Different distribution of thiaminpyrophosphatase activity in neuronal and glial cell enriched fractions from human and rat brain: an isoelectric focusing investigation.

Basic and applied histochemistry Pub Date : 1990-01-01
U Laforenza, P Mazzarello, C Patrini, M Poloni, G P Casadei, G Rindi
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Abstract

Enriched fractions of neuronal and glial cells from rat and human (autoptical specimens of frontal cortex) brains were assayed for thiaminpyrophosphatase (TPPase) activity by isoelectric focusing (IEF). Glial and neuronal fractions yielded about 0.16 and 0.03 g/g of wet tissue respectively. Purity was estimated to be 85-90%. The isolated fractions were homogenized in the presence of 1% Triton X-100, and IEF was carried out on thin layer polyacrylamide gel in a pH range of 3.5-9.5, using Ampholine PAG plates. TPPase activity of the protein bands was assayed by laser-densitometry, after incubation with thiaminpyrophosphate in an appropriate buffer and PbS staining. IEF analysis showed that TPPase activity was present almost exclusively in the neuronal fraction (at levels 16-fold higher than those found in the glial fraction). After IEF, TPPase activity was present in 10 distinct protein bands with different isoelectric points. These preliminary data suggest that TPPase may be involved in the neuronal activity of rat and human brain.

硫胺焦磷酸酶活性在人脑和大鼠脑神经元和胶质细胞富集组分中的不同分布:等电聚焦研究。
用等电聚焦(IEF)法测定了大鼠和人脑(额叶皮质自解剖标本)中富集的神经元和胶质细胞的硫胺焦磷酸酶(TPPase)活性。胶质和神经元部分分别产生约0.16和0.03 g/g的湿组织。纯度估计为85-90%。分离的部分在1% Triton X-100存在下均质,用Ampholine PAG板在pH为3.5-9.5的薄层聚丙烯酰胺凝胶上进行IEF。在适当的缓冲液中与焦磷酸硫胺孵育并PbS染色后,用激光密度法测定蛋白带的TPPase活性。IEF分析显示,TPPase活性几乎只存在于神经元部分(其水平比神经胶质部分高16倍)。经IEF后,TPPase活性出现在10个不同等电点的蛋白条带上。这些初步数据提示,TPPase可能参与了大鼠和人脑的神经元活动。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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