Shh and Fgf8 act synergistically to drive cartilage outgrowth during cranial development

Much of the skeleton and connective tissue of the vertebrate head is derived from cranial neural crest. During development, cranial neural crest cells migrate from the dorsal neural tube to populate the forming face and pharyngeal arches. Fgf8 and Shh, signaling molecules known to be important for craniofacial development, are expressed in distinct domains in the developing face. Specifically, in chick embryos these molecules are expressed in adjacent but non-overlapping patterns in the epithelium covering crest-derived mesenchyme that will give rise to the skeletal projections of the upper and lower beaks. It has been suggested that these molecules play important roles in patterning the developing face. Here, we directly examine the ability of FGF8 and SHH signaling, singly and in combination, to regulate cranial skeletogenesis, both in vitro and in vivo. We find that SHH and FGF8 have strong synergistic effects on chondrogenesis in vitro and are sufficient to promote outgrowth and chondrogenesis in vivo, suggesting a very specific role for these molecules in producing the elongated beak structures during chick facial development.     

FGF8 signaling is chemotactic for cardiac neural crest cells

Cardiac neural crest cells migrate into the pharyngeal arches where they support development of the pharyngeal arch arteries. The pharyngeal endoderm and ectoderm both express high levels of FGF8. We hypothesized that FGF8 is chemotactic for cardiac crest cells. To begin testing this hypothesis, cardiac crest was explanted for migration assays under various conditions. Cardiac neural crest cells migrated more in response to FGF8. Single cell tracing indicated that this was not due to proliferation and subsequent transwell assays showed that the cells migrate toward an FGF8 source. The migratory response was mediated by FGF receptors (FGFR) 1 and 3 and MAPK/ERK intracellular signaling. To test whether FGF8 is chemokinetic and/or chemotactic in vivo, dominant negative FGFR1 was electroporated into the premigratory cardiac neural crest. Cells expressing the dominant negative receptor migrated slower than normal cardiac neural crest cells and were prone to remain in the vicinity of the neural tube and die. Treating with the FGFR1 inhibitor, SU5402 or an FGFR3 function-blocking antibody also slowed neural crest migration. FGF8 over-signaling enhanced neural crest migration. Neural crest cells migrated to an FGF8-soaked bead placed dorsal to the pharynx. Finally, an FGF8 producing plasmid was electroporated into an ectopic site in the ventral pharyngeal endoderm. The FGF8 producing cells attracted a thick layer of mesenchymal cells. DiI labeling of the neural crest as well as quail-to-chick neural crest chimeras showed that neural crest cells migrated to and around the ectopic site of FGF8 expression. These results showing that FGF8 is chemotactic and chemokinetic for cardiac neural crest adds another dimension to understanding the relationship of FGF8 and cardiac neural crest in cardiovascular defects.     

成纤维细胞生长因子8(FGF8)研究进展

成纤维细胞生长因子8(fibroblast growth factor 8,FGF8)是成纤维细胞生长因子(FGFs)家族的成员之一。其在人胚胎时期多种组织内进行表达,对各种器官的形成中起着重要的作用。在正常成人体内,FGF8的表达水平受到严格的限制,然而在某些癌细胞或炎症部位中大量表达,特别是在激素类癌症的发生和发展中起着重要的作用。因此应用FGF8抗体治疗激素类癌症,为临床提供了新的治疗途径。     

Fibroblast growth factor 2 regulates dopaminergic neuron development in vivo

Fibroblast growth factor 2 (FGF-2) is a neurotrophic factor participating in regulation of proliferation, differentiation, apoptosis and neuroprotection in the central nervous system. With regard to dopaminergic (DA) neurons of substantia nigra pars compacta (SNpc), which degenerate in Parkinson's disease, FGF-2 improves survival of mature DA neurons in vivo and regulates expansion of DA progenitors in vitro. To address the physiological role of FGF-2 in SNpc development, embryonic (E14.5), newborn (P0) and juvenile (P28) FGF-2-deficient mice were investigated. Stereological quantification of DA neurons identified normal numbers in the ventral tegmental area, whereas the SNpc of FGF-2-deficient mice displayed a 35% increase of DA neurons at P0 and P28, but not at earlier stage E14.5. Examination of DA marker gene expression by quantitative RT-PCR and in situ hybridization revealed a normal patterning of embryonic ventral mesencephalon. However, an increase of proliferating Lmx1a DA progenitors in the subventricular zone of the ventral mesencephalon of FGF-2-deficient embryos indicated altered cell cycle progression of neuronal progenitors. Increased levels of nuclear FgfR1 in E14.5 FGF-2-deficient mice suggest alterations of integrative nuclear FgfR1 signaling (INFS). In summary, FGF-2 restricts SNpc DA neurogenesis in vivo during late stages of embryonic development.     

Loss of floor plate Netrin-1 impairs midline crossing of corticospinal axons and leads to mirror movements

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How does Fgf signaling from the isthmic organizer induce midbrain and cerebellum development?

The mesencephalic/rhombomere 1 border (isthmus) is an organizing center for early development of midbrain and cerebellum. In this review, we summarize recent progress in studies of Fgf signaling in the isthmus and discuss how the isthmus instructs the differentiation of the midbrain versus cerebellum. Fgf8 is shown to play a pivotal role in isthmic organizer activity. Only a strong Fgf signal mediated by Fgf8b activates the Ras-extracellular signal-regulated kinase (ERK) pathway, and this is sufficient to induce cerebellar development. A lower level of signaling transduced by Fgf8a, Fgf17 and Fgf18 induce midbrain development. Numerous feedback loops then maintain appropriate mesencephalon/rhombomere1 and organizer gene expression.     

诱导人脐血间充质干细胞分化为多巴胺能神经元样细胞的研究

目的：探讨人羊膜上皮细胞条件培养液(ACM)及SHH,FGF8诱导人脐血间充质干细胞（CB-MSCs）分化为多巴胺( dopamine,DA )能神经元样细胞的作用及机制。方法：利用沉降红细胞、密度梯度离心和贴壁筛选法纯化CB-MSCs,加入ACM及SHH,FGF8分为CON,ACM,SHH+FGF8,ACM+SHH+FGF8四组诱导48h后,免疫细胞化学染色鉴定分化细胞的DA神经元表型。应用高压液相色谱技术测定细胞培养上清中DA含量,并应用Realtime PCR方法观察DA能神经元发育过程中的相关基因En-1,Foxa2,Lmx1b,Gli-1,Pitx3,Nurr1以及Ngn2的表达。结果：加诱导剂诱导48h后,各诱导组的TH和DAT的表达量均比对照组高。早期的转录因子En1,Foxa2 在各组中均有表达;而Pitx3,Lmx1b表达在各诱导组中;Lmx1b在ACM+SHH+FGF8组中表达最高;Pitx3在各诱导组中均有表达且无明显差异。Nurr1,Ngn2在SHH+FGF8组中表达很强。结论：ACM及SHH,FGF8可诱导脐血间充质干细胞分化为DA能神经元样细胞,其作用机制与多巴胺能神经元发育过程中的各种基因En-1,Foxa2,Lmx1b,Gli-1,Pitx3,Nurr1以及Ngn2相关。     

Role of Pax-5 in the regulation of a mid-hindbrain organizer's activity

The mes-metencephalic boundary (isthmus) has been suggested to act as an organizer in the development of the optic tectum. Pax-5 was cloned as a candidate for regulator of the organizing center. Isthmus-specific expression of Pax-5 and analogy with the genetic cascade in Drosophila suggest that Pax-5 may be at a higher hierarchical position in the gene regulation cascade of tectum development. To examine this possibility, a gain-of-function experiment on Pax-5 was carried out. In ovo electroporation on E2 chick brain with the eucaryotic expression vector that encodes chick Pax-5 cDNA was used. Not only was a considerable amount of Pax-5 expressed ectopically in the transfected brain, but irregular bulging of the neuroepithelium was induced in the diencephalon and mesencephalon. At Pax-5 misexpressing sites, uptake of BrdU was increased. Histological examination of E7 transfected brain revealed that Pax-5 caused transdifferentiation of diencephalon into the tectum-like structure. In the bulges of the E7 mesencephalon, differentiation of laminar structure was repressed when compared to the normal side. In transfected embryos, En-2, Wnt-1 and Fgf8 were up-regulated ectopically, and Otx2 was down-regulated in the diencephalon to mesencephalon. Moreover, Ephrin-A2, which is expressed specifically in the tectum with a gradient highest at the caudal end, is suggested to be involved in pathfinding of the retinal fibers, and was induced in the bulges. When the mouse Fgf8 expression vector was electroporated, Pax-5 and chick Fgf8 were also induced ectopically. These results suggest that Pax-5, together with Fgf8, hold a higher position in the genetic hierarchy of the isthmus organizing center and regulate its activity.     

FGF signaling enhances a sonic hedgehog negative feedback loop at the initiation of spinal cord ventral patterning

A prevalent developmental mechanism for the assignment of cell identities is the production of spatiotemporal concentration gradients of extracellular signaling molecules that are interpreted by the responding cells. One of such signaling systems is the Shh gradient that controls neuronal subtype identity in the ventral spinal cord. Using loss and gain of function approaches in chick and mouse embryos, we show here that the fibroblast growth factor (FGF) signaling pathway is required to restrict the domains of ventral gene expression as neuroepithelial cells become exposed to Shh during caudal extension of the embryo. FGF signaling activates the expression of the Shh receptor and negative pathway regulator Patched 2 (Ptch2) and therefore can enhance a negative feedback loop that restrains the activity of the pathway. Thus, we identify one of the mechanisms by which FGF signaling acts as a modulator of the onset of Shh signaling activity in the context of coordination of ventral patterning and caudal axis extension. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 956–971, 2016     

Patterning through differential endoreduplication in epithelial organogenesis of the chordate,Oikopleura dioica

The contributions that control of cell proliferation and cell growth make to developmental regulation of organ and body size remain poorly explored, particularly with respect to endocycles in polyploid tissues. The epithelium of the marine chordate Oikopleura dioica is composed of a fixed number of cells grouped in territories according to gene-specific expression and nuclear sizes and shapes. As the animal grows 10-fold during the life cycle, epithelial cells increase in size differentially as a function of their spatial position. We show that this cellular pattern reflected differences in ploidy levels ranging from 34 to 1,300 C. The diverse ploidy levels in defined cellular fields resulted both from different timing of entry into endocycles and from cell-specific regulation of endocycle lengths. Throughout the life cycle, differential cell size and ploidy increases were accompanied by field-specific profiles of progressive reductions in G-phase duration. Endocycles were asynchronous among cells of a given epithelial territory, but at the resolution of individual cells, both DNA replication timing and ploidy levels were bilaterally symmetric. The transparent, accessible, oikoplastic epithelium is a model of choice for the study of endoreduplication in the context of pattern formation and growth.     

EGFR-dependent suppression of synaptic autophagy is required for neuronal circuit development

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