脂肪源性干细胞穴位注射对大鼠脑缺血/再灌注后海马中血管生成的影响

目的探讨脂肪源性干细胞(ADSC)穴位注射对大鼠脑缺血/再灌注损伤后海马中血管生成的影响。方法24只SD大鼠随机分为假手术组、模型组、对照组(穴位注射生理盐水)和治疗组(穴位注射ADSC),每组6只。线栓法制作大鼠大脑中动脉缺血/再灌注模型,治疗组在大鼠大椎、双侧内关穴位注射PKH-26标记ADSC细胞悬液,对照组相同穴位注射同剂量生理盐水。缺血72 h后行神经功能损害评分(NSS),采用免疫组织化学法及实时荧光定量聚合酶链反应检测血管内皮生长因子(VEGF)蛋白和mRNA表达,免疫组织化学法进行CD34染色计数微血管密度(MVD)。结果脑缺血后72 h,治疗组大鼠海马区可见PKH-26标记的ADSC表达,而其他3组未发现红色荧光信号。模型组、对照组和治疗组NSS评分高于假手术组(P<0.05);与模型组比较,对照组和治疗组NSS评分显著降低(P<0.05),其中治疗组NSS评分低于对照组(P<0.05)。假手术组海马区中未见VEGF蛋白和mRNA表达,模型组、对照组和治疗组VEGF蛋白和mRNA表达及MVD较假手术组显著增加(P<0.05),其中治疗组VEGF蛋白和mRNA表达及MVD较模型组和对照组显著增高(P<0.05),而模型组和对照组3个指标比较差异无统计学意义(P>0.05)。结论穴位注射ADSC对大鼠脑缺血/再灌注有神经保护作用,其作用机制可能与促进海马VEGF表达,增加MVD有关。

Effect of acupoint injection with adipose-derived stem cell on angiogenesis after cerebral ischemia-reperfusion in rats

Objective To investigate the effect of acupoint injection with adipose-derived stem cell(ADSC) on angiogenesis after cerebral ischemia-reperfusion in rats.Methods A total of twenty-four SD rats were randomly divided into sham operation group,model group,control group(saline acupoint injection) and treatment group(ADSC acupoint injection),six rats in each group.The cerebral ischemia-reperfusion model of middle cerebral artery in rats was established by suture occlusion method.ADSC suspensions were marked with PKH-26 which was infused into acupoints of "Dazhui"and"Neiguan" in the treatment group,and saline was infused in the control group.The neurological function score were observed by neurological severity scores(NSS)at 72 hours after reperfusion.The expression of VEGF was detected by using imunohistochemistry and real-time fluorescent quantitative-polymerase chain reaction(RT-PCR) and microvessel density(MVD) was evaluated by immunohistochemistry with the specific antibody CD34.Results The ADSC which was labeled by PKH-26,was observed in hippocampus in the treatment group,and no red fluorescence signal was observed in the other three groups at 72 hours after reperfusion.Compared with sham operation group,the NSS of model group,control group and treatment group increased(P<0.05);the NSS of control group and treatment group were lower than the model group(P<0.05),and the NSS of treatment group was lower than control group(P<0.05).There was no the protein and mRNA expressions of VEGF in hippocampus region in sham operation group.Compared with sham operation group,and the protein and mRNA expressions of VEGF,MVD were significantly enhanced in model group,control group and treatment group(P<0.05).Compared with model group and control group,the protein and mRNA expressions of VEGF,MVD of the treatment group were significantly increased(P<0.05),however,there was no difference of the expression of VEGF,MVD between model group and control group(P>0.05).Conclusion ADSC acupoint injection could improve the neurological function in rats after cerebral ischemia-reperfusion,which may be related to up-regulating VEGF and MVD in hippocampus region.