Treatment of oligodendrocytes with antisense deoxyoligonucleotide directed against CREB mRNA: Effect on the cyclic AMP‐dependent induction of myelin basic protein expression

Abstract

We have shown previously that in oligodendrocytes, the transcription factor cyclic AMP response element binding protein (CREB) is maximally expressed immediately prior to the most rapid period of myelination in rat brain. We have begun to investigate the role of this protein during myelination by downregulating CREB synthesis in cultured oligodendrocytes using an antisense deoxyoligonucleotide directed against CREB mRNA. Neonatal oligodendrocytes were grown for 4 days in a chemically defined medium (CDM) after which intracellular delivery of CREB antisense oligonucleotide was facilitated by using a liposome preparation. Control cultures were treated in a similar manner but in the presence of CREB sense oligomer. Immediately after transfection, cells were cultured for 3 days in CDM in the presence or absence of the cyclic AMP (cAMP) analogue N6, O21-dibutyryl cAMP (db-cAMP). In these cultures, myelin basic protein (MBP) expression was investigated by immunocytochemistry and Western blot analysis. Treatment of control cultures with db-cAMP resulted in a significant increase in the number of MBP positive cells which was abolished when the cells were treated with CREB antisense oligonucleotide. MBP positive cells in control cultures treated with db-cAMP have extended and highly branched MBP positive processes. In contrast, MBP positive cells in either control cultures grown in the absence of db-cAMP or cultures grown in the presence of db-cAMP but treated with CREB antisense oligonucleotide showed shorter and less complex processes and the MBP immunoreactivity appeared to be concentrated in the cell body. These observations suggest that CREB is at least one of the mediators in the induction of oligodendrocyte differentiation by cAMP.

Neurotrophin‐3 and a CREB‐mediated signaling pathway regulate Bcl‐2 expression in oligodendrocyte progenitor cells

“…1999). In support of this hypothesis, we found that in committed OLGs, CREB plays a crucial role in the cyclic AMP‐mediated stimulation of myelin basic protein (MBP) expression (Sato‐Bigbee and DeVries 1996; Afshari et al . 2001).…”

“…We have previously used this technique to successfully block CREB expression at different stages of OLG development (Sato‐Bigbee and DeVries 1996; Johnson et al . 2000; Afshari et al .…”

Relationship between cell cycle molecules and onset of oligodendrocyte differentiation

“…The possible involvement of a coactivator has also been proposed in the case of cAMP. Several studies from Sato-Bigbee et al have shown convincingly that CREB plays an important role in the events induced by cAMP and leading to differentiation (Sato-Bigbee, 1996;. The long latency between start of treatment and detection of the effect, however, and the absence of a canonical CREB binding site on the MBP promoter, argue in favor of an indirect model of regulation.…”

“…Invariably, however, there is an obligate relationship between proliferation and differentiation, because cell cycle exit is a prerequisite for oligodendrocyte differentiation. Among the extracellular factors promoting differentiation of oligodendrocyte progenitors are: transforming growth factor ␤ (TGF-␤) (McKinnon et al, 1993), cyclic-AMP (McMorris, 1985;Sato-Bigbee and DeVries, 1996;Ye et al, 1992;Sato-Bigbee and Yu, 1993;Zhang and Miskimins, 1993;Ghiani et al, 1999b), ␤-adrenergic receptor agonists (Ghiani et al, 1999a) and thyroid hormone (Barres et al, 1994;Ibarrola et al, 1996;Ahlgren et al, 1997). In vitro, for instance, TGF ␤ has been shown to induce oligodendrocyte differentiation and inhibit progenitor proliferation (McKinnon et al, 1993).…”

Molecular mechanisms involved in the actions of apotransferrin upon the central nervous system: Role of the cytoskeleton and of second messengers

“…Oligodendroglial cell differentiation is stimulated by cAMP followed by activation of protein kinase A (PKA; Mc Morris, 1983; Pleasure et al, 1986; Raible and Mc Morris, 1990). Developing OLGc express elevated levels of the transcription factor CREB, which mediates cAMP‐dependent stimulation of MBP expression and the outgrowth of cell processes in developing OLGc (Sato‐Bigbee et al, 1994; Sato‐Bigbee and De Vries, 1996). Protein kinase activity may affect oligodendrocyte differentiation (Vartanian et al, 1986; Althaus et al, 1991; Bambrick and Braun, 1991).…”