Fgf9 signaling regulates inner ear morphogenesis through epithelial-mesenchymal interactions

The mammalian inner ear comprises the cochleovestibular labyrinth, derived from the ectodermal otic placode, and the encasing bony labyrinth of the temporal bone. Epithelial-mesenchymal interactions are thought to control inner ear development, but the modes and the molecules involved are largely unresolved. We show here that, during the precartilage and cartilage stages, Fgf9 is expressed in specific nonsensory domains of the otic epithelium and its receptors, Fgfr1(IIIc) and Fgfr2(IIIc), widely in the surrounding mesenchyme. To address the role of Fgf9 signaling, we analyzed the inner ears of mice homozygous for Fgf9 null alleles. Fgf9 inactivation leads to a hypoplastic vestibular component of the otic capsule and to the absence of the epithelial semicircular ducts. Reduced proliferation of the prechondrogenic mesenchyme was found to underlie capsular hypoplasticity. Semicircular duct development is blocked at the initial stages, since fusion plates do not form. Our results show that the mesenchyme directs fusion plate formation and they give direct evidence for the existence of reciprocal epithelial-mesenchymal interactions in the developing inner ear. In addition to the vestibule, in the cochlea, Fgf9 mutation caused defects in the interactions between the Reissner's membrane and the mesenchymal cells, leading to a malformed scala vestibuli. Together, these data show that Fgf9 signaling is required for inner ear morphogenesis.     

Notch signaling during cell fate determination in the inner ear

In the inner ear, Notch signaling has been proposed to specify the sensory regions, as well as regulate the differentiation of hair cells and supporting cell within those regions. In addition, Notch plays an important role in otic neurogenesis, by determining which cells differentiate as neurons, sensory cells and non-sensory cells. Here, I review the evidence for the complex and myriad roles Notch participates in during inner ear development. A particular challenge for those studying ear development and Notch is to decipher how activation of a single pathway can lead to different outcomes within the ear, which may include changes in the intrinsic properties of the cell, Notch modulation, and potential non-canonical pathways.     

Forced activation of Wnt signaling alters morphogenesis and sensory organ identity in the chicken inner ear

Components of the Wnt signaling pathway are expressed in the developing inner ear. To explore their role in ear patterning, we used retroviral gene transfer to force the expression of an activated form of beta-catenin that should constitutively activate targets of the canonical Wnt signaling pathway. At embryonic day 9 (E9) and beyond, morphological defects were apparent in the otic capsule and the membranous labyrinth, including ectopic and fused sensory patches. Most notably, the basilar papilla, an auditory organ, contained infected sensory patches with a vestibular phenotype. Vestibular identity was based on: (1) stereociliary bundle morphology; (2) spacing of hair cells and supporting cells; (3) the presence of otoliths; (4) immunolabeling indicative of vestibular supporting cells; and (5) expression of Msx1, a marker of certain vestibular sensory organs. Retrovirus-mediated misexpression of Wnt3a also gave rise to ectopic vestibular patches in the cochlear duct. In situ hybridization revealed that genes for three Frizzled receptors, c-Fz1, c-Fz7, and c-Fz10, are expressed in and adjacent to sensory primordia, while Wnt4 is expressed in adjacent, nonsensory regions of the cochlear duct. We hypothesize that Wnt/beta-catenin signaling specifies otic epithelium as macular and helps to define and maintain sensory/nonsensory boundaries in the cochlear duct.     

Canonical Wnt signaling negatively regulates branching morphogenesis of the lung and lacrimal gland

Key gene families such as FGFs and BMPs are important mediators of branching morphogenesis. To understand whether Wnt genes, and in particular, the canonical Wnt signaling pathway also function in the branching process, we have used a combination of experimental and genetic gain and loss of function approaches to perturb the levels of canonical Wnt signaling in two arborized structures, the lung and the lacrimal gland. Here, we show that the addition of Wnt3a conditioned medium or LiCl strongly represses growth and proliferation of the lung and lacrimal gland, a result that was confirmed in vivo using a dominant stable mutation of beta-catenin conditionally expressed in the lacrimal gland epithelium. In agreement with these data, knockdown of Wnt signaling with beta-catenin morpholinos results in a greater number of branches and increased cell proliferation. In addition, we show that canonical Wnt signaling is able to modulate the levels of Fgf10 and suppress BMP-induced proliferation in the lacrimal gland. Thus, canonical Wnt signaling negatively regulates branching morphogenesis providing a balance to FGFs and BMPs which positively regulate this process. This multilayered control of growth and proliferation ensures that branched structures attain the morphology required to function efficiently.     

A regulatory role of Wnt signaling pathway in the hematopoietic differentiation of murine embryonic stem cells

One of the most important issues in stem cell research is to understand the regulatory mechanisms responsible for their differentiation. An extensive understanding of mechanism underlying the process of differentiation is crucial in order to prompt stem cells to perform a particular function after differentiation. To elucidate the molecular mechanisms responsible for the hematopoietic differentiation of embryonic stem cells (ESCs), we investigated murine ES cells for the presence of hematopoietic lineage markers as well as Wnt signaling pathway during treatments with different cytokines alone or in combination with another. Here we report that Wnt/beta-catenin signaling is down-regulated in hematopoietic differentiation of murine ES cells. We also found that differentiation induced by the interleukin-3, interleukin-6, and erythropoietin combinations resulted in high expression of CD3e, CD11b, CD45R/B220, Ly-6G, and TER-119 in differentiated ES cells. A high expression of beta-catenin was observed in two undifferentiated ES cell lines. Gene and protein expression analysis revealed that the members downstream of Wnt in this signaling pathway including beta-catenin, GSK-3beta, Axin, and TCF4 were significantly down-regulated as ES cells differentiated into hematopoietic progenitors. Our results show that the Wnt/beta-catenin signaling pathway plays a role in the hematopoietic differentiation of murine ESCs and also may support beta-catenin as a crucial factor in the maintenance of ES cells in their undifferentiated state.     

BMP/SMAD signaling regulates the cell behaviors that drive the initial dorsal-specific regional morphogenesis of the otocyst

During development of the otocyst, regional morphogenesis establishes a dorsal vestibular chamber and a ventral auditory chamber, which collectively constitute the membranous labyrinth of the inner ear. We identified the earliest morphogenetic event heralding the formation of the vestibular chamber, a rapid thinning and expansion of the dorsolateral wall of the otocyst, and showed that this process is generated by changes in otocyst cell shape from columnar to squamous, as opposed to changes in other cell behaviors, such as localized changes in cell proliferation or cell death. Moreover, we showed that thinning and expansion of the dorsolateral otocyst is regulated by BMP/SMAD signaling, which is both sufficient and necessary for localized thinning and expansion. Finally, we showed that BMP/SMAD signaling causes fragmentation of E-cadherin in the dorsolateral otocyst, occurring concomitantly with cell shape change, suggesting that BMP/SMAD signaling regulates cell-cell adhesion during the initial morphogenesis of the otocyst epithelium. Collectively, our results show that BMP signaling via SMADs regulates the cell behaviors that drive the initial dorsal-specific morphogenesis of the otocyst, providing new information about how regional morphogenesis of a complex organ rudiment, the developing membranous labyrinth, is initiated.     

Nuclear beta-catenin-dependent Wnt8 signaling in vegetal cells of the early sea urchin embryo regulates gastrulation and differentiation of endoderm and mesodermal cell lineages

The entry of beta-catenin into vegetal cell nuclei beginning at the 16-cell stage is one of the earliest known molecular asymmetries seen along the animal-vegetal axis in the sea urchin embryo. Nuclear beta-catenin activates a vegetal signaling cascade that mediates micromere specification and specification of the endomesoderm in the remaining cells of the vegetal half of the embryo. Only a few potential target genes of nuclear beta-catenin have been functionally analyzed in the sea urchin embryo. Here, we show that SpWnt8, a Wnt8 homolog from Strongylocentrotus purpuratus, is zygotically activated specifically in 16-cell-stage micromeres in a nuclear beta-catenin-dependent manner, and its expression remains restricted to the micromeres until the 60-cell stage. At the late 60-cell stage nuclear beta-catenin-dependent SpWnt8 expression expands to the veg2 cell tier. SpWnt8 is the only signaling molecule thus far identified with expression localized to the 16-60-cell stage micromeres and the veg2 tier. Overexpression of SpWnt8 by mRNA microinjection produced embryos with multiple invagination sites and showed that, consistent with its localization, SpWnt8 is a strong inducer of endoderm. Blocking SpWnt8 function using SpWnt8 morpholino antisense oligonucleotides produced embryos that formed micromeres that could transmit the early endomesoderm-inducing signal, but these cells failed to differentiate as primary mesenchyme cells. SpWnt8-morpholino embryos also did not form endoderm, or secondary mesenchyme-derived pigment and muscle cells, indicating a role for SpWnt8 in gastrulation and in the differentiation of endomesodermal lineages. These results establish SpWnt8 as a critical component of the endomesoderm regulatory network in the sea urchin embryo.     

A blockade in Wnt signaling is activated following the differentiation of F9 teratocarcinoma cells

Aberrant activation of the Wnt signaling pathway is a common event in human tumor progression. Wnt signaling has also been implicated in maintaining a variety of adult and embryonic stem cells by imposing a restraint to differentiation. To understand the effect of Wnt signaling on the differentiation of epithelial cells, we used mouse teratocarcinoma F9 cells as a model. The F9 cells can be differentiated into visceral endoderm (VE) resembling absorptive columnar epithelial cells. We performed comparative gene expression analysis on retinoic acid-differentiated and undifferentiated F9 cells and confirmed that markers of VE and intestinal epithelium were induced upon differentiation. The induction of these markers by retinoic acid was reduced in the presence of Wnt, although Wnt alone did not change their expression. This suggests that Wnt signaling inhibited the differentiation of F9 cells by altering gene expression. This inhibition was also reflected in the morphology of the F9 cells as their apical-basal polarity was disrupted by inclusion of Wnt during differentiation. These results support a model in which Wnt modulates the expression of genes required for normal terminal differentiation of the stem cells. However, it follows that progenitor cells must escape from Wnt signaling to attain the differentiated state. Accordingly, we found that differentiated F9 cells no longer responded to Wnt and that a blockade in Wnt signaling occurred upstream of Axin. Consistent with this, Wnt negative regulators, such as Dickkopf-1 and Disabled-2, were induced upon the differentiation of F9 cells. We propose that a similar system to produce Wnt inhibitors regulates homeostasis of certain stem cell compartments in vivo.     

Wnt信号通路在毛细胞分化和再生过程中的作用

Wnt信号通路在生物发育和维持内环境稳态过程中起着重要作用。Wnt配体通过与Frizzle受体结合参与体轴的形成、细胞分化和细胞命运决定等生命活动。在小鼠内耳发育过程中,Wnt信号通路扮演了重要角色：在内耳发育早期阶段,参与听基板的特化和听泡的形成;在内耳发育后期阶段,调控毛细胞分化及毛细胞纤毛束的定向。本文综述了Wnt信号通路在内耳毛细胞发育分化及再生过程中的研究进展,以期为从事相关领域的科研人员提供参考。     

FGF信号通路在内耳发育调控和毛细胞再生中的作用

内耳是感受听觉和平衡觉的复杂器官。在内耳发育过程中,成纤维生长因子(fibroblast growth factor, FGF)信号通路参与了听基板的诱导、螺旋神经节(statoacoustic ganglion, SAG)的发育以及Corti器感觉上皮的分化。FGF信号开启了内耳早期发育的基因调控网络,诱导前基板区域以及听基板的形成。正常表达的FGF信号分子可促进听囊腹侧成神经细胞的特化,但成熟SAG神经元释放的过量FGF5可抑制此过程,形成负反馈环路使SAG在稳定状态下发育。FGF20在Notch信号通路的调控下参与了前感觉上皮区域向毛细胞和支持细胞的分化过程,而内毛细胞分泌的FGF8可调控局部支持细胞分化为柱细胞。人类FGF信号通路异常可导致多种耳聋相关遗传病。此外,FGF信号通路在低等脊椎动物毛细胞自发再生以及干细胞向内耳毛细胞诱导过程中都起到了关键作用。本文综述了FGF信号通路在内耳发育调控以及毛细胞再生中的作用及其相关研究进展,以期为毛细胞再生中FGF信号通路调控机制的阐明奠定理论基础。     

Wnt信号通路参与外周免疫调节的研究进展

Wnt信号通路最初是由于其在动物胚胎发育和形态发生过程中的作用而引起了人们的注意。过去二十多年来,人们又发现Wnt通路参与干细胞的分化及多种疾病的发生,这使它成为研究的一个热点。近年来的研究表明,Wnt通路与免疫系统也有密切的联系,不仅参与各种免疫细胞的发育分化,还能调控外周免疫细胞的功能。该文就对Wnt信号通路在外周免疫系统中的研究进展作一综述。