Paraxis is required for somite morphogenesis and differentiation in Xenopus laevis

Background : In most vertebrates, the segmentation of the paraxial mesoderm involves the formation of metameric units called somites through a mesenchymal‐epithelial transition. However, this process is different in Xenopus laevis because it does not form an epithelial somite. Xenopus somitogenesis is characterized by a complex cells rearrangement that requires the coordinated regulation of cell shape, adhesion, and motility. The molecular mechanisms that control these cell behaviors underlying somite formation are little known. Although the Paraxis has been implicated in the epithelialization of somite in chick and mouse, its role in Xenopus somite morphogenesis has not been determined. Results : Using a morpholino and hormone‐inducible construction approaches, we showed that both gain and loss of function of paraxis affect somite elongation, rotation and alignment, causing a severe disorganization of somitic tissue. We further found that depletion or overexpression of paraxis in the somite led to the downregulation or upregulation, respectively, of cell adhesion expression markers. Finally, we demonstrated that paraxis is necessary for the proper expression of myotomal and sclerotomal differentiation markers. Conclusions : Our results demonstrate that paraxis regulates the cell rearrangements that take place during the somitogenesis of Xenopus by regulating cell adhesion. Furthermore, paraxis is also required for somite differentiation. Developmental Dynamics 244:973–987, 2015. © 2015 Wiley Periodicals, Inc.     

BMP regulation of myogenesis in zebrafish

In amniotes, BMP signaling from lateral plate and dorsal neural tube inhibits differentiation of muscle precursors in the dermomyotome. Here, we show that BMPs are expressed adjacent to the dermomyotome during and after segmentation in zebrafish. In addition, downstream BMP pathway members are expressed within the somite during dermomyotome development. We also show that zebrafish dermomyotome is responsive to BMP throughout its development. Ectopic overexpression of Bmp2b increases expression of the muscle precursor marker pax3, and changes the time course of myoD expression. At later stages, overexpression increases the number of Pax7+ myogenic precursors, and delays muscle differentiation, as indicated by decreased numbers of MEF2+ nuclei, decreased number of multi‐nucleated muscle fibers, and an increased myotome angle. In addition, we show that while BMP overexpression is sufficient to delay myogenic differentiation, inhibition of BMP does not detectably affect this process, suggesting that other factors redundantly inhibit myogenic differentiation. Developmental Dynamics 239:806–817, 2010. © 2010 Wiley‐Liss, Inc.     

Extrinsic versus intrinsic cues in avian paraxial mesoderm patterning and differentiation.

Somitic and head mesoderm contribute to cartilage and bone and deliver the entire skeletal musculature. Studies on avian somite patterning and cell differentiation led to the view that these processes depend solely on cues from surrounding tissues. However, evidence is accumulating that some developmental decisions depend on information within the somitic tissue itself. Moreover, recent studies established that head and somitic mesoderm, though delivering the same tissue types, are set up to follow their own, distinct developmental programmes. With a particular focus on the chicken embryo, we review the current understanding of how extrinsic signalling, operating in a framework of intrinsically regulated constraints, controls paraxial mesoderm patterning and cell differentiation.     

Noncyclic Notch activity in the presomitic mesoderm demonstrates uncoupling of somite compartmentalization and boundary formation

To test the significance of cyclic Notch activity for somite formation in mice, we analyzed embryos expressing activated Notch (NICD) throughout the presomitic mesoderm (PSM). Embryos expressing NICD formed up to 18 somites. Expression in the PSM of Hes7, Lfng, and Spry2 was no longer cyclic, whereas Axin2 was expressed dynamically. NICD expression led to caudalization of somites, and loss of Notch activity to their rostralization. Thus, segmentation and anterior-posterior somite patterning can be uncoupled, differential Notch signaling is not required to form segment borders, and Notch is unlikely to be the pacemaker of the segmentation clock.     

四孔螺钉肋骨接骨板治疗多发性肋骨骨折的力学分析

目的分析自主设计开发的新型四孔螺钉肋骨接骨板在治疗多发性肋骨骨折中的力学特性并验证其临床疗效。方法使用有限元方法,分析四孔螺钉肋骨接骨板固定后断骨的最大位移与骨折断面间隙,并与目前临床常用的六爪肋骨接骨板的固定效果进行对比,最后回顾临床数据验证其治疗多发性肋骨骨折的临床疗效。结果有限元分析结果表明,使用四孔螺钉肋骨接骨板固定后,断骨的最大位移降低,断面间隙缩小,固定效果优于六爪肋骨接骨板。但螺钉孔处存在应力集中,可能会损伤皮质骨。回顾分析复旦大学附属浦东医院30例使用四孔螺钉肋骨接骨板治疗多发性肋骨骨折的临床资料,术后患者胸痛症状明显减轻,呼吸恢复顺畅,术后CT显示骨折复位良好,验证了四孔螺钉肋骨接骨板的临床疗效。结论四孔螺钉肋骨接骨板对断骨固定良好,且对于大曲率肋骨的骨折情形适用性强,适用于治疗多发性肋骨骨折。     

Xenopus Rnd1 and Rnd3 GTP‐binding proteins are expressed under the control of segmentation clock and required for somite formation

The process of segmentation in vertebrates is described by a clock and wavefront model consisting of a Notch signal and an fibroblast growth factor‐8 (FGF8) gradient, respectively. To further investigate the segmentation process, we screened gene expression profiles for downstream targets of the segmentation clock. The Rnd1 and Rnd3 GTP‐binding proteins comprise a subgroup of the Rho GTPase family that show a specific expression pattern similar to the Notch signal component ESR5, suggesting an association between Rnd1/3 and the segmentation clock. Rnd1/3 expression patterns are disrupted by overexpression of dominant‐negative or active forms of Notch signaling genes, and responds to the FGF inhibitor SU5402 by a posterior shift analogous to other segmentation‐related genes, suggesting that Rnd1/3 expressions are regulated by the segmentation clock machinery. We also show that antisense morpholino oligonucleotides to Rnd1/3 inhibit somite segmentation and differentiation in Xenopus embryos. These results suggest that Rnd1/3 are required for Xenopus somitogenesis. Developmental Dynamics 238:2867–2876, 2009. © 2009 Wiley‐Liss, Inc.     

Molecular evolution and expression of zebrafish St8SiaIII, an alpha-2,8-sialyltransferase involved in myotome development.

Enzymes of the St8Sia family, a subgroup of the glycosyltransferases, mediate the transfer of sialic acid to glycoproteins or glycolipids. Here, we describe the cloning of the zebrafish St8SiaIII gene and study its developmental activity. A conserved synteny relationship among vertebrate chromosome regions containing St8SiaIII loci underscores an ancient duplication of this gene in the teleost fish lineage and a specific secondary loss of one paralog in the zebrafish. The single zebrafish St8SiaIII enzyme, which is expected to function as an oligosialyltransferase, lacks maternal activity, is weakly expressed during nervous system development, and shows a highly dynamic expression pattern in somites and somite-derived structures. Morpholino knock-down of St8SiaIII leads to anomalous somite morphologies, including defects in segment boundary formation and myotendious-junction integrity. These phenotypes hint for a basic activity of zebrafish St8SiaIII during segmentation and somite formation, providing novel evidence for a non-neuronal function of sialyltransferases during vertebrate development.     

斑马鱼类胰岛素生长因子信号途径及作用机制研究进展

斑马鱼是研究早期发育中类胰岛素生长因子（insulin-like growth factors, IGF）信号途径的模式生物,斑马鱼IGF信号系统主要包括IGF配体、受体、结合蛋白（IGFBPs）。IGF配体包括IGF-1,IGF-2。受体为IGF-1R,该受体有两种全长结构（igf-1ra和igf-1rb）。结合蛋白包括IGFBP-1,IGFBP-2,IGFBP-3,IGFBP-5和 IGFBP-6,它们的结构特征已经阐明,从斑马鱼模式动物中获得的信息不仅可以为斑马鱼胚胎发育生物学提供新的观点,还可以进一步加深我们对一般意义上的脊椎动物的生长和发育的理解。本文就近年来斑马鱼IGF系统的进展作一综述。     

Role of fragile histidine triad protein expression in pathogenesis of malignant pleural mesothelioma

AIM:To investigate the relationship of fragile histidine triad (FHIT) and Ki-67 expression with clinicopathological variables of patients with malignant pleural mesothelioma (MPM). METHODS: Formalin-fixed, paraffin-embedded tissue sections of 30 asbestos induced MPM (epithelial and biphasic) patients were examined for FHIT and Ki-67 expression using immunohistochemical techniques and results were compared with clinicopathological variables. RESULTS: Immunohistochemical study results were as follows: 12 (40%) cases showed low FHIT expression and 18 (60%) showed high expression. There was no significant relationship between FHIT and age, gender or histological subtypes (p > 0.05). Ki-67 expression was 'low' in 13 (43.3%) cases and 'high' in 17 (56.7%) cases. No correlation could be demonstrated between Ki-67 expression and age, gender or histological subtypes (p > 0.05). No significant association was observed between FHIT and Ki-67 expression in MPM. CONCLUSION: The results support the role of FHIT as a tumour suppressor gene in asbestos induced MPM. There is no significant correlation between FHIT and cell proliferation marker expressions in malignant pleural mesothelioma. Therefore, it can be concluded that loss of FHIT does not interfere with tumour proliferation. This can be accepted as evidence for an early role of FHIT loss in carcinogenesis; however, it needs to be strengthened by further studies.     

BMP-4 and Noggin signaling modulate dorsal fin and somite development in the axolotl trunk.

BMP-4, a member of the TGF-beta superfamily of growth factors, is involved in various developmental processes. We investigated the effects of BMP-4 and its antagonist Noggin on axolotl trunk development. Implantation of BMP-4-coated microbeads caused inhibition of muscle and dorsal fin formation in the vicinity of the microbeads. At some distance, myotomes developed with reduced height but increased width, which was accompanied by increased cell proliferation. These effects could be modulated by co-implanting Noggin-coated beads. Immunostaining of Pax7 further revealed that although the dermomyotome was absent in the vicinity of BMP-4-coated beads, at some distance from them, it was thicker than in controls, indicating that moderate amounts of BMP-4 stimulate this layer of undifferentiated cells. In contrast, Noggin generally inhibited the dermomyotome, possibly indicating premature differentiation of dermomyotome cells. We conclude that BMP-4 and Noggin are involved in the regulation of cell proliferation and differentiation during somite development.     

痰液和胸腹水沉积物切片在肿瘤脱落细胞学中的作用

目的　探讨痰和胸腹水沉积物切片在肿瘤细胞学诊断中的意义。方法　对 183例痰液和 2 5 4例胸腹水沉积物进行涂片和石蜡包埋连续切片 ,部分病例做了特染、免疫组化检查。结果　在组织学证实的 96例肺癌中 ,痰液涂片加连续切片的联合检查阳性者 92例 ,阳性率 95 83% ,假阴性 4例。联合检查、切片和涂片三者间差异有显著性 (P <0 0 1)。在组织学证实的 16 2例胸腹腔癌瘤所致的胸腹水沉积物连续切片 ,全部为阳性 ,而在涂片中阳性为 135例 ,两者之间差异有显著性 (P <0 0 1)。结论　痰液和胸腹水沉积物切片避免了单一涂片取材的局限性 ,切片上细胞相对集中 ,易于观察肿瘤的组织形态 ,可增加肿瘤脱落细胞学的诊断价值。     

The Role of Sdf‐1α signaling in Xenopus laevis somite morphogenesis

Background: Stromal derived factor‐1α (sdf‐1α), a chemoattractant chemokine, plays a major role in tumor growth, angiogenesis, metastasis, and in embryogenesis. The sdf‐1α signaling pathway has also been shown to be important for somite rotation in zebrafish (Hollway et al., 2007). Given the known similarities and differences between zebrafish and Xenopus laevis somitogenesis, we sought to determine whether the role of sdf‐1α is conserved in Xenopus laevis. Results: Using a morpholino approach, we demonstrate that knockdown of sdf‐1α or its receptor, cxcr4, leads to a significant disruption in somite rotation and myotome alignment. We further show that depletion of sdf‐1α or cxcr4 leads to the near absence of β‐dystroglycan and laminin expression at the intersomitic boundaries. Finally, knockdown of sdf‐1α decreases the level of activated RhoA, a small GTPase known to regulate cell shape and movement. Conclusion: Our results show that sdf‐1α signaling regulates somite cell migration, rotation, and myotome alignment by directly or indirectly regulating dystroglycan expression and RhoA activation. These findings support the conservation of sdf‐1α signaling in vertebrate somite morphogenesis; however, the precise mechanism by which this signaling pathway influences somite morphogenesis is different between the fish and the frog. Developmental Dynamics 243:509–526, 2014. © 2013 Wiley Periodicals, Inc.     

The role of the notochord in vertebral column formation

The backbone or vertebral column is the defining feature of vertebrates and is clearly metameric. Given that vertebrae arise from segmented paraxial mesoderm in the embryo, this metamerism is not surprising. Fate mapping studies in a variety of species have shown that ventromedial sclerotome cells of the differentiated somite contribute to the developing vertebrae and ribs. Nevertheless, extensive studies in amniote embryos have produced conflicting data on exactly how embryonic segments relate to those of the adult. To date, much attention has focused on the derivatives of the somites, while relatively little is known about the contribution of other tissues to the formation of the vertebral column. In particular, while it is clear that signals from the notochord induce and maintain proliferation of the sclerotome, and later promote chondrogenesis, the role of the notochord in vertebral segmentation has been largely overlooked. Here, we review the established role of the notochord in vertebral development, and suggest an additional role for the notochord in the segmental patterning of the vertebral column.     

Gateway compatible vectors for analysis of gene function in the zebrafish.

The recent establishment of recombination-based cloning systems has greatly facilitated the analysis of gene function by allowing rapid and high-efficiency generation of plasmid constructs. However, the use of such an approach in zebrafish requires the availability of recombination-compatible plasmids that are appropriate for functional studies in zebrafish embryos. In this work, we describe the construction and validation of Gateway compatible vectors based on commonly used zebrafish plasmids. We have generated pCS-based plasmids that allow rapid generation of both N-terminal and C-terminal fusion proteins, and we demonstrate that mRNA synthesized from these plasmids encodes functional native or fusion proteins in injected zebrafish embryos. In parallel, we have established similar Gateway plasmids containing Tol2 cis elements that promote efficient integration into the zebrafish genome and allow expression of native or fusion proteins in a tissue-specific manner in the zebrafish embryo. Finally, we demonstrate the use of this system to rapidly identify tissue-specific cis elements to aid the establishment of blood vessel-specific transgenic constructs. Taken together, this work provides an important platform for the rapid functional analyses of open reading frames in zebrafish embryos.     

Temporal and spatial patterning of axial myotome fibers in Xenopus laevis

An image of an X. laevis tadpole showing rhodamine‐labeled transplanted presomitic mesoderm cells (red) in which a subset of cells have differentiated into myotome fibers (shown in orange). The differentiated myotome fibers are stained with the antibody 12/101 (green) and the notochord is stained with Tor 70 (blue). The image was digitally duplicated twice and fused at the anterior end. From Krneta‐Stankic et al., Developmental Dynamics 239:1162–1177, 2010.     

Temporal and spatial patterning of axial myotome fibers in Xenopus laevis

Somites give rise to the vertebral column and segmented musculature of adult vertebrates. The cell movements that position cells within somites along the anteroposterior and dorsoventral axes are not well understood. Using a fate mapping approach, we show that at the onset of Xenopus laevis gastrulation, mesoderm cells undergo distinct cell movements to form myotome fibers positioned in discrete locations within somites and along the anteroposterior axis. We show that the distribution of presomitic cells along the anteroposterior axis is influenced by convergent and extension movements of the notochord. Heterochronic and heterotopic transplantations between presomitic gastrula and early tail bud stages show that these cells are interchangeable and can form myotome fibers in locations determined by the host embryo. However, additional transplantation experiments revealed differences in the competency of presomitic cells to form myotome fibers, suggesting that maturation within the tail bud presomitic mesoderm is required for myotome fiber differentiation. Developmental Dynamics 239:1162–1177, 2010.© 2010 Wiley‐Liss, Inc.     

Characterization of the laminin gene family and evolution in zebrafish

Laminins are essential components of all basement membranes and are fundamental to tissue development and homeostasis. Humans possess at least 16 different heterotrimeric laminin complexes formed through different combinations of alpha, beta, and gamma chains. Individual chains appear to exhibit unique expression patterns, leading to the notion that overlap between expression domains governs the constitution of complexes found within particular tissues. However, the spatial and temporal expression of laminin genes has not been comprehensively analyzed in any vertebrate model to date. Here, we describe the tissue‐specific expression patterns of all laminin genes in the zebrafish, throughout embryonic development and into the “post‐juvenile” period, which is representative of the adult body form. In addition, we present phylogenetic and microsynteny analyses, which demonstrate that the majority of our zebrafish sequences are orthologous to human laminin genes. Together, these data represent a fundamental resource for the study of vertebrate laminins. Developmental Dynamics 240:422–431, 2011. © 2011 Wiley‐Liss, Inc.