肌源神经营养因子(MDNF)对体外培养的大鼠脊髓运动神经元缺氧时p53表达的影响

本研究目的是观察缺氧时肌源神经营养因子(MDNF)对体外培养的大鼠脊髓运动神经元p53蛋白表达的影响。从成鼠和胎鼠骨骼肌中提取MDNF。在胎鼠脊髓运动神经元培养第7 d时将其分成对照组和实验组(包括成鼠MDNF组和胎鼠MDNF组),取100μl MDNF(0.1μg/ml)加入培养大鼠脊髓运动神经元的培养液中,对照组加入等量的PBS。然后用液体石蜡封闭液面造成神经元缺氧损伤,缺氧3 d后,应用Nissl染色、原位末端标记、免疫组化方法,对损伤的大鼠脊髓运动神经元进行检测。结果表明:培养的大鼠脊髓运动神经元缺氧3 d后,经MDNF孵育可使脊髓运动神经元TUNEL、p53表达均较对照组明显减弱,神经元损伤程度减轻,神经元存活数明显高于对照组,且胎鼠MDNF的效果更好。提示:大鼠脊髓运动神经元缺氧后可出现神经细胞凋亡,但胎鼠MDNF较成鼠MDNF更能减弱缺氧时脊髓运动神经元p53的表达,抑制缺氧后神经元的死亡。

EFFECTS OF MUSCLE-DERIVED NEUROTROPHIC FACTOR ON p53 EXPRESSION IN RAT SPINAL CORD MOTONEURON AFTER ANOXIA IN VITRO

The present study was to observe the effects of Muscle-derived Neurotrophic Factor(MDNF) on p53 expression in rat spinal cord motoneurons after anoxia in vitro. MDNF was respectively extracted from adult and fetal rat, 100μl MDNF(0. 1μg/ ml) was added into culture medium for motoneurons from spinal cords of embryonic rats. The equal volume of PBS was added into the medium for control. Then liquid paraffin was added into the medium to create the anoxia injury neurons. Three days later, spinal cord motoneurons were stained by Nissl's method, terminal deoxynucleotidyl transferase-mediated dUPT nick end labeling (TUNEL) and immunohistochemical antiserum of p53. The results showed that the number of the living neurons and the arbitrary densities of p53 immunoreactive (p53-IR) as well as TUNEL positive neurons were observed. It was found that TUNEL motoneurons in MDNF group were much more than that in control, the arbitrary densities of p53 immunoreactive (p53-IR) and TUNEL motoneurons in MDNF group significantly decreased in comparison with control, the effects of MDNF from fetal rat were better than the ones from adult rat. The present results indicate that cell apoptosis is observed; we presume that MDNF from fetal rat can decrease p53 expression of rat spinal cord motoneurons after anoxia in vitro much more than MDNF from adult and prevent the death of rat spinal cord motoneurons after anoxia in vitro.