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Characterisation of LaNt regulation of Basement Membrane organisation in wound repair and angiogenesis.

LaNt 在伤口修复和血管生成中对基底膜组织的调节特征。

基本信息

批准号：
BB/L020513/1
负责人：
Kevin Hamill
金额：
$ 43.77万
依托单位：
University of Liverpool
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2014
资助国家：
英国
起止时间：
2014 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FL020513%2F1
关键词：
Characterisation LaNt regulation Basement Membrane

项目摘要

In this work I propose to study the LaNt family of proteins which were recently identified and which I believe are important for processes such as wound repair, blood vessel growth and the spread of tumours. Through these studies, a deeper understanding of these processes will be obtained and this, in turn, may lead to identification of new treatment approaches for conditions such as chronic or slow-healing skin ulcers and cancer.The different tissues of the body are composed of defined combinations of specialised cell types and a mixture of proteins and sugars outside the cells, termed the extracellular matrix (ECM). Some of the cell types reside in and contribute to the production of this ECM, whereas others cover the outer (epithelia) and inner (endothelia) surfaces of regions of ECM as sheets of cells. Directly beneath these cell sheets, as well as surrounding nerves and muscles, there is an organised region of ECM termed basement membrane (BM). BMs provide the anchorage point for cells and are therefore important for stress resistance and structural integrity. In addition, BMs support the different behavioural requirements of a wide range of cell types at different times, including acting as the road upon which the skin cells migrate to close wounds. A major component of all BMs is the laminin family of proteins. Laminins assemble into cross shaped molecules that associate with one another to form a network. Formation of this network has been shown to involve a small region at the very end of the short arms of the laminin cross, which is termed a LN domain. The importance of this interaction is exemplified by a number of genetic diseases where specific defects in LN domains impact the laminin network and BM organisation resulting in skin blistering, eye defects, kidney failure or muscular dystrophy. However, despite this knowledge, the ways in which laminin networks form, how network organisation changes during different cellular processes and what drives those changes is yet to be fully understood.This project will focus on the LaNts which have been demonstrated to play a role in cell attachment and migration and which my preliminary data indicate is likely to be through regulating BM formation. Like the laminins, the LaNt (Laminin N-terminus) also contain a LN domain, this suggests that they can interact directly with laminins and modify the ways in which laminin networks are organised. Importantly, there are also tissue specific differences within the laminin family and these differences are likely to mean that the impact of the LaNts is cell type specific. This may also mean that LaNts play different roles during blood vessel growth or wound repair than during normal tissue function. In order to characterise the roles of LaNts in BM formation and the impact they have on cell behaviour and tissue function, this project will pursue 3 aims.In Aim 1, I will use proteins in solution to directly assess the ability of LaNts to interact with laminins and other BM proteins and to determine their impact on network formation. In Aim 2, I will use skin and corneal epithelial cells and blood vessel endothelial cells to study the impact of changing LaNt protein levels on cell behaviour. Specifically, we will determine if the BM deposited by cells changes in response to increasing or decreasing LaNt levels and we will assess cell movement speeds, how strongly they attach and how rapidly they divide on the different substrates.In Aim 3, I will use three dimensional models of skin, eye and of blood vessel growth to study LaNt roles in these more complex tissue models.Together the data obtained from these studies will dramatically expand what is known about LaNts, about laminin network formation, about BM organisation and ultimately about wound repair, blood vessel growth and tumour progression. In the longer term, this may lead to identification of new treatment strategies or new drug targets.

在这项工作中，我建议研究最近发现的LaNt蛋白家族，我认为这对伤口修复，血管生长和肿瘤扩散等过程很重要。通过这些研究，将获得对这些过程的更深入的了解，这反过来可能会导致识别新的治疗方法，如慢性或缓慢愈合的皮肤溃疡和癌症。身体的不同组织由特定的细胞类型和细胞外的蛋白质和糖的混合物组成，称为细胞外基质（ECM）。一些细胞类型存在于该ECM中并有助于该ECM的产生，而其他细胞类型作为细胞片覆盖ECM区域的外表面（上皮细胞）和内表面（内皮细胞）。直接在这些细胞片以及周围的神经和肌肉之下，存在ECM的组织化区域，称为基底膜（BM）。BM为细胞提供锚定点，因此对于抗应力和结构完整性很重要。此外，BM在不同时间支持各种细胞类型的不同行为要求，包括充当皮肤细胞迁移到闭合伤口的道路。所有BM的主要组分是层粘连蛋白家族蛋白质。层粘连蛋白组装成十字形分子，相互结合形成网络。该网络的形成已被证明涉及层粘连蛋白交叉短臂末端的小区域，其被称为LN结构域。这种相互作用的重要性由许多遗传疾病例证，其中LN结构域中的特定缺陷影响层粘连蛋白网络和BM组织，导致皮肤起泡、眼缺陷、肾衰竭或肌营养不良。然而，尽管有这些知识，层粘连蛋白网络形成的方式，网络组织如何在不同的细胞过程中发生变化，以及是什么驱动了这些变化还没有完全理解。本项目将重点关注LaNts，这些LaNts已被证明在细胞附着和迁移中发挥作用，我的初步数据表明可能是通过调节BM形成。与层粘连蛋白一样，LaNt（层粘连蛋白N-末端）也含有LN结构域，这表明它们可以直接与层粘连蛋白相互作用并改变层粘连蛋白网络的组织方式。重要的是，在层粘连蛋白家族内也存在组织特异性差异，这些差异可能意味着LaNts的影响是细胞类型特异性的。这也可能意味着LaNts在血管生长或伤口修复过程中发挥的作用与正常组织功能不同。为了阐明LaNts在BM形成中的作用以及它们对细胞行为和组织功能的影响，本项目将追求3个目标：在目标1中，我将使用溶液中的蛋白质直接评估LaNts与层粘连蛋白和其他BM蛋白相互作用的能力，并确定它们对网络形成的影响。在目标2中，我将使用皮肤和角膜上皮细胞以及血管内皮细胞来研究改变LaNt蛋白水平对细胞行为的影响。具体来说，我们将确定细胞沉积的BM是否会随着LaNt水平的增加或减少而变化，我们将评估细胞移动速度，它们在不同基质上的附着强度和分裂速度。在目标3中，我将使用皮肤的三维模型，研究LaNt在这些更复杂的组织模型中的作用。从这些研究中获得的数据将极大地扩展我们所知道的关于LaNts、关于层粘连蛋白网络形成、关于BM组织以及最终关于伤口修复、血管生长和肿瘤进展。从长远来看，这可能会导致新的治疗策略或新的药物靶点的确定。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Alpha actinin-1 regulates cell-matrix adhesion organization in keratinocytes: consequences for skin cell motility.

DOI：
10.1038/jid.2014.505
发表时间：
2015-04
期刊：
JOURNAL OF INVESTIGATIVE DERMATOLOGY
影响因子：
6.5
作者：
Hamill, Kevin J.;Hiroyasu, Sho;Colburn, Zachary T.;Ventrella, Rosa V.;Hopkinson, Susan B.;Skalli, Omar;Jones, Jonathan C. R.
通讯作者：
Jones, Jonathan C. R.

Mass Spectrometry Reveals a-2-HS-Glycoprotein as a Key Early Extracellular Matrix Protein for Conjunctival Cells.

质谱分析揭示 a-2-HS-糖蛋白是结膜细胞早期关键的细胞外基质蛋白。