大鼠胚胎小脑提取液神经营养活性成分的分析

本实验观察了不同胎龄大鼠的小脑提取液（ｃｅｒｅｂｅｌｌｕｍ ｅｘｔｒａｃｔｓ， ＣＥ）对体外培养小脑及脊髓神经细胞的影响。并采用聚丙烯酰胺凝胶电泳（ＳＤＳ ＰＡＧＥ）法鉴定了各胎龄的ＣＥ 的蛋白成分差异。制备不同蛋白浓度的１４、１６、１８、２０ ｄ 胎鼠（Ｅ１４，Ｅ１６，Ｅ１８，Ｅ２０）及新生１ ｄ 鼠（Ｐ１）的ＣＥ，加入原代培养的Ｅ１８ 胎鼠小脑及脊髓神经细胞悬液中，２４ ｈ 后以微量快速自动比色法检测细胞活性，发现实验各胎龄鼠的ＣＥ均能促进培养小脑及脊髓神经元的存活，并有蛋白浓度依赖关系，各胎龄都有各自不同的最适浓度，Ｅ１４、Ｅ１６ 和Ｅ１８ 的ＣＥ 显示了更强的神经营养活性，与对照组及其它实验组相比有极显著性差异（Ｐ＜ ０．００１）。以蛋白浓度为０．４ ｇ／Ｌ的Ｅ１８ 的ＣＥ培养新生３ ｄ 鼠（Ｐ３）脊髓细胞的结果表明，此ＣＥ可促进培养脊髓神经元的成熟和分化，抑制胶质细胞的增殖。电泳结果表明，Ｅ１６ 与Ｅ１８ 的ＣＥ中含有Ｐ１ 的ＣＥ中所没有的蛋白带，分子量分别约为２５．８ ｋＤ、４５．１ ｋＤ、８９．１ｋＤ 和１７９．５ ｋＤ，为寻找新的神经营养因子提供了线索。

STUDY ON NEUROTROPHIC COMPONENTS IN CEREBELLUM EXTRACTS OF EMBRYONIC RAT

The effects of cerebellum extracts (CE) from different embryonic ages on cultured rat cerebellar and spinal cells were studied. The different protein components of CE were proved by SDS polyacrylamide gel electrophoresis(SDS PAGE). CE from 14 d,16 d,18 d, 20 d embryonic rats(E14～20) and P1 newborn rats were made into different concentrations of protein, then were added to cultured cerebellar and spinal cells of E18 rats. The neuronal survival was determined by the auto colorimetric assay. The result showed that CE of all experimental ages could promote the survival of cerebellar and spinal neurons, and there was a protein concentration dependent relation, with different optimal concentration in different embryonic ages. E14, E16 and E18 CE showed more prominent neurotrophic activities, that made significant differences(P<0.001) with the control and other experimental groups. E18 CE with the concentration of 0.4 mg/ml was found to promote the maturation and differentiation of neurons, but inhibit the survival of glial cells in the culture of P3 spinal cell. The result of SDS PAGE showed that E16 and E18 but not P1 CE shared the same protein bands with molecular weights of about 25.8 kD, 45.1 kD, 89.1 kD, and 179.5 kD which might be target derived neurotrophic factors(TNTFs). (Figures 2～5 on plate 34)