大鼠脑缺血再灌注损伤脑匀浆上清液对骨髓基质细胞的影响

Effects of rat cerebral ischemia-reperfusion injury homogenate supernatant liquid on bone marrow stromal cells

目的研究缺血再灌注损伤脑匀浆上清液对大鼠骨髓基质细胞(BMSCs)形态、活力、神经元特异性烯醇化酶(NSE)、电生理和神经营养因子(NTFs)表达情况的影响。方法将体外培养的大鼠BMSCs分别置于实验组、假手术组及空白对照组培养条件中,采用组织学、台盼蓝染色、免疫组织化学、膜片钳、半定量RT-PCR等方法,观察细胞形态改变,测定活性,鉴定NSE表达,记录全细胞膜电流,检测NGF、GDNF、BDNF和bFGF在各时间点mRNA表达情况。结果实验组细胞形态发生神经元样改变,细胞活力良好,NSE阳性率、外向K+电流峰值及电流密度、NTFs的表达水平高于其他组,但未引出动作电位。结论大鼠BMSCs在脑缺血再灌注损伤的细胞微环境中生长良好,可以诱导分化为神经元样细胞,但无神经元的电生理特征,NTFs的表达能力明显增强。

【Objective】To research the morphous,viability,NSE,electrophysiology and expression of neurotrophic factors（NTFs） in bone marrow stromal cells（BMSCs） of rat on the effects of cerebral ischemia-reperfusion injury homogenate supernatant liquid.【Methods】BMSCs were cultured under the condition of experimental group,sham operation group and blank control group.Inverted microscope was used to observe morphological changes,Trypan blue dyeing was used to detect viability,indirect immunocytochemisty staining was used to assessed the expression of NSE,patch clamp was used to record whole cell current,Semi-Quantitative RT-PCR was used to detect mRNA expression levels of NGF,GDNF,BDNF and bFGF at 6,12,18,24,48 and 72 h.【Results】BMSCs under the condition of experimental group had morphological changes showed as neuron-like.Viability was fine.Positive rate of NSE,peak and density of outward K＋ currents and expression levels of NTFs were significantly higher compared with the other groups,but no active potential.【Conclusions】Rat BMSCs grew well in cell microenvironment of cerebral ischemia-reperfusion injury,had an advantage to the expression levels of NTFs,can be induce into neuron-like cell,but had no electrophysiological properties like mature neurons.