ISSN:
1573-6903
Schlagwort(e):
Astrocytes
;
primary culture
;
differentiation
;
ethanolamine base exchange enzyme
Quelle:
Springer Online Journal Archives 1860-2000
Thema:
Medizin
Notizen:
Abstract The enzymatic activities of ethanolamine base exchange (EBEE) and CDP-ethanolamine: 1,2-diacylglycerol ethanolamine phosphotransferase (EPT) were investigated during the growth of rat astrocyte primary cultures. From the 16th day, cells ceased to divide (2.0×106 cells per culture dish); the total phospholipid (PL) content increased 1.5 fold between the 16th and 24th day (0.20 to 0.30 μmol per mg protein) but the amount of ethanolamine phospholipid (28% of PL content) remained constant. Whereas the specific activity (pmol/ min × mg protein) of EPT reached a plateau at 16 days in culture and remained constant (400) thereafter, that of EBEE increased up to the 19th day (190) and decreased gradually to a basal level (75) at the 24th day. EBEE activity was not detected in plasma membranes isolated from 16, 19 and 24 days astrocyte cultures. Sub-cellular fractionation and determination of EBEE specific activities showed that (1) the 104×103 g fraction (P4) was 4.8 and 8.8 fold enriched at the 16th day and 24th day respectively as compared to the whole cell homogenate (50 and 75). (2) the 7×103 g (P2) and 17×103 g (P3) fractions were 8.4 and 7.0 fold enriched respectively at the 19 day in culture. The percentages of the enzymatic activity in the different subcellular fractions were 30, 57.2 and 25.7 for P2 and 39.2, 2.6 and 39.8 for P4 at 16, 19 and 24 days in culture respectively. The activity remained constant in P3 (23%) and was negligible in P1 (6%). Ultrastructual studies revealed that P2 and P3 were enriched in mitochondria while P4 contained essentially microsomes. P4 was enriched in glucose-6-phosphatase activity (G-6-P microsomal marker) and P2 and P3, in monoaminooxidase (MAO) and succinate dehydrogenase (SDH) (mitochondrial markers); G-6-P, MAO and SDH in the different subcellular fractions remained constant from the 16th to the 24th day. These data indicate (1) that the rate and profile of EPT and EBEE activities differed during the differentiation of astrocyte culture; (2) that EBEE activity, except at the 19th day in culture, was mainly localized in a microsomal subcellular fraction; (3) that at the 19th day the optimal EBEE activity observed in whole cell homogenate correlates with an enrichment of this activity in an enriched mitochondrial subcellular fraction.
Materialart:
Digitale Medien
URL:
http://dx.doi.org/10.1007/BF00993249
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