目的:观察不同剂量银杏内酯B(GB)对缺氧缺血性脑损伤(HIBD)新生大鼠内源性神经干细胞增殖分化的影响.方法:清洁级7d龄SD大鼠96只,随机分为假手术组、模型组、低剂量组及高剂量组,后3组采用经典Rice法制作HIBD动物模型,模型制作4h后低剂量组与高剂量组分别按5mg/kg、10mg/kg腹腔注射GB,其他两组分别注射等量生理盐水,每日1次,共5d.每组随机分别在造模后第3,7,14,28天处死,单标、双标免疫组化技术观察4组大鼠海马齿状回颗粒下层区(SGZ)溴脱氧尿嘧啶核苷(BrdU+)及皮质BrdU+/nestin+(聚蛋白)、BrdU+/NSE+(神经元特异性烯醇化酶)、BrdU+/GFAP+(胶质纤维酸性蛋白)阳性细胞的表达,并计数分析.结果:HIBD后BrdU+、皮质BrdU+/nestin+、BrdU+/NSE+、BrdU+/GFAP+细胞数增加,低剂量组、高剂量组均高于模型组,GB高剂量组的阳性细胞数高于GB低剂量组.结论:GB能提高HIBD新生鼠内源性神经干细胞增殖、分化的能力,提示其可以促进神经发生.

Objective To observe the effects of ginkgolide B (GB) on mRNA expression of foxgl and proliferation of cells in brain tissue of newborn rats with hypoxic-ischemic brain damage (HIBD).Methods A total of 128 clean 7-day-old healthy SD rats were randomly divided into sham operation group,the model group,the low-GB dose and the high-GB dose treatment groups.Classic Rice method were used to establish HIBD models in the latter 3 groups.Four hours after operation,and GB in dose of 5 mg/kg and 10 mg/kg was given to rats in the low and the high dose treatment groups by intraperitoneal injection postoperatively,once a day for 5 days,while sham operation and model groups were treated with equal physiological saline.All groups were respectively sacrificed on 3 d,7 d,14 d,28 d respectively.Quantitative real-time fluorescent polymerase chain reaction was employed to detect expression of Foxg1 gene.Then the number of 5-bromodeoxyuridine positive cell in subgranular zone was investigated by immunolluorescent stairning.Results The Foxg1 mRNA expression was observed 3 days after HIBD,peaked on 7th day,and then declined gradually; the levels of Foxg1 mRNA in the 2 treatment groups were higher than that of the HIBD group (all P ＜ 0.01) ; The expression of Foxgl at 7 d,14 d,28 d,in high-dose group were higher than those in the low-dose group (all P ＜ 0.01).The number of 5-bromodeoxyuridine positive cell was increased after HIBD,and the levels in the low-and the high-dose treatment groups were all higher than that of the model group (all P ＜ 0.05) ; the number of positive cell in high-dose treatment groups were higher than that in the low-dose treatment groups (P ＜ 0.05).Conclusions GB can promote the expression of Foxg1 gene and improve the proliferation of cells in Brain tissue after HIBD,which shows more significant efficacy in high-dose group than in low-dose group.

Aim: To observe the effect of ginkgolide B ( GB) on mRNA expression of Caspase-3 and vascular endothe-lial growth factor ( VEGF) in brain tissue of hypoxic-ischemic brain damage ( HIBD) newborn rats. Methods: A total of 96 clean 7-day-old health SD rats were randomly divided into sham operation group,the model group,low and high dose GB treatment groups. Classic Rice method was used to establish HIBD model in the latter 3 groups. 4 h after operation,GB at dose of 5 mg / kg and 10 mg / kg was given to rats in the low and high dose GB treatment groups by intraperitoneal injection postoperation,once a day for 5 days,sham operation and model groups were given equal physiological saline. 6 rats in each group were sacrificed at the 3rd,7th,14th and 28th day after model establishment,respectively. Quantitative real-time fluo-rescent PCR was employed to detect the expressions of Caspase-3 and VEGF mRNA in brain tissue. Results: The expres-sions of Caspase-3 and VEGF mRNA had no significant differences among 4 time points in sham operation group ( F =0. 134,0. 382,P 0. 05) ,but in the model group,low and high dose GB treatment groups,the expressions of Caspase-3 and VEGF mRNA reached the peak at the 3rd day,then decreased,and reached the level of the sham operation group at the 28th day after operation( F = 109. 356,33. 281,9. 703 and 39. 945,103. 903,60. 956,P 0. 001) . At the 3rd,7th,14th day after operation,the expressions of Caspase-3 and VEGF mRNA had significant differences among the 4 groups ( F = 128. 865 and 97. 912,40. 058 and 57. 121,24. 184 and 8. 621,P 0. 001) ; compared with sham operation group,the ex-pressions of Caspase-3 and VEGF mRNA in model group were higher,while the expression of Caspase-3 mRNA was lower and that of VEGF mRNA was higher in low and high dose GB treatment groups when compared with the model group( P 0. 05) ,and the changes of high dose GB treatment group was more significant( P 0. 05) . Conclusion: GB can repair brain damage after HIBD through down-regulating Caspase-3 expression and up-regulating VEGF expression.

Objective To observe the effect on Foxg1 gene expression in the subgranular zone (SGZ) of cerebral tissue from neonatal rats with hypoxic-ischemic brain damage (HIBD) after transplantation of neural stem cells (NSCs) derived from umbilical cord blood.Methods Mononuclear cells separated from umbilical cord blood by density gradient centrifugation were cultured with orientated induction to differentiate the NSCs.The neuronal phenotype was identified using immunocytochemical methods.A total of 150 Sprague-Dawley rats were randomly divided into a sham-operation group,an HIBD group and an HIBD-NSCs group.Rats in the HIBD group and the HIBD-NSCs group were subject to ligation of the left carotid artery and then kept in a box under 8％ oxygen and 92％ nitrogen for 2.5 hours to establish the HIBD animal model.The artery was separated but not ligated in the sham operation group,which was not subjected to hypoxia.Twenty-four hours after the operation,the cultivated NSCs were transplanted by caudal vein injection into the rats in the HIBD-NSCs group.Rats were then sacrificed on the 3rd,7th,14th,21st and 28th days after the operation.Foxg1 gene expression in the SGZ was examined using in-situ hybridization methods.Results The number of Nestin-positive cells peaked on the 6th day of cultivation and then decreased by the 9th day.The Foxg1 gene was expressed in the SGZs of each group.The expression increased by the 3rd day after surgery in the HIBD and HIBD-NSCs groups,and peaked on 7th day after the operation,then declined gradually.The average expression level of Foxg1 in the HIBD group was significantly lower than that in the HIBD-NSCs group on the 7th day and thereafter.Conclusions Human umbilical cord blood mesenchymal stem cells can be induced and differentiated into neural stem cells.Foxg1 genes can still be present in the SGZ after birth.HIBD can induce the expression of Foxg1 genes.Transplanting NSCs can promote the expression of Foxg1 genes and improve morphological and functional recovery after HIBD,at least in neonatal rats.

Objective To study the protective therapeutic effect of insulin-like growth factor-1(IGF-1)on endogenous neural stem cells of newborn rats with hypoxic-ischemic brain damage(HIBD),by intranalsal delivery of medicine via the olfactory pathway.Methods Ninety SD rats at 7 days were randomly divided into 3 groups: sham-operated group,model saline control group,and IGF-1-treated group.The HIBD models were induced by Rice method.Intranasal delivery of 0.1 mL saline was applied to model saline control group,and IGF-1-treated group was delivered by intranasally with IGF-1 2.5 μg(dissolved with 0.1 mL saline) after HIBD 1 hour,in sham-operated group the common carotid arteries were only split without ligation or hypoxia management.All animals were killed respectively on the 1st,3rd,5th,7th,14th day after hypoxia.Immunohistochemistry was used to detect the expressions of BrdU and Nestin in sub ventricular zone of newborn rats;the numbers of BrdU positive cells,Nestin positive cells and BrdU-Nestin double lable cells;Hematoxylin-eosin staining was used to learn the changes in histological structure with brain injury.Results Model saline control group: after HIBD,BrdU positive cells,Nestin positive cells and BrdU-Nestin double lable cells became to increase on the 1st day,the number of cells reached top on the 3rd day,then reduced obviously on the 7th day and only a few positive cells were in SVZ on the 14th day;IGF-1-treated group:3 kind of positive cells increased visibly on the 1st day,added sharply on the 3rd day and became highest on the 5th day,then began to lessen on the 7th day,on the 14th day,the cells decreased distinctly.Compared with model saline control group,the expression of cells in IGF-1-treated group were higher at the same time(Pa0.05);There was no proliferation appearance in sham-operated group,compared with the other 2 groups(P0.05).Conclusion The IGF-1 deliveried intranasally have significant neuroprotective and therapeutic effect for HIBD.