Objective:To investigate whether astragaloside (AST) IV can improve spatial learning and memory abilities by alleviating oxidative stress damage to the frontal cortex and hippocampus in vascular dementia (VD) rats induced by chronic cerebral ischemia.Methods:Totally, 72 adult male Wistar rats were randomly assigned to four groups: sham-operated group ( n=12), model group ( n=20), AST-IV 10 mg group ( n=20), and AST-IV 20 mg group ( n=20); chronic cerebral ischemia-induced VD models in the later three groups were established by permanent bilateral common carotid artery occlusion (BCCAO); 3 h after BCCAO, these rats were administered with saline, 10 mg/kg AST-IV, or 20 mg/kg AST-IV once daily for a consecutive 90 d. Ninety-four d after modeling, spatial learning and memory abilities were assessed by Morris water maze; the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT), and malondialdehyde (MDA) levels were measured by enzyme linked immunosorbent assay (ELISA). The levels of lipid peroxidation and oxidative DNA damage were assessed by immunohistochemical staining for 4-hydroxynonenal (4-HNE) and 8-hydroxy20-deoxyguanosine (8-OhdG), respectively. Results:(1) On the 3 rd, 4 th and 5 th d of place navigation test, the escape latency in rats of the model group was significantly longer than that in the sham-operated group, and that in the AST-IV 20 mg group was significantly shorter than that in the model group ( P<0.05); spatial probe test showed that the time percentage of rats spending in platform region in the model group (20.3%±1.7%) was significantly smaller than that in the sham-oprated group (48.2%±3.6%), and that in the AST-IV 20 mg group (39.7%±3.2%) was significantly larger than that in the model group ( P<0.05). (2) As compared with those in the sham-operated group, the SOD, GSH-Px and CAT activities were statistically decreased while MDA level was significantly increased in the frontal cortex and hippocampal CA1 area of rats in the model group ( P<0.05); as compared with those in the model group, the SOD, GSH-Px and CAT activities were statistically increased while MDA level was significantly decreased in the frontal cortex and hippocampal CA1 area of rats in the AST-IV 20 mg group ( P<0.05). (3) As compared with those in the model group, the numbers of 4-HNE and 8-oHdG positive cells in the frontal cortex and hippocampal CA1 area of rats in the AST-IV 20 mg group were significantly smaller ( P<0.05). Conclusion:Intraperitoneal injection of high dose AST-IV can ameliorate oxidative damage in the frontal cortex and hippocampal CA1 area in chronic cerebral ischemia-induced VD models, and has the potential to reverse spatial learning damages and memory dysfunction.

Objective:To investigate the effect of TRIM52 antisense RNA 1 (TRIM52-AS1) on injury of brain cortex neurons induced by hypoxia/re-oxygenation (H/R) and its possible mechanism in rats.Methods:(1) The cortical neurons were cultured in vitro and divided into control group and model group. In the model group, H/R-induced cell injury models were prepared. Real-time quantitative PCR (RT-qPCR) was used to detect the expressions of TRIM52-AS1 and micro RNA (miR)-28-5p in neurons of the two groups. (2) The cortical neurons were divided into control group, model group, small interfering (si)-TRIM52-AS1 transfection group, and nonsense sequence transfection group. Cells in the model group were prepared for H/R-induced cell damage models. After cells in the latter two groups were transfected with si-TRIM52-AS1 or its nonsense control sequence for 6 h, they were prepared for H/R-induced cell damage models. RT-qPCR, CCK-8, and flow cytometry were used to detect the TRIM52-AS1 expression, and proliferation and apoptosis of neurons in the 4 groups; enzyme-linked immunosorbent assay (ELISA) and Western blotting were used to detect the lactate dehydrogenase (LDH), malondialdehyde (MDA) and superoxide dismutase (SOD) levels and protein expressions of Cyclin D1, Bcl-2 and Bax. (3) The cortical neurons were divided into miR-28-5p transfection group and nonsense sequence transfection group. After the cells were respectively transfected with miR-28-5p or its nonsense control sequence for 6 h, they were prepared for H/R-induced cell damage models. RT-qPCR, CCK-8, and flow cytometry were used to detect the TRIM52-AS1 expression, and proliferation and apoptosis of neurons in the two groups; ELISA and Western blotting were used to detect the LDH, MDA and SOD levels and protein expressions of Cyclin D1, Bcl-2 and Bax. (4) The cortical neurons were divided into wild-type TRIM52-AS1+miR-28-5p mimic transfection group, wild-type TRIM52-AS1+miR-28-5p nonsense control sequence transfection group, mutant TRIM52-AS1+miR-28-5p mimic transfection group, and mutant TRIM52-AS1+miR-28-5p nonsense control sequence transfection group. After the cells were co-transfected for 6 h, the culture medium was replaced with fresh medium and they were cultured for another 12 h, and then, the luciferase activity of cells in each group was detected by dual luciferase reporter gene experiment. (5) Cortical neurons were divided into nonsense sequence transfection group, miR-28-5p inhibitor transfection group, nonsense sequence+si-TRIM52-AS1 transfection group, and miR-28-5p inhibitor+si-TRIM52-AS1 transfection group, and these cells were transfected with miR-28-5p inhibitor nonsense control sequence, miR-28-5p inhibitor, miR-28-5p inhibitor nonsense control sequence+si-TRIM52-AS1, miR-28-5p inhibitor+si-TRIM52-AS1; 6 h after transfection, H/R-induced cell damage models were prepared. RT-qPCR, CCK-8, and flow cytometry were used to detect the miR-28-5p expression, proliferation and apoptosis of neurons in the two groups, respectively; ELISA and Western blotting were used to detect the LDH, MDA and SOD levels and protein expressions of Cyclin D1, Bcl-2 and Bax. Results:(1) As compared with those in the control group, the TRIM52-AS1 expression statistically increased, but the miR-28-5p expression significantly decreased in the model group ( P<0.05). (2) As compared with the control group, the model group and nonsense sequence transfection group had significantly increased LDH content in the culture supernatant, statistically decreased MDA and SOD content in the cells, significantly decreased A value and protein expressions of Cyclin D1 and Bcl-2, but significantly increased apoptosis rate and Bax protein expression ( P<0.05). As compared with the model group and nonsense sequence transfection group, si-TRIM52-AS1 transfection group had significantly decreased si-TRIM52-AS1 expression, statistically decreased LDH content in the culture supernatant and MDA content in the cells, significantly increased SOD content in the cells, significantly increased A value and protein expression of Cyclin D1 and Bcl-2, significantly decreased apoptosis rate and Bax protein expression ( P<0.05). (3) As compared with the nonsense sequence transfection group, the miR-28-5p transfection group had significantly increased miR-28-5p expression, significantly decreased LDH content in the culture supernatant and MDA content in the cells, and significantly increased SOD content in the cells, significantly increased A value and protein expressions of Cyclin D1 and Bcl-2, but significantly decreased apoptotic rate and Bax protein expression ( P<0.05). (4) The luciferase activity of the wild-type TRIM52-AS1+miR-28-5p mimic transfection group was significantly lower than that of the wild-type TRIM52-AS1+miR-28-5p nonsense control sequence transfection group (0.43±0.04 vs. 1.00±0.09, P<0.05). (5) As compared with the nonsense sequence transfection group, the miR-28-5p inhibitor transfection group had significantly decreased miR-28-5p expression, significantly increased LDH content in the culture supernatant and MDA content in the cells, significantly decreased SOD content in the cells, significantly decreased A value and protein expressions of Cyclin D1 and Bcl-2, but significantly increased apoptosis rate and Bax protein expression ( P<0.05). As compared with the nonsense sequence+si-TRIM52-AS1 transfection group, the miR-28-5p inhibitor+si-TRIM52-AS1 transfection group had significantly decreased miR-28-5p expression, significantly increased LDH content in the culture supernatant and MDA content in the cells, significantly decreased SOD content in the cells, significantly decreased A value and protein expressions of Cyclin D1 and Bcl-2, but significantly increased apoptosis rate and Bax protein expression ( P<0.05). Conclusion:TRIM52-AS1 down-regulation may inhibit H/R-induced oxidative stress and apoptosis of rat brain cortical neurons by negatively regulating the miR-28-5p expression, which reduces neuronal cell damage.