关于Akt信号通路在淋巴管发育过程中的功能和相关分子机制的研究

论文摘要

淋巴管系统在维持哺乳动物体液平衡和转运大分子或免疫细胞的过程中扮演着非常重要的角色。从淋巴管内皮细胞发育成为成熟的带有正常平滑肌细胞覆盖和瓣膜的淋巴管系统是一个非常复杂的过程,受到各种不同基因的调控。近几十年来,借助于转基因和基因敲除技术,淋巴管相关的研究也取得了长足的发展,很多基因以及相关的信号通路都被发现在淋巴管发育过程中起作用。丝/苏氨酸蛋白激酶Akt（也被称之为蛋白激酶B, PKB）被认为是生物体内最重要的信号通路之一,参与调控很多细胞功能,例如细胞存活、细胞迁移、细胞分化和增殖等。在哺乳动物体内,哺乳动物体内存在Akt1、Akt2和Akt3,这三者由不同的基因编码,但在结构上却非常相似。之前的报道指出Akt能够介导生长因子的刺激信号,对血管内皮细胞的功能有很重要的影响,并且在斑马鱼和鸡的血管发育过程中Akt也起到重要的作用。然而与此同时,关于Akt在淋巴管发育方面的作用没有报道。在本研究课题中,我们选择了Akt基因敲除的小鼠作为分析的对象。首先我们发现在单个Akt基因敲除小鼠体内,淋巴管发育都能进行。但是,Akt1敲除小鼠的皮肤里的毛细淋巴管明显比野生型小鼠要细,而Akt2或Akt3敲除小鼠的毛细淋巴管却和野生型小鼠类似。进一步的研究发现,Akt1敲除小鼠皮肤毛细淋巴管的淋巴管内皮细胞数明显要少于野生型小鼠。其次,Akt1敲除小鼠的淋巴管瓣膜形成也有缺陷。在Akt1敲除小鼠皮肤中,初级的收集淋巴管缺少瓣膜,但在皮肤深层的主收集淋巴管中发现有瓣膜的存在。对淋巴管功能进行分析发现,Akt1敲除小鼠的淋巴管的吸收和回流作用相比于野生型小鼠显得比较弱。同时,Akt1敲除小鼠的收集淋巴管在某些区域有异常的扩张,并且我们也发现Akt1敲除小鼠中收集淋巴管的平滑肌细胞覆盖明显比野生型小鼠要少,而且排列也很松散而且杂乱。最后我们发现如果用表达VEGF-C的重组腺病毒去感染Akt1基因敲除小鼠,能够和在野生型小鼠中一样诱导大量的淋巴管新生。此外,在双Akt基因敲除的小鼠中（Akt1-/-,Akt3+/-）也有淋巴管的发育,但存在类似于Akt1基因敲除小鼠的淋巴管发育缺陷。综合这些信息表明,Akt在淋巴管发育过程中是具有重要作用的,但三个Akt基因在淋巴管发育的过程中有代偿互补的作用,而相比于其它两个Akt基因（Akt2,Akt3）,Akt1在淋巴管发育中是最为重要的信号介导因子。

论文目录

• Abstract
• 中文摘要
• Abbreviations
• Chapter 1:Review: Molecular regulation of vascular development
• 1. Blood and Lymphatic Vascular System
• 1.1 The Blood Vascular System
• 1.1.1 Vasculogenesis and Angiogenesis
• 1.1.2 Arterial-Venous Specification
• 1.2 The Lymphatic Vascular System
• 1.2.1 LECs Budding from Cardinal Vein
• 1.2.2 Primary Lymph Sac Formation
• 1.2.3 Lymphatic-Blood Vessel Separation
• 1.2.4 Lymphatic Vessel Maturation
• 2. Molecular Regulation of Vascular Development
• 2.1 Vascular endothelial growth factors and receptors
• 2.1.1 Vascular endothelial growth factors
• 2.1.1.1 VEGF-A
• 2.1.1.2 VEGF-C
• 2.1.2 Vascular endothelial growth factor receptors
• 2.1.2.1 VEGFR-1 and VEGFR-2
• 2.1.2.2 VEGFR-3
• 2.1.2.3 Neuropilins,co-receptors for VEGFRs
• 2.2 Angiopoietins and Ties
• 2.3 Notch signaling
• 2.4 Other molecules regulating vascular development
• 2.5 Molecular identity of blood and lymphatic endothelial calls
• 3. Akt signaling pathway in vascular development
• 3.1 Three Akt isoforms
• 3.2 Akt is a signal transducer for PI3K
• 3.3 Akt related mice models and their phenotypes
• 3.4 Akt in vascular system
• 3.4.1 Akt in endothelial cell functions
• 3.4.2 Akt in vascular development
• References
• Chapter 2:Dissection of Akt signaling pathway inlymphatic vascular development
• Introduction
• Material and Methods
• Results
• -/- mice showed thinner lymphatic capillariesas a result of decreased LECs number'>Section 2.1 Akt1^-/- mice showed thinner lymphatic capillariesas a result of decreased LECs number
• -/- mice'>2.1.1 Decrease of lymphatic capillary size in Akt1^-/-mice
• -/- null mice'>2.1.2 Developmental angiogenesis not altered in Akt1^-/- null mice
• -/- lymphatics'>2.1.3 Decreased LECs number in Akt1^-/-lymphatics
• -/- mice'>2.1.4 LECs proliferation and apoptosis not altered in Akt1^-/-mice
• -/- mice'>2.1.5 No obvious change of eNOS phosphorylation in Akt1^-/-mice
• 2.1.6 Conclusion and brief discussion
• -/- mice'>Section 2.2 Defective lymphatic vascular maturation in Akt1^-/-mice
• -/- mice'>2.2.1 Defective lymphatic valve formation in Akt1^-/-mice
• 2.2.2 Dilated collecting lymphatic vessels in Akt1 deficient mice
• -/- mice'>2.2.3 Abnormal vSMC coverage with collecting lymphatics in Akt1^-/-mice
• 2.2.4 Conclusion and brief discussion
• Section 2.3 Compensatory effects between Akt isoforms
• 2.3.1 Akt acts as downstream effecter for VEGF-C/VEGFR-3 signaling
• -/- mice'>2.3.2 Induction of lymphangiogenesis by VEGF-C in Akt1^-/-mice
• 2.3.3 Detection of lymphatic vessels in Akt compound knockout mice
• 2.3.4 Conclusion and brief discussion
• Discussion
• References
• Original Publication
• Acknowledgements
• 致谢