异丙酚促进体外培养的成年大鼠海马神经干细胞增殖

目的异丙酚对于成年神经干细胞增殖能力的影响目前尚不明确,本研究对异丙酚对体外培养的大鼠成年神经干细胞的作用及其可能的分子机制进行了研究。方法体外培养的大鼠成年神经干细胞给予BrdU后,分别用10、50、100μg/ml的异丙酚进行处理,并同时设立溶媒对照和空白对照。24h后,采用免疫染色显示BrdU阳性细胞,DAPI复染细胞核。细胞存活率采用细胞计数法和MTT法测定。人工计数BrdU阳性细胞和固缩细胞核的比例。用Fura2-AM检测胞浆内Ca2＋浓度。分别用免疫荧光和免疫印迹法检测细胞内CREB和磷酸化CREB的表达位置和水平。结果低浓度（10μg/ml）异丙酚不能促进大鼠成年神经干细胞的增殖,而中浓度（50μg/ml）和高浓度异丙酚（100μg/ml）则呈剂量依赖性的促进其增殖。异丙酚提高了BrdU阳性细胞的比例。细胞死亡率在处理前后维持在低水平上（〈2%）。异丙酚显著提高了大鼠成年神经干细胞胞浆内的游离Ca2＋浓度。异丙酚处理前后,大鼠成年神经干细胞均能表达CREB和磷酸化CREB,但是异丙酚干预后,大鼠成年神经干细胞内磷酸化CREB表达水平显著增高。结论异丙酚可促进体外培养的大鼠成年神经干细胞的增殖水平,这是细胞存活率升高的主要原因,可能与胞浆内Ca2＋浓度以及CREB磷酸化水平的升高有关。

Propofol Promotes Proliferation of Cultured Adult Rat Hippocampal Neural Stem Cells

Objective The effect of propofol on the proliferation of adult neural stem cells （ANSCs） is unclear. We investigated the effect of propofol on cultured rat adult neural stem cells and the underlying molecular mechanisms. Methods Rat adult neural stem cells were treated with propofol （10,50,100 μg / ml）,vehicle or untreated. After 24 hours of treatment,BrdU incorporation method was performed and the nuclei were counterstained with 4＇ ,6-diamidino-2-phenylindole （DAPI）. The number of cells was enumerated by counting the viable cells and by the 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay. The proportions of BrdU-positive cells and pyknotic nuclei were also determined. Cytoplasmic calcium concentrations were determined with Fura 2-AM. The expression of CREB and phospho-CREB in cells was determined by immunostaining method. Phosphorylation level of CREB was analyzed by Western blot. Quantitative data were analyzed by ANOVA or student＇s t-test. P values less than 0.05 were considered as significant. Results Propofol was found to dose-dependently promote the proliferation of ANSCs at the concentrations of 50,100 μg / ml. Propofol increased the percentages of BrdU-positive cells. Percentages of cell death was found to maintain at a low level （2%）. Propofol also elevated the concentration of cytoplasmic free calcium in ANSCs. Basal expression of CREB and phospho-CREB was found in ANSCs. Propofol was found to up-regulate the phosphorylation level of CREB in ANSCs. Conclusion Propofol can promote the proliferation of ANSCs in vitro. Elevation of cytoplasmic calcium and the subsequent up-regulation of phosphorylation of CREB in ANSCs might contribute to the growth promotion.