Bmp7 Regulates the Survival, Proliferation, and Neurogenic Properties of Neural Progenitor Cells during Corticogenesis in the Mouse

Bone morphogenetic proteins (BMPs) are considered important regulators of neural development. However, results mainly from a wide set of in vitro gain-of-function experiments are conflicting since these show that BMPs can act either as inhibitors or promoters of neurogenesis. Here, we report a specific and non-redundant role for BMP7 in cortical neurogenesis in vivo using knockout mice. Bmp7 is produced in regions adjacent to the developing cortex; the hem, meninges, and choroid plexus, and can be detected in the cerebrospinal fluid. Bmp7 deletion results in reduced cortical thickening, impaired neurogenesis, and loss of radial glia attachment to the meninges. Subsequent in vitro analyses of E14.5 cortical cells revealed that lack of Bmp7 affects neural progenitor cells, evidenced by their reduced proliferation, survival and self-renewal capacity. Addition of BMP7 was able to rescue these proliferation and survival defects. In addition, at the developmental stage E14.5 Bmp7 was also required to maintain Ngn2 expression in the subventricular zone. These data demonstrate a novel role for Bmp7 in the embryonic mouse cortex: Bmp7 nurtures radial glia cells and regulates fundamental properties of neural progenitor cells that subsequently affect Ngn2-dependent neurogenesis

Involvement of twisted gastrulation in T cell-independent plasma cell production

Bone morphogenetic protein (BMP) signaling is increasingly implicated in immune cell differentiation and function; however, direct in vivo evidence for such a role is still missing. In this article, we report that Twisted gastrulation (TWSG1), an extracellular regulator of BMP signaling, is expressed in activated B cells and regulates T-independent B cell responses in the mouse. Twsg1-deficient B cells mount stronger T-independent type 2 responses reflected as increased IgM levels and numbers of Ag-specific IgM-secreting cells. BCR stimulation of Twsg1-deficient B cells results in hyperproliferation, hyperresponsiveness, and decreased apoptosis, whereas TLR stimulation results in hyperproliferation and increased IgG3 production. These changes are reflected on the molecular level by increased transcription of Bcl-6, Pax5, and the BMP-responsive gene Id-2. The TWSG1 effects on B cells appear to be cell intrinsic, suggesting that Twsg1 expression in B cells serves to interpret BMP signals on a per-cell basis. In summary, our observations on the role of TWSG1 in B cell function is opening new paths toward the exploration of the role of BMP signaling in immunological processes

BMP7 controls radial glia survival.

<p>(A) Absence of Bmp7 affects RG and neural progenitor cells. Expression of Nestin (i, ii), RC2 (iii, iv), Sox2 (v, vi), Pax6 (vii, viii) in wt or <i>Bmp7</i>^Δ/Δ E14.5 cortices. The <i>Bmp7</i>^Δ/Δ cortex appears less organized (i–iv). The RGC make poor contact to the meninges (iii, iv). Sox2 and Pax6 expression are lost or diminished in the VZ/SVZ. (B) Counts of cell spreads of E14.5 cortices reveal a reduction of NeuN and Pax6-positive cells in <i>Bmp7</i>^Δ/Δ cortical cells (black bar) when compared to wt littermate cells (open bar). (C, D) The number of PCNA-positive cells (C) and cells having incoporated BrdU (D) are reduced in <i>Bmp7</i>^Δ/Δ cortical cells (black bar) when compared to wt littermate cells (open bar). (E) <i>Bmp7</i>^Δ/Δ cortical cells (black bar) showed increased numbers of Caspase-3-positive cells when compared to wt littermate cells, which was corrected following treatment with rBMP7. (F–H) Neurospheres derived from wt, <i>Bmp7^wt/</i>^Δ heterozygote, or <i>Bmp7</i>^Δ/Δ cortical cells (F) are fewer (G) and smaller (H) indicating that Bmp7 affects the survival and self-renewal properties of neural progenitor cells in the developing cortex. *p < 0.05 by Student's t test.</p

<i>Bmp7</i> null embryos suffer from microencephaly.

<p>(A) <i>Bmp7</i> expression as monitored by lacZ-reporting in the mouse E14.5 cortex. Strong expression is seen in the medial regions, such as hem (H) and choroid plexus (CPl), the pial membrane (Pi) and meninges (Me). No expression is apparent in the ventricular (VZ), the subventricular (SVZ), and intermediate (IZ) zones, and the cortical plate (CP) (inlay). (B) Bmp7 protein (lower panels; the staining with DAPI is shown in the upper panels) is observed in the marginal zone (M), the CP and VZ of wild-type embryos (E14.5) and is lost in the <i>Bmp7</i>-deficient mouse embryo. (C) <i>Bmp7</i>-deletion results in microencephaly with a thinner cortex and a less clearly defined cortical plate, most prominent in medial (A, A′) and lesser in more lateral regions. (D) Other Bmp proteins present in the developing neocortex are not significantly altered following <i>Bmp7</i> deletion. The anti-Bmp6 antibody used here appears to cross-react with Bmp7, resulting in a second upper band, which is lost upon <i>Bmp7</i>^Δ/Δ deletion.</p

Normal layering of the E14.5 neural cortex in <i>Bmp7</i>-deficient mouse embryos.

<p>Comparison of wild-type (wt, panels A, C, E, G, I) and <i>Bmp7</i>^Δ/Δ (panels B, D, F, H, J) cortical sections for expression of <i>Reelin</i> (A, B), presence of Map2 protein (C, D), Svet1 mRNA (E, F), and Tbr2 protein (G, H), and Pax6 mRNA (I, J), demonstrating their respective correct location even in the absence of <i>Bmp7</i>. The cortical plate (CP) may appear somewhat reduced in the Map2-stained sections (C, D).</p

Bmp7 regulates <i>Ngn2</i> steady-state transcript levels in the E14.5 cortex.

<p>Expression of the pro-neural genes <i>Ngn1</i> (A, B), <i>Ngn2</i> (C, D), <i>NeuroD</i> (E, F) in wild-type (wt; A, C, E) and <i>Bmp7</i>^Δ/Δ (B, D, F) E14.5 cortices. Whereas <i>Ngn1</i> and <i>NeuroD</i> expression appear normal, <i>Ngn2</i> is significantly reduced (transiently, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034088#pone.0034088.s003" target="_blank">Figure S3</a>) in the absence of <i>Bmp7</i>.</p

Reduced cortical plate thickness in the absence of <i>Bmp7</i>.

<p>Colocalization of Tbr1 (green) and Ctip2 (red) reveals a reduction of post-mitotic neurons in the CP marked as Tbr1/Ctip2 double positive cells (yellow). Ctip2 staining in the SVZ remained unaltered.</p