Molecular Mechanisms of Arterial-Venous Differentiation

动静脉分化的分子机制

• 批准号: 8645705
• 负责人: Saulius Sumanas
• 金额: $ 37.49万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8645705
• 关键词: Adult; Affect; Angioblast; Arteries; Biological Models; Blood Circulation; Blood Vessels; CD44 Antigens; Cells; Data; Defect; Development; Diabetic Retinopathy; Disease; Embryo; Endothelial Cells; Exhibits; Genes; Hemorrhage; Hyaluronan; Hyaluronic Acid; Image; Knowledge; Lead; Location; Maps; Modeling; Molecular; Notch Signaling Pathway; Pathway interactions; Pattern; Phosphorylation; Play; Process; Role; Signal Transduction; Signaling Molecule; Staging; Stem cells; Stroke; System; Testing; Time; Transgenic Organisms; Translating; Vascular Diseases; Vascular Endothelial Growth Factors; Veins; Venous; Venous Malformation; Vertebrates; Wound Healing; Zebrafish; angiogenesis; base; cell motility; experience; hyaluronan synthase 1; loss of function; malformation; migration; morphogens; notch protein; novel; overexpression; prevent; progenitor; public health relevance; receptor; tissue repair; transcription factor; tumor; tumorigenesis; vasculogenesis

DESCRIPTION (provided by applicant): Arterial and venous (A-V) patterning is critical for the establishment of functional embryonic and adult vasculature. Proper A-V differentiation is critical for the formation of functional vessels during tissue repair processes such as wound healing and is associated with a number of pathological conditions including A-V malformations and tumor induced angiogenesis. Therefore understanding the mechanisms of A-V differentiation will lead to new treatments in tumorigenesis, wound repair and multiple vascular disorders. While it is difficult to study A-V patterning in mammalian systems, transparent zebrafish embryos are easily accessible for observation and experimental manipulations. During formation of the major vessels, vascular endothelial progenitor cells (EPCs) assume arterial or venous identity prior to the initiation of circulation. However, it is not understood how EPCs choose among the arterial or venous fates if they are all exposed to the same signaling molecules such as Vegf and Hh. We have found that the expression timing of an evolutionarily conserved master regulator of vasculogenesis Etsrp / Etv2 is one of the critical factors in A-V differentiation. Furthermore, arterial and venous progenitors may originate at different spatial locations, the inner and the outer lines and thus experience different Vegf concentrations. We hypothesize that the arterial- venous fate of EPCs is determined by the combination of etsrp-dependent timing of Vegf receptor flk1 and hyaluronan (HA) receptor stabilin 2 (stab2) expresion and their spatial location within Vegf gradient. To demonstrate that the timing of etsrp expression affects A-V differentiation, photoactivatable morpholinos will be used to inhibit etsrp function at different developmental stages in zebrafish embryos. Fate mapping and time- lapse imaging will be performed to characterize the cell movements and to determine the arterial-venous fates of the inner and outer lines of EPCs. To determine if Vegf and Hh gradients play a role in the activation of Notch signaling and arterial differentiation within the inner line angioblasts, overexpression and loss of function approaches will be used to inhibit Vegf, Hh and Notch signaling combined with lineage tracing to determine the A-V cell fates. To determine if Etsrp downstream target Stab2 functions as a receptor for HA and induces activation of Notch signaling during A-V differentiation, A-V defects in Stab2 and HA synthase Has2 knockdown embryos will be investigated. It will be analyzed if HA-Stab2 signaling leads to Tyr-phosphorylation of Stab2 and ERK phosphorylation, resulting in Notch pathway activation and arterial marker expression. Upon completion of this study, we will have identified the molecular mechanism how the timing of etsrp expression and Vegf gradient lead to the differential expression of arterial and venous genes. The acquired knowledge will be critical in our understanding of molecular mechanisms of A-V differentiation and will have an impact on developing treatments for multiple diseases and pathological conditions related to vasculature formation which include A-V malformations, diabetic retinopathy, wound healing and tumorigenesis.!

描述（由申请人提供）：动脉和静脉（A-V）图案对于建立功能性胚胎和成人脉管系统至关重要。适当的A-V分化对于在组织修复过程（例如伤口愈合）中形成功能血管至关重要，并且与许多病理条件有关，包括A-V畸形和肿瘤诱导的血管生成。因此，了解A-V分化的机制将导致肿瘤发生，伤口修复和多种血管疾病的新治疗方法。 尽管很难在哺乳动物系统中研究A-V模式，但透明的斑马鱼胚胎很容易访问进行观察和实验操作。在主要血管形成期间，血管内皮祖细胞（EPC）在循环启动之前假设动脉或静脉认同。但是，如果EPC都暴露于相同的信号分子（例如VEGF和HH），则尚不了解EPC在动脉命运中的选择。我们发现，进化保守的血管生成主要调节剂ETSRP / ETV2的表达时间是A-V分化的关键因素之一。此外，动脉和静脉祖细胞可能起源于不同的空间位置，内部和外线，因此经历了不同的VEGF浓度。我们假设EPC的动脉静脉命运是由VEGF受体FLK1和透明质酸（HA）受体稳定蛋白2（Stab2）Explision及其在VEGF梯度内的空间位置组合的ETSRP依赖性计时确定的。为了证明ETSRP表达的时机会影响A-V分化，可光活化的形态学将用于抑制斑马鱼胚胎不同发育阶段的ETSRP功能。将执行命运映射和时间流失成像，以表征细胞运动并确定EPC的内部和外部线的动脉命运。为了确定VEGF和HH梯度是否在内线血管生成细胞内的Notch信号传导和动脉分化的激活中发挥作用，将使用过表达和功能方法的丧失来抑制VEGF，HH和Notch信号与谱系信号结合与谱系相结合以确定A-V细胞命运。为了确定eTSRP下游目标stab2是否充当HA的受体并在A-V分化过程中诱导Notch信号的激活，将研究Stab2和HA合酶HAS2敲低胚胎中的A-V缺陷。如果HA-STAB2信号传导导致Stab2和ERK磷酸化的Tyr磷酸化，从而导致Notch途径激活和动脉标记物表达，则将分析。 这项研究完成后，我们将确定ETSRP表达和VEGF梯度的时机如何导致动脉和静脉基因的差异表达。获得的知识将在我们对A-V分化的分子机制的理解中至关重要，并将影响与脉管形成有关的多种疾病和病理状况的治疗，包括A-V畸形，糖尿病性视网膜病变，伤口愈合和肿瘤。