal filament blockage. The middle cerebral arteries were blocked for 1 h and then reperfused for 24 h. Neurological performances of all rats were scored with Bederson's standard. The brain tissues of the rats were stained and the average infarct volume was calculated. TUNEL method was used to determine the number of apoptotic neurons, and the expression of fractalkine was detected by the method of immunohistochemistry.RESULTS: The score of neurological performances in bFGF group was 2.23±0.59, lower than that in ischemia/reperfusion group (3.18±0.65). The number of apoptotic neurons in bFGF group (13.22±1.35) was lower than that in ischemia/reperfusion group (17.28±1.01, P<0.05), which was the lowest in sham operation group (0.91±0.65). Compared with sham operation group, the expression of fractalkine in ischemia/reperfusion group was decreased. The expression of fractalkine in bFGF group was mainly higher than that in ischemia/reperfusion group (P<0.05).CONCLUSION: Up-regulation of fractalkine may be one of the molecular mechanisms of bFGF to protect neurons against ischemia/reperfusion injury.     

Effect of IL-13 on expression of IL-1β in acute renal ischemia/ reperfusion injury in rats

AIM:To investigate the effects of IL-13 on expression of IL-1β in acute renal ischemia/reperfusion injury.METHODS:Fifty-seven male Wistar rats were randomly divided into 8 group: normal group, sham operation group, ischemia group, ischemia/reperfusion injury group(I/R), normal saline(NS)-treated group 1(C-1), NS-treated group 2(C-2), IL-13-treated group1(T-1)and IL-13-treated group 2(T-2).Rats were subjected to 45 min bilateral renal ischemia followed by reperfusion. rmIL-13 (1.5 μg/50 g body weight )was injected into the renal arteries through the abdominal aorta before ischemia(T-1) or immediately afterischemia(T-2).The serum level of IL-1β and the renal expression of IL-1β were determined in each group at 24 h post-ischemia. In addition, BUN, Cr and renal histology were also measured.RESULTS:(1)The serum level of IL-1β, gene expression and protein production of IL-1β in kidney decreased markedly in IL-13-treated groups.(2)Renal function and histology were significantly improved in IL-13-treated groups, renal injury scores decreased significantly.(3)A positive correlation were found between the serum level of IL-1β and BUN, SCr(r=0.708, P＜0.01;r=0.770, P＜0.01).CONCLUSION:These data suggest that IL-13 inhibit the expression of IL-1βand improve func-tion and histology of kidney in acute renal ischemia/reperfusion injury.     

Effects of baicalin on glial fibrillary acidic protein and nuclear factor-κB expression in juvenile rat hippocampus after status convulsion

AIM: To study the effects of baicalin (BC) on glial fibrillary acidic protein (GFAP) and nuclear factor-κB (NF-κB) expression and neuronal apoptosis in juvenile rat hippocampus after status convulsion (SC). METHODS: One hundred and ninety five juvenile male Sprague-Dawley rats were randomly divided into 3 groups: normal saline pretreatment group (NS group), SC group and SC with BC pretreatment group (BC group). Each of these 3 groups would be subdivided into 5 subgroups sacrificed at 4 h, 12 h, 24 h, 48 h and 72 h after SC. The rat SC model was prepared by lithium-pilocarpine chemical method. The protein expression of GFAP and NF-κB was detected by the method of immunohistochemistry. The mRNA expression of GFAP was detected by RT-PCR. The neuronal apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL). RESULTS: Compared with NS group, the GFAP positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of GFAP was significantly reduced in BC group (P<0.05). Compared with NS group, the NF-κB positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of NF-κB was significantly reduced in BC group. RT-PCR showed that the expression trend of GFAP mRNA was similar to that of the protein. Compared with NS group, the TUNEL positive cells in the hippocampus CA1 area in SC group increased significantly 12 h after SC (P<0.01), and reached a peak at 48 h. After the intervention with BC, the TUNEL positive cells decreased significantly between 12~48 h after SC (P<0.05 or P<0.01), but the number of TUNEL positive cells remained significantly greater than that in NS group (P<0.05). CONCLUSION: The expression of GFAP and NF-κB in the hippocampus increased after SC in rats. Baicalin decreases the expression of GFAP and NF-κB in hippocampus of rats with pilocarpine-induced seizures, and reduces the number of neuronal apoptosis, suggesting that baicalin may protect against the brain damage caused by status convulsion.     

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.