This study investigated the effects of epidermal growth factor (EGF) on the labeling of various RNA species and of nuclear proteins in primary rat astroglial cell cultures. After 12 hours of EGF treatment in serumfree medium or chemically defined medium, significant increase in RNA labeling, and also in acid-soluble radioactivity and RNA content, was observed. The ratio RNAIDNA was significantly higher in EGFtreated cultures compared with controls. Ribosomal RNAs (28s and BS), polyadenylated, and nonpolyadenylated RNAs showed a higher specific radioactivity in EGF-treated cultures. Among the nuclear proteins, the labeling of basic proteins was enhanced by EGF treatment, whereas that of total nuclear acidic protein (NHPs) was less modified, except for some NHPs separated by gel electrophoresis with a molecular weight (MW) approximately 95-83 and 44 kd, which were significantly more labeled in EGF-treated cultures.
basal medium supplemented with fetal calf serum (FCS) develop into a highly enriched astroglial culture. It is well known that serum contains many undefined components, some of which may interfere with the effects mediated by the added factors.
In the present study, cells were grown in 10% FCS and then, during EGF treatment, shifted to three different media: (I) FCS-supplemented medium (SSM); (2) serumfree basal nutrient medium without any supplement (SFM); (3) chemically defined medium (CDM) supplemented with insulin, transferrin, and fatty acid-free bovine serum albumin. Indeed, basal nutrient medium supplemented with insulin, transferrin, and BSA has been shown able to support the growth of cultured astroglial cells for few weeks (Weibel et al., 1984).
The results demonstrated that EGF treatment increases RNA and protein synthesis in astroglial cell cultures, thus showing that EGF affects the macromolecular events occurring in glial cell proliferation and differentiation. Kev words: epidermal growth factor, protein labeling, Poly(A)=RNA and Poly(A)+RNA, basic and acidic nuclear proteins, astroglial cell cultures MATERIALS AND METHODS Materials EGF (prepared from mouse submaxillary glands), INTRODUCTION bovine pancreas insulin, human transferrin, and fatty It has been shown recently that EGF is a potent acid-free bovine serum albumin were purchased from mitogen for cultured astrocytes (Simpson et al., 1982), Sigma Chemical Company (St. Louis, USA). DMEM, which suggests that this peptide may have a role in the penicillin G, streptomycin sulfate, and FCS were purdevelopment of the central nervous system. Specific, saturable high-affinity receptors for EGF are present in Abbreviations used: DMEM, Dulbecco's modified Eagle's mecultured rat cerebral astrocytes (Simpson et al., 1982). dium; EGF, epidermal growth factor; FGF, fibroblast growth factor;
The aim of the present study was to investigate the FCS, fetal calf serum; GFAP, glial fibrillary acidic protein; PBS, phosphate buffered saline solution; SDS, sodium dodecyl sulphate; astroglial Cells in primary culture. In particular, the effect semm-supplemented medium; CDM, Chemically defined medium; of EGF treatment on the labeling of poly(A)-RNA, SFM, serum-free medium; MW, molecular weight; NHP, nonhistone poly(A)+mRNA and nuclear proteins was investigated. protein.
Cells dissociated from cerebral hemispheres of