Identification and functional characterization of thromboxane A₂ receptors in Schwann cells

Abstract

Previous reports have demonstrated the presence of functional thromboxane A 2 (TP) receptors in astrocytes and oligodendrocytes. In these experiments, the presence and function of TP receptors in primary rat Schwann cells (rSC) and a neuro®brosarcoma-derived human Schwann cell line (T265) was investigated. Immunocytochemical and immunoblot analyses using polyclonal anti-TP receptor antibodies demonstrate that both cell types express TP receptors. Treatment with the stable thromboxane A 2 mimetic U46619 (10 mM) did not stimulate intracellular calcium mobilization in rSC, whereas T265 cells demonstrated a calcium response that was inhibited by prior treatment with TP receptor antagonists. U46619 also stimulated CREB phosphorylation on Ser133 in T265 cells and, to a lesser extent, in rSC. To identify potential mechanisms of CREB phosphorylation in rSC, we monitored intracellular cAMP levels following U46619 stimulation. Elevated levels of cAMP were detected in both rSC (20-fold) and T265 (15-fold) cells. These results demonstrate that TP receptor activation speci®cally stimulates CREB phosphorylation in T265 cells, possibly by a calciumand/or cAMP-dependent mechanism. In contrast, TP receptor activation in rSC stimulates increases in cAMP and CREB phosphorylation but does not elicit changes in intracellular calcium.

A Novel Nuclear Signaling Pathway for Thromboxane A2 Receptors in Oligodendrocytes: Evidence for Signaling Compartmentalization during Differentiation

“…Studies from our laboratory have previously characterized the localization and functional effects of TPR activation in both OLGs and Schwann cells (5,3,25,30,41). These results demonstrated that stimulation of TPRs promotes proliferation of OPCs and prolongs survival of mature OLGs (25).…”

Transcriptional profiling in an MPNST-derived cell line and normal human Schwann cells

“…The T265 cell line is widely used for cell culture studies of Schwann cells in MPNSTs (Muja et al, 2001;Badache and De Vries, 1998;Muir et al, 2001;Miller et al, 2003;Fieber et al, 2003). This cell line has characteristics consistent with Schwann cells that have been transformed into a malignant phenotype because of the loss of neurofibromin in the auto-somaldominant condition NF1; however, many genes are thought to be regulated abnormally in NF1 and in this cell line, which makes it difficult to design properly controlled experiments for studies of NF1 in vitro.…”

Thromboxane A2Promotes Interleukin-6 Biosynthesis Mediated by an Activation of Cyclic AMP-Response Element-Binding Protein in 1321N1 Human Astrocytoma Cells

“…4D), suggesting the involvements of both PKA and p38 MAPK/MSK pathways. Although the mechanism remains unclear, cyclic AMP elevation and PKA activation have been shown in response to TP agonist (Muja et al, 2001;Miggin and Kinsella, 2002). On the other hand, p38 MAPK activation by TXA 2 has been reported in platelets and isolated rat mesenteric resistance arteries (Bolla et al, 2002;Minuz et al, 2002).…”

“…IL-6 gene expression is regulated by multiple transcription factors, including CREB, activator protein-1 (AP-1), nuclear factor-B (NF-B), CCAAT enhancerbinding protein-␤, and glucocorticoid receptor in complex ways (Obara and Nakahata, 2002). Because CREB activation by U46619 has been reported (Muja et al, 2001), we determined whether CREB is involved in the U46619-induced IL-6 gene expression. CRE activity was measured by reporter gene assay.…”

“…TP-mediated extracellular signal-regulated kinase (ERK) activation in differentiated 1321N1 cells depends on the activation of phosphatidylcholine-specific phospholipase C (Kobayashi et al, 2000). It has also been shown that TXA 2 analog elevates cyclic AMP concentration in several cell types, such as Schwann cells (Muja et al, 2001), even though there is currently no evidence for the interaction between TPs and G␣ s . In that report, cyclic AMP-response element-binding protein (CREB) is activated, which is assumed to be by a cyclic AMP-dependent mechanism.…”