摘要
目的探讨人脐血细胞(HUCBCs)静脉移植治疗自发性高血压(SHR)大鼠缺血再灌注脑损伤疗效及其相关机制。方法人脐血细胞体外分离、扩增并以4,6-2胺-2苯基吲哚(DAPi)标记。68只雄性SHR大鼠经线栓法建立大鼠大脑中动脉栓塞(MCAO)模型,并随机分为2组:治疗组和对照组。术后48h后经尾静脉治疗组注射1×106HUCBCs,对照组注射HUCBCs细胞培养基。于移植后3、14、28d,Bederson评分观察大鼠神经功能状况;图像分析法检测计算各组TTC染色脑梗死面积;荧光显微镜对治疗组脑组织DAPi染色阳性细胞计数;免疫组化法检测脑组织内脑源性神经营养因子(BDNF)染色阳性细胞数以及逆转录聚合链式反应(RT-PCR)检测移植后14、28d缺血侧脑组织BDNFmRNA表达水平。结果(1)移植后3d2组Bederson评分无显著差别,移植后14、28d治疗组显著低于对照组(P<0.05)。(2)移植后3d两组脑梗死面积无显著差别,移植后14、28d治疗组显著低于对照组(P<0.05)。(3)治疗组移植后3d梗死侧见少量DAPi阳性染色细胞,移植后14、28d梗死侧DAPi阳性染色细胞增加(P<0.05)。(4)免疫组化显示治疗组各时间点BDNF阳性细胞数较对照组增加(P<0.05)。(5)移植后14、28d治疗组缺血侧脑组织BDNFmRNA表达水平显著增加。结论HUCBCs静脉移植改善SHR大鼠脑缺血再灌注损伤的神经功能并缩小脑缺血损伤面积,其保护作用机制可能与经静脉移植的HUCBCs迁入脑组织内生存与增殖,并促进缺血侧脑组织BDNF表达水平增高相关。
Objective To investigate the effects and possible mechanisms of the intravenous transplantation of human umbilical cord blood cells （ HUCBCs ） on cerebral ischemic - reperfusion injury of spontaneously hypertensive rats （SHR）. Method HUCBCs were abstracted,purified and cultured in vitro and their nucleus were labeled by diamidino - phenyl- indole（DAPi）. The middle cerebral artery occlusion （MCAO） model was established by suture embolic method. 68 rats with MCAO were divided randomly into two groups ： treated and control groups. Each of the treated group was injec- ted intravenously with HUCBCs （1 × 10^6） at 48h after MACO, while each of the control group was injected intravenously with the same volume of cell culture medium. On 3d, 14d ,28d after transplantation, the recovery of neurological deficits was measured by Bederson neurological test. Quantitative histological determinations of infarct area were performed using stand- ard Trc staining and quantitative image analysis. The tracing and counting of DAPi positively stained cell was conducted under the inverted fluorescence microscope. The counting of brain -derived neurotrophic factors（BDNF） positively stained cells was detected with immunohistochemisry ,and reverse transcriptase polymerase chain reaction （RT- PCR） was used to reveal the expression level of the BDNF mRNA in the ischemic cerebral tissue on 14d and 28d after transplantation. Resuits （ 1 ） No difference was found in the Bederson scores between the two groups on 3d after transplantation, however, the score was significantly lower in the treated group on both 14d and 28d （P 〈0.05）. （2）There was no difference in the percent of cerebral infarcted area between two groups on 1 d after transplantation, but on 14d and 28d after treatment, the infarcted areas in the treated group were smaller significantly （ P 〈 0.05 ）. （ 3 ） A small quantity of DAPi positively stained cells in the ischemic encephalon of the treated group was found on 3 d after transplantation, then the number of DAPi positive cells was increased on 14d and 28d（ P 〈 0.05 ）. （4） At all examination,the number of BDNF positive cells located in the ische- mic encephalon of the treated group was significantly increased compared to that of the control group （P 〈 0.05 ）. （5）Com- pared to that of control group on 14d and 28d,the levels of BDNA mRNA expression were increased in the ischemic cerebral tissue of the treated group. Conclusions The intravenous transplantation of HUCBCs could obviously ameliorate neurologi- cal behavior and reduce the cerebral infarcted size in the cerebral ischemic reperfusion injury of SHRs. The possible mecha- nism for neurological protection in cerebral ischemic-reperfusion injury may be correlated to the survival and proliferation of HUCBCs and increased expression level of BDNF in the ischemic brain.
出处
《中风与神经疾病杂志》
CAS
CSCD
北大核心
2010年第1期16-19,共4页
Journal of Apoplexy and Nervous Diseases
基金
国家自然科学基金项目(30572079)
关键词
人脐血细胞
静脉移植
自发性高血压大鼠
腑缺血再灌注损伤
Human umbilical cord blood cells
Intravenous transplantation
Spontaneously hypertensive rat
Cerebral ischemic-reperfusion injury
