Astroglial-Conditioned Media and Growth Factors Modulate Proliferation and Differentiation of Astrocytes in Primary Culture

Abstract

Astroglial conditioned media (ACM) influence the development and maturation of cultured nerve cells and modulate neuron-glia interaction. To clarify mechanisms of astroglial cell proliferation/differentiation in culture, incorporation of [methyl-3H]-thymidine or [5,6-3H]-uridine in cultured astrocytes was assessed. Cultures were pre-treated with epidermal growth factor (EGF), insulin (INS), insulin-like growth factor-I (IGF-I), and basic fibroblast growth factor (bFGF) and subsequently with ACM. DNA labeling revealed a marked stimulatory effect of ACM from 15 days in vitro (DIV) cultures in 30 DIV astrocytes after 12 h pre-treatment with growth factors. The main effects were found after INS or EGF pre-treatment in 30 DIV cultures. ACM collected from 15 or 60 or 90 DIV increased RNA labeling of 15 and 30 DIV astrocyte cultures, being the highest value that of 30 DIV cultures added with ACM from 90 DIV. The findings of increased DNA labeling after EGF or INS pre-treatment in 30 DIV cultures, followed by addition of ACM from 15 DIV cultures, suggest that these phenomena may depend by extra cellular signal-regulated kinase 1 (ERK1) activation.

Effect of Acetylcholine Precursors on Proliferation and Differentiation of Astroglial Cells in Primary Cultures

“…Primary astroglial cell cultures were characterized by Glial Fibrillary Acidic Protein (GFAP) and glutamine synthetase activity as previously reported [3,5,8]. This analysis revealed that approximately 90% of cells were GFAPpositive, indicating that cultures were enriched in astrocytes (data not shown).…”

“…In our previous researches [5,6,9] we reported findings showing growth rate and differentation of astroglial cells in culture. In particular, it has been evaluated DNA and RNA lobeling and ERK1 immunoactiviy in astrocyte cultures preatrated with growth factors (GFs) and treated with astroglial conditioned media (ACM) [5] and it has been found an up and down regulation of nucleic acid labelling.…”

“…Primary cultures of astrocytes were performed from newborn albino rat brains (1-2-day-old Wistar strain) according to the methods previously reported [5]. Briefly, cerebral tissues, after dissection and careful removal of the meninges, were mechanically dissociated through sterile meshes of 82-mm pore size (Nitex).…”

“…-dependent-cross linking of the carboxamide moiety of a peptide-bound glutamine either to the e-amino group of a peptide-bound lysine, or to polyamines, with liberation of ammonia [11,17]. TG-2 is the only member of TGs playing a role in cell signaling transduction, differentiation and apoptosis [5,19]. Cell cycle-related proteins including cyclins [20] play a crucial role during astroglial cell proliferation in culture and may be considered, as an important marker to evaluate the involvement of different compounds on growth, development and maturation of our in vitro model.…”

Growth Factors and Steroid Mediated Regulation of Cytoskeletal Protein Expression in Serum-Deprived Primary Astrocyte Cultures

“…Primary cultures of astrocytes were prepared from newborn albino rat brains (1-2-day-old Wistar strain) as previously described [7,14,15]. In particular, cerebral tissues, after dissection and careful removal of the meninges, were mechanically dissociated through sterile meshes of 82 mm pore size (Nitex).…”