Open in a separate window However, it remains unclear how BMP4 signaling inhibits remyelination. demyelinated lesions typically contain oligodendrocyte progenitor cells (OPCs) and premyelinating OLs that have stalled in their differentiation, implicating blocked OL differentiation as a major contributing factor to remyelination failure (Chang et al., 2002; Kuhlmann et al., 2008). Although the full complement of factors that inhibit OL differentiation and remyelination in the context of MS are yet to be completely elucidated, they most likely include a variety of inhibitory signals present within the lesion environment as well as an absence of positive signals Reparixin (Fancy et al., 2011; Kotter et al., 2011; Franklin et al., 2012). Thus, blocking the action of inhibitory factors is regarded as a leading strategy to promote endogenous CNS remyelination (Franklin and Gallo, 2014). The bone morphogenetic proteins (BMPs) are a group of secreted proteins that are part of the larger transforming growth factor- (TGF-) superfamily (Chen et al., 2004) and play critical roles in neural development and gliogenesis (Bond et al., 2012; Cole et al., 2016). Of the 20 BMPs, BMP4 has a prominent role in promoting astroglial and inhibiting oligodendroglial specification (Gomes et al., 2003; Grinspan, 2015). transgenic overexpression of BMP4 Rabbit polyclonal to DPF1 led to an increase in the number of astrocytes and a Reparixin decrease in the number of oligodendrocytes in the murine CNS (Gomes et al., 2003). In the context of demyelinating disease, BMP4 mRNA is detected in human Reparixin demyelinated MS lesions (Deininger et al., 1995), and is expressed by astrocytes, microglia, and infiltrating immune cells (Harnisch et al., 2019). Astrocytes also express a high level of BMP4 in chronic lesions that have failed to remyelinate (Harnisch et al., 2019). Through using the cuprizone-induced murine model of CNS demyelination, we have previously found that BMP4 mRNA is upregulated in the mouse corpus callosum (CC) following a demyelinating insult (Cate et al., 2010). Furthermore, we demonstrated that inhibiting BMP4 signaling following cuprizone-induced CNS demyelination via infusion of its extracellular antagonist noggin resulted in more mature oligodendrocytes and more remyelinated axons Reparixin (Cate et al., 2010; Sabo et al., 2011). However, in addition to BMP4, noggin also inhibits other BMPs such as BMP2, 7, 13, and 14 (Krause et al., 2011). Due to the promiscuous inhibitory effect of noggin and the potential effects it exerted on oligodendroglia, astrocytes, and microglia, the precise influence that the inhibition of BMP4 exerts on remyelination and the cell type mediating the effect remain unclear. BMP4 signals through membrane-bound receptor complexes composed of two type I receptors and two type II receptors. While several type I receptors exist, BMP4 gets the biggest affinity for the BMP type I receptors BMPRIA (also called ALK3) and BMPRIB (also known as ALK6; Liu et al., 1995; Knaus and Sebald, 2001). In the presence of BMP4, BMPRIA and BMPRIB initiate signaling via phosphorylation of SMAD1, SMAD5, and SMAD8 (Cuny et al., 2008) and the pharmacological inhibitor LDN-193189 selectively blocks phosphorylation SMAD1/5/8 (Cuny et al., 2008). To specifically interrogate the influence that BMP4 signaling exerted on remyelination, we infused LDN-193189 into the brain following cuprizone-induced demyelination and found it significantly enhanced oligodendroglial differentiation and their subsequent remyelination following the Reparixin demyelinating insult in which LDN-193189 significantly enhanced OPC differentiation and myelination. Further, by using a tamoxifen-dependent inducible conditional knockout (KO) mouse strategy (cuprizone experiments and postnatal day 5 (P5) to P7 mice of either sex were used for experiments. C57BL/6 mice were purchased from the Animal Resource Center (Canning Vale, WA, Australia). mouse line (provided by Dr. Kaylene Young, University of Tasmania, Hobart, TAS, Australia; Rivers et al., 2008) with allele from the start of the sequence to the end of exon 2, rendering it untranscribable (Mishina et al., 1995). All animals used for this study were bred at the Core Animal Services facility of the Florey Institute of Neuroscience and Mental Health. All chemicals were obtained from Sigma-Aldrich, unless otherwise indicated. Cuprizone protocol Cuprizone-mediated demyelination was induced by feeding 8- to 10-week-old female mice (C57BL/6) powdered feed (Barastoc) containing 0.2% cuprizone (w/w: bis-cyclohexanoneoxaldihydrazone) for 5 weeks, as previously described (Cate et al., 2010; Sabo et al., 2011). Mice were then.