肌源神经营养因子(MDNF)对体外培养大鼠脊髓运动神经元划伤后Bcl-2表达的影响

EFFECTS OF MUSCLE-DERIVED NEUROTROPHIC FACTOR ON BCL-2 EXPRESSION OF RAT SPINAL CORD MOTONEURONS AFTER SCRATCH INJURY IN VITRO

为检测大鼠肌源神经营养因子对体外培养大鼠脊髓运动神经元划痕损伤后 Bcl-2表达的影响 ,本实验从成鼠和乳鼠骨骼肌中提取出肌源神经营养因子。在胎鼠脊髓运动神经元培养第 7d时将其随机分成对照组和实验组 (成鼠组和乳鼠组 ) ,取 10 0μl肌源神经营养因子 ( 0 .1μg/ ml)加入实验组 ,对照组加入等量的 PBS,同时进行划痕损伤。损伤后第 3d计数各组存活的运动神经元 ,行 Nissl染色和免疫组化反应 ,计数 Bcl-2免疫反应 ( Bcl-2 -IR)阳性神经元数量 ,并在图像分析仪上对 Bcl-2 -IR阳性神经元作光密度分析。结果发现 ,损伤后第 3d,实验组运动神经元存活数 ,Bcl-2 -IR阳性神经元数和平均光密度均明显高于同期对照组 ,而乳鼠肌源神经营养因子组功效优于成鼠肌源神经营养因子组。结果提示 ,大鼠肌源神经营养因子对体外培养的受损运动神经元具有增强 Bcl-2表达、减少凋亡和损伤修复等作用 。

In order to study the effect of Muscle-derived neurotrophic factor (MDNF) on Bcl-2 expression of rat spinal cord motoneurons after scratch injury in vitro, MDNF was respectively extracted from adult and neonatal rat, scratch injury was executed on 7th day of serum culture and at the same time, 100 μl MDNF(0.1 μg/ml) was added into culture medium for neurons from spinal cords of embryonic rats. The equal volume of PBS was added into the medium for control. On 3rd day after scratch injury, spinal cord motoneurons were stained by Nissl's method and immunohistochemical antiserum of Bcl-2. The number and the mean optical densities of Bcl-2 immunoreactive (Bcl-2-IR) positive neurons were observed. It was found that not only the number of survival and Bcl-2-IR positive neurons but also the mean optical densities of Bcl-2-IR positive neurons in MDNF was significantly increased on 3rd day after scratch injury, as compared with control. The effects of MDNF from neonatal rat were better than those from adult rat on 3rd day after scratch injury. Our results showed that MDNF can increase Bcl-2 expression and decrease motoneuron apoptosis induced by injury in vitro. MDNF from neonatal rat has a longer effect than that from adult rat on injured motoneurons. (Figures 1～4 on plate 23)