Slit/Robo通路参与调控小鼠黄体细胞凋亡

本文旨在研究Slit/Robo家族成员在小鼠卵巢组织中的表达及其功能。用real-time PCR检测Slit/Robo家族成员在小鼠卵巢中的mRNA表达丰度,用免疫组织化学方法检测Slit2/Robo1在卵巢组织中的定位,用real-time PCR和免疫组织化学方法检测Slit2/Robo1在不同时期黄体组织中表达的变化,并用Slit/Robo信号通路的阻断剂ROBO1/Fc chimera在体外研究其在小鼠黄体组织中的功能。结果显示,在Slit/Robo家族成员中,配体Slit2和受体Robo1在小鼠卵巢组织中的表达丰度最高,Slit2和Robo1表达定位于小鼠的黄体细胞。与发情前期卵巢相比,间情期卵巢组织中Slit2和Robo1的mRNA表达水平均显著上调(P 0.01, P 0.001)。与妊娠黄体相比,晚期黄体Slit2和Robo1 mRNA表达水平均显著上调。阻断Slit/Robo信号通路后,晚期黄体细胞的凋亡率显著下降(P 0.05)。以上结果提示,Slit/Robo家族成员主要表达于晚期黄体组织,并参与调控黄体细胞的凋亡过程。     

《自然－通讯》：内源性大麻素含量增高或损胎儿脑发育

人体自身会产生一种类似大麻的化学物质,称为内源性大麻素。奥地利研究人员发现,如果孕妇体内的内源性大麻素含量增高,胎儿的大脑发育可能会受到损害。奥地利维也纳医科大学的研究人员在新一期《自然－通讯》上报告说,在胎儿大脑发育过程中,蛋白Slit及其受体Robo是重要的信号分子。附着在Robo受体上的Slit可以调节神经轴突的方向控制,从而引导胎儿大脑回路的形成。     

神经轴突导向分子Robo的功能及其分子作用机制研究进展(英文)

神经轴突导向分子Robo是进化上高度保守的跨膜蛋白。Robo及其配体Slit对神经轴突导向、神经细胞迁移、肿瘤转移、血管生成、肺脏、肾脏、心脏的形态发生以及卵巢、性腺的发育等多项生命活动具有调节作用。Robo功能的实现主要通过其Ig1结构域与其配体Slit的LRR-2结构域结合,同时也通过与多种信号分子如硫酸肝素蛋白多糖(heparan sulfate proteoglycans,HSPGs)、GTP酶激活蛋白(GTPase-activating proteins,GAPs)、酪氨酸激酶Abelson等结合发挥作用。robo基因的表达受到Hox、Midline、Nkx2.9等转录因子的调节,另外,转录后水平上的选择性剪接和转录产物的转运等调控也影响Robo的功能。本文对Robo蛋白的结构与功能以及分子作用机制等研究进展进行了综述,以期为神经发育研究和神经系统疾病与癌症防治提供新思路。     

ROBO4 与缺血性脑血管病相关的研究进展

Roundabout(Robo)蛋白是神经轴突导向分子家族Slit蛋白的单次跨膜受体,属于一种神经细胞粘附分子。Robo蛋白在神经系统已被确认具有重要轴突导向功能。近年来研究发现,血管新生的内皮细胞表面只特异性地表达Robo4,且Robo4对内皮细胞迁移、病理性血管生成和血管完整性都具有调节作用。缺血性脑血管病是人类致残甚至死亡的主要疾病之一,由于短暂或持续的脑血流减少而造成脑细胞损伤,因此,恢复脑血流、促进血管再生对脑功能恢复至关重要。Robo4对血管方面的作用为我们进一步研究及了解其在血管生成中的机制提供重要依据,也为缺血性脑血管病的治疗提供新的发展方向。     

BMP9/ALK1/Endoglin信号通路参与调节血管形成与稳态维持

在发育过程中,新生血管的形成与稳态维持是机体生命活动的重要基础.而血管生成机制复杂,参与并促进血管生成的因子众多.此过程发生异常直接与血管类疾病、炎症、癌症等疾病密切相关.除了VEGF/VEGFR,Ang-Tie2,DLL4-Notch和PDGF-BB/PDGFRβ等主要调节血管形成的传导通路外,BMP9/ALK1/Endoglin通路在其中也起着重要的作用.此通路中二聚化配体BMP9作为信号分子,与受体ALK1和Endoglin结合后激活受体,进而调控下游靶基因的表达.本文旨在对此信号通路中关键节点的分子结构和作用机制进行阐述,并对未来的机制研究与药物开发进行展望.     

Wnt5a及其信号通路与血管新生的研究进展

Wnt5a是Wnt蛋白家族中的成员之一,在细胞成熟、胚胎发育等过程中发挥着重要作用。研究表明Wnt5a的表达调控及其信号通路与血管新生密切相关,并且在血管新生性相关疾病中发挥了重要作用。本文从Wnt5a与其相关信号转导通路对血管新生的影响以及分子机制等方面进行阐述和展望,旨在为以Wnt5a为靶点进行血管新生性疾病的防治提供理论依据。     

神经迁移因子在血管系统中的表达与功能

神经迁移因子是近10年来在发育神经生物学中的研究热点,主要由ephrin、neuropilin、Slit和netrin四大家族成员构成,其主要功能是吸引或排斥神经元轴突的迁移,在神经系统中发挥着重要作用。现在,越来越多的实验证据表明：神经迁移因子的作用不仅仅局限在神经系统发育过程中,在血管发生或新生血管形成中同样具有不可替代的功能。     

Wnt信号通路与皮肤创面愈合的关系

创面愈合是由炎性细胞、细胞因子等多种因素共同参与,涉及组织修复、再生、重建的一个复杂有序的病理生理过程。皮肤慢性创面的愈合仍然是临床研究的重点与热点,随着分子生物学的发展,对皮肤创面愈合机制的认识也逐渐深入。Wnt信号通路是一条由Wnt蛋白及其受体、调节蛋白等组成的高度保守的信号通路,参与细胞增殖、凋亡、分化等多种生物学过程。Wnt信号通路作为参与皮肤愈合的信号通路之一,被认为具有调控皮肤及其附属器的发育、诱导皮肤附件的形态发生、调节毛囊的周期生长、促进创面血管新生及上皮重塑等多方面的功能。因此本文试从炎性细胞、成纤维细胞、干细胞、血管新生、表皮新生与毛囊新生等方面对Wnt信号通路与皮肤创面愈合的关系作一综述。     

Wnt/β-catenin和NF-κB信号通路对肿瘤血管生成中VEGFs/VEGFRs的调控作用

血管内皮生长因子(vascular endothelial growth factors,VEGFs)及其受体(vascular endothelialgrowth factor receptors,VEGFRs)在肿瘤的发生、发展、转移等过程中发挥了重要的作用,尤其是在肿瘤血管生成方面。而对其抑制剂的研究已经成为肿瘤防治的热点和发展方向,Wnt/β-catenin和NF-κB信号通路对肿瘤血管生成中也起着重要作用。本文就这两条信号通路对肿瘤血管生成中VEGF/VEGFRs的调控作用作一综述。     

胚胎发育中血管新生的研究

在胚胎发育中,心血管系统最早发育并发挥运输氧和营养物质的功能。在原肠运动时期,中胚层细胞在相邻内胚层细胞信号的诱导下分化产生内皮细胞,从而开始形成血管系统。血管新生是形成完整血管系统的重要过程,主要包括出芽式血管新生和套叠式血管新生两种方式。出芽式血管新生最为普遍,主要包括基底膜的降解、内皮细胞的迁移和增殖、管腔形成和血管的成熟与稳定四步。由于血管新生对胚胎发育以及许多生理过程均发挥重要作用,血管新生受到多条信号通路的精密调控。该文从内皮细胞的来源、血管新生的过程及信号调控三个方面就近年来胚胎发育中血管新生的研究作简要综述。     

A Slit/miR-218/Robo regulatory loop is required during heart tube formation in zebrafish

Members of the Slit family of secreted ligands interact with Roundabout (Robo) receptors to provide guidance cues for many cell types. For example, Slit/Robo signaling elicits repulsion of axons during neural development, whereas in endothelial cells this pathway inhibits or promotes angiogenesis depending on the cellular context. Here, we show that miR-218 is intronically encoded in slit2 and slit3 and that it suppresses Robo1 and Robo2 expression. Our data indicate that miR-218 and multiple Slit/Robo signaling components are required for heart tube formation in zebrafish and that this network modulates the previously unappreciated function of Vegf signaling in this process. These findings suggest a new paradigm for microRNA-based control of ligand-receptor interactions and provide evidence for a novel signaling pathway regulating vertebrate heart tube assembly.     

Role of Slit/Robo Signaling pathway in Bone Metabolism

Slit/Robo signals were initially found to play an essential role in nerve development as axonal guidance molecules. In recent years, with in-depth study, the role of Slit/Robo in other life activities, such as tumor development, angiogenesis, cell migration, and bone homeostasis, has gradually been revealed. Bone is an organ with an active metabolism. Bone resorption and bone formation are closely related through precise spatiotemporal coordination. There is much evidence that slit, as a new bone coupling factor, can regulate bone formation and resorption. For example, Slit3 can promote bone formation and inhibit bone resorption through Robo receptors, which has excellent therapeutic potential in metabolic bone diseases. Although the conclusions of some studies are contradictory, they all affirm the vital role of Slit/Robo signaling in regulating bone metabolism. This paper reviews the research progress of Slit/Robo signaling in bone metabolism, briefly discusses the contradictions in the existing research, and puts forward the research direction of Slit/Robo in the field of bone metabolism in the future.     

Activation of endothelial roundabout receptor 4 reduces the severity of virus-induced keratitis

Antiangiogenic molecules exert a feedback control to restrain pathological angiogenesis, which includes physical binding or inhibition of angiogenic signaling in blood vessel endothelial cells. The latter is the case in which Slit2 ligand-dependent activation of the blood vessel endothelial cell receptor roundabout 4 (Robo4) occurs. In this study, we demonstrate that Robo4 receptors are upregulated following HSV infection of the eye on the majority of the new blood vessel endothelial cells that occur in the corneal stroma. However, expression levels of the ligand for Robo4 receptors, Slit2, was not significantly increased during the disease process, and the knockdown of Slit2 gene expression using lentiviral short hairpin RNAs had no effect on the extent of pathological angiogenesis. In contrast, providing additional Slit2 protein by subconjunctival administration resulted in significantly reduced angiogenesis. The Slit2 binding to Robo4 was shown to block the downstream vascular endothelial growth factor signaling molecules Arf 6 and Rac 1 and reduce the antiapoptotic molecule Bcl-xL in blood vessel endothelial cells. Our results indicate that augmenting the host Robo4/Slit2 system could provide a useful therapeutic approach to control pathological angiogenesis associated with HSV induced stromal keratitis.     

The axonal repellent Slit2 inhibits pericyte migration: potential implications in angiogenesis

The Slit family of secreted proteins acts through the Roundabout (Robo) receptors to repel axonal migration during central nervous system development. Emerging evidence shows that Slit/Robo interactions also play a role in angiogenesis. The effect of Robo signaling on endothelial cells has been shown to be context-dependent. However, the role of Slit/Robo in pericytes has been largely unexplored. The aim of this study was to determine the effect of Slit2 on primary human pericytes and to address the underlying mechanisms, including the receptors potentially implicated. We demonstrate that both Robo1 and Robo4 are expressed by human pericytes. In the presence of their ligand Slit2, spontaneous and PDGF-induced migration of pericytes was impaired. This antimigratory activity of Slit-2 correlated with the inhibition of actin-based protrusive structures. Interestingly, human pericyte interaction with immobilized Slit2 was inhibited in the presence of anti-Robo1 and anti-Robo4 blocking antibodies, suggesting the implication of both receptors. These results add new insights into the role of Slit proteins during the angiogenic process that relies on the directional migration not only of endothelial cells but also of pericytes.     

Slit2 Inactivates GSK3β to Signal Neurite Outgrowth Inhibition

Slit molecules comprise one of the four canonical families of axon guidance cues that steer the growth cone in the developing nervous system. Apart from their role in axon pathfinding, emerging lines of evidence suggest that a wide range of cellular processes are regulated by Slit, ranging from branch formation and fasciculation during neurite outgrowth to tumor progression and to angiogenesis. However, the molecular and cellular mechanisms downstream of Slit remain largely unknown, in part, because of a lack of a readily manipulatable system that produces easily identifiable traits in response to Slit. The present study demonstrates the feasibility of using the cell line CAD as an assay system to dissect the signaling pathways triggered by Slit. Here, we show that CAD cells express receptors for Slit (Robo1 and Robo2) and that CAD cells respond to nanomolar concentrations of Slit2 by markedly decelerating the rate of process extension. Using this system, we reveal that Slit2 inactivates GSK3β and that inhibition of GSK3β is required for Slit2 to inhibit process outgrowth. Furthermore, we show that Slit2 induces GSK3β phosphorylation and inhibits neurite outgrowth in adult dorsal root ganglion neurons, validating Slit2 signaling in primary neurons. Given that CAD cells can be conveniently manipulated using standard molecular biological methods and that the process extension phenotype regulated by Slit2 can be readily traced and quantified, the use of a cell line CAD will facilitate the identification of downstream effectors and elucidation of signaling cascade triggered by Slit.     

Robo4 is a vascular-specific receptor that inhibits endothelial migration

Guidance and patterning of axons are orchestrated by cell-surface receptors and ligands that provide directional cues. Interactions between the Robo receptor and Slit ligand families of proteins initiate signaling cascades that repel axonal outgrowth. Although the vascular and nervous systems grow as parallel networks, the mechanisms by which the vascular endothelial cells are guided to their appropriate positions remain obscure. We have identified a putative Robo homologue, Robo4, based on its differential expression in mutant mice with defects in vascular sprouting. In contrast to known neuronal Robo family members, the arrangement of the extracellular domains of Robo4 diverges significantly from that of all other Robo family members. Moreover, Robo4 is specifically expressed in the vascular endothelium during murine embryonic development. We show that Robo4 binds Slit and inhibits cellular migration in a heterologous expression system, analogous to the role of known Robo receptors in the nervous system. Immunoprecipitation studies indicate that Robo4 binds to Mena, a known effector of Robo-Slit signaling. Finally, we show that Robo4 is the only Robo family member expressed in primary endothelial cells and that application of Slit inhibits their migration. These data demonstrate that Robo4 is a bona fide member of the Robo family and may provide a repulsive cue to migrating endothelial cells during vascular development.     

Cleaved Slit directs embryonic muscles

The formation of functional musculoskeletal system relies on proper connectivity between muscles and their corresponding tendon cells. In Drosophila, larval muscles are born during early embryonic stages, and elongate toward tendons that are embedded within the ectoderm in later. The Slit/Robo signaling pathway had been implicated in the process of muscle elongation toward tendons. Here we discuss our recent findings regarding the critical contribution of Slit cleavage for immobilization and stabilization of the Slit signal on the tendon cells. Slit cleavage produces 2 polypeptides, the N-terminal Slit-N, which is extremely stable, undergoes oligomerization, and associates with the tendon cell surfaces, and the C-terminal Slit-C, which rapidly degrades. Slit cleavage leads to immobilization of Slit signaling on tendons, leading to a short-range repulsion, which eventually arrest further muscle elongation. Robo2, which is co-expressed with Slit by the tendon cells facilitates Slit cleavage. This activity does not require the cytoplasmic signaling domain of Robo2. We suggest that Robo2-dependent Slit cleavage, and the formation of Slit-N oligomers on the tendon cell surfaces direct muscle elongation, and provide a stop signal for the approaching muscle, through binding to Robo and Robo3 receptors expressed by the muscles.     

Cleaved Slit directs embryonic muscles

The formation of functional musculoskeletal system relies on proper connectivity between muscles and their corresponding tendon cells. In Drosophila, larval muscles are born during early embryonic stages, and elongate toward tendons that are embedded within the ectoderm in later. The Slit/Robo signaling pathway had been implicated in the process of muscle elongation toward tendons. Here we discuss our recent findings regarding the critical contribution of Slit cleavage for immobilization and stabilization of the Slit signal on the tendon cells. Slit cleavage produces 2 polypeptides, the N-terminal Slit-N, which is extremely stable, undergoes oligomerization, and associates with the tendon cell surfaces, and the C-terminal Slit-C, which rapidly degrades. Slit cleavage leads to immobilization of Slit signaling on tendons, leading to a short-range repulsion, which eventually arrest further muscle elongation. Robo2, which is co-expressed with Slit by the tendon cells facilitates Slit cleavage. This activity does not require the cytoplasmic signaling domain of Robo2. We suggest that Robo2-dependent Slit cleavage, and the formation of Slit-N oligomers on the tendon cell surfaces direct muscle elongation, and provide a stop signal for the approaching muscle, through binding to Robo and Robo3 receptors expressed by the muscles.     

Slit-Robo signaling mediates lymphangiogenesis and promotes tumor lymphatic metastasis

The Slit family of guidance cues binds to Roundabout (Robo) receptors to modulate neuronal, leukocytic, and endothelial migration. Slit-Robo signaling had been reported to function as chemoattractive signal for vascular endothelial cells during angiogenesis. In this study, we found that Robo1 was expressed in lymphatic endothelial cells to mediate the migration and tube formation of these cells upon Slit2 stimulation, which were specifically inhibited by the function-blocking antibody R5 to Slit2/Robo1 interaction. To further explore the lymphangiogenic effect and significance mediated by Slit-Robo signaling, we intercrossed Slit2 transgenic mice with a non-metastatic RIP1-Tag2 mouse tumor model, and found that transgenic overexpression of Slit2 significantly enhanced tumor lymphangiogenesis and subsequently promoted mesenteric lymph node metastasis of pancreatic islet tumors. Taken together, our findings reveal that through interacting with Robo1, Slit2 is a novel and potent lymphangiogenic factor and contributes to tumor lymphatic metastasis.