cAMP-induced Astrocytic Differentiation of C6 Glioma Cells Is Mediated by Autocrine Interleukin-6

Abstract

Elevation in the level of intracellular cAMP is known to induce the astrocytic differentiation of C6 glioma cells by unknown mechanisms. In this report, we show that cAMP-induced autocrine interleukin 6 (IL-6) promoted astrocytic differentiation of C6 cells. Treatment of cells with N 6 ,2 -O-dibutyryl cAMP (Bt 2 AMP) and theophylline caused the delayed phosphorylation of signal transducer and activator of transcription 3 (STAT3), as well as the expression of an astrocyte marker, glial fibrillary acidic protein (GFAP). Overexpression of the dominant-negative form of STAT3 leads to the suppression of GFAP promoter activity, suggesting that STAT3 activity was essential for cAMP-induced GFAP promoter activation. On the other hand, the IL-6 gene was quickly induced by Bt 2 AMP/theophylline, and subsequent IL-6 protein secretion was stimulated. In addition, recombinant IL-6 induced GFAP expression and STAT3 phosphorylation. Most importantly, treatment with IL-6-neutralizing antibody dramatically reduced the cAMP-induced GFAP expression and STAT3 phosphorylation and reversed the cellular morphological changes that had been caused by Bt 2 AMP/theophylline. Taken together, these results indicated that Bt 2 AMP/ theophylline lead to delayed STAT3 activation via autocrine IL-6. These processes subsequently led to the induction of GFAP. IL-6 secretion is thus thought to be a key event in controlling the astrocytic differentiation of C6 cells.Astrocyte differentiation occurs largely during the postnatal period (1), and the timing of the differentiation is regulated by various extracellular cues and cell-intrinsic programs (2). The extracellular stimuli known to promote astrocyte differentiation are fetal calf serum (3), bone morphogenic protein 2 (4), a neuropeptide, pituitary adenylate cyclase-activating polypeptide (5), and the interleukin-6 (IL-6) 1 family of cytokines (6). The IL-6 family of cytokines, oncostatin M and cardiotrophin-1, are known to activate a downstream transcription factor, signal transducer and activator of transcription 3 (STAT3), and then the activated STAT3 binds to the promoter region of the glial fibrillary acidic protein (GFAP) gene, an astrocyte marker, thereby inducing the expression of GFAP (7,8). STAT3 belongs to the STAT family of transcription factors, which play crucial roles in various intracellular signaling cascades involved in proliferation and differentiation (9, 10). In response to activation of cell-surface receptors, STAT3 is phosphorylated on its Tyr 705 residue by Janus kinase (JAK), and this leads to STAT3 dimerization, subsequent nuclear translocation, and the transactivation of its target genes (11). Bone morphogenic protein 2 synergistically acts with leukemia inhibitory factor, another IL-6 family of cytokines, to induce astrocytogenesis by promoting the complex formation of respective downstream transcription factors, Smads and STAT3, which are bridged by transcriptional coactivator p300 (12).The above-mentioned reports have extensively elucidated the mechanisms of cytokine...

Activation of tumor suppressor protein PTEN and induction of apoptosis are involved in cAMP-mediated inhibition of cell number in B92 glial cells

“…3B). Thus, cAMP has been shown to alter the morphology of primary cortical precursor cells and glioma cells [13,22]. Our results suggested that cAMP acts in B92 cells to induce morphological changes within 72 h of the stimuli (Fig.…”

“…Therefore, intracellular cAMP seems to play an important role in controlling differentiation and proliferation [13,22]. However, the signaling cascade involved in the cAMP-regulated proliferation has not yet been fully elucidated.…”

Immediate early gene IEX‐1 induces astrocytic differentiation of U87‐MG human glioma cells

“…Cells expressing high level of IEX-1 were no longer flat but rather rounded with longer processes, characteristic of cells exhibiting astrocytic phenotypes. The levels of S-100 protein and GFAP, glial differ- entiation markers [Tabuchi et al, 1981;Takanaga et al, 2004], increased directly proportional to that of IEX-1. From these results, we speculated that IEX-1 might be implicated in differentiation of glioma cells.…”

“…In the present study, we have attempted to test the possibility that IEX-1 might enhance apoptosis of human glioma cells induced by therapeutic treatments, such as chemotherapeutic agents and g-ray. Unexpectedly, however, we found that U87-MG human glioma cells overexpressing IEX-1 underwent morphological changes to astrocytic phenotype and increase in glial differentiation marker proteins, S-100 [Tabuchi et al, 1981] and glial fibrillary acidic protein (GFAP) [Takanaga et al, 2004]. Elevated cAMP and the consequent activation of protein kinase A (PKA) appeared to differentiate U87-MG glioma cells to express astrocytic phenotype, as previously observed in rat C6 glioma cell line [Willingham, 1976;Yoshimura et al, 1997;Takanaga et al, 2004], which is often used as a model system of glial cell differentiation [Harris et al, 1980].…”

Cholera toxin induces malignant glioma cell differentiation via the PKA/CREB pathway

“…Cholera toxin catalyzes ADP-ribosylation of Gs protein and results in accumulation of cellular cAMP (11,12). Ample evidence indicates that cAMP-elevating stimuli such as N-substituted cAMP analogues and cAMP-increasing reagents can induce cell differentiation in gliomas (13,14). Guerrant et al (15) reported that active whole cholera toxin, but not inactive choleragenoid, produces elevation of cAMP and parallel morphological changes in CHO cells.…”