Construction of a novel Digital Scanning Lightsheet Microscope and its application in measuring 3D cell behaviour and movement in embryos

新型数字扫描光片显微镜的构建及其在测量胚胎 3D 细胞行为和运动中的应用

• 批准号: BB/G015082/1
• 负责人: Kees Weijer
• 金额: $ 80.79万
• 依托单位: University of Dundee
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2009
• 资助国家: 英国
• 起止时间: 2009 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FG015082%2F1
• 关键词: Construction; novel; Digital; Scanning; Lightsheet

Understanding embryonic development is one of the big challenges of Life Science Research The first stage in the embryonic development of all higher organisms is the formation of the zygote from the fusion of a sperm with an egg cell, which is typically followed by a series of rapid cell divisions in which many cells are generated. Due to asymmetries present in the cytoplasm of the egg, or external asymmetries provided by the mother, different cells initiate distinct gene expression programs, which allow the cells to proliferate, undergo programmed cell death, differentiate and cell move. For proper development to occur these processes have to be coordinated precisely in 3 dimensional space and time, which is achieved by extensive cell-cell communication. Cell-cell communication can involve signaling through direct cell-cell contacts (short-range) or through secretion of signaling molecules that can diffuse in the space in between the cells and travel a relatively long distance. There are only a limited number of forms and shapes that can be generated by these processes and in many cases cells will need to move from the place where they produced to the site where they are required. This is particularly important in the process of gastrulation and during the formation and wiring of the nervous system. Gastrulation is a critical stage in embryogenesis where the main body plan of the embryo is laid down and the axes of symmetry emerge. It involves large-scale long-range cell movements during which cells of the three germlayers (ectoderm, mesoderm, and endoderm) take up the correct topological positions in the embryo. The endoderm is located innermost in the embryo and adult and endoderm cells form the lining of the digestive tract and associated glands (liver pancreas etc). The endoderm is surrounded by the mesoderm that will give rise to the muscles and the skeleton. The mesoderm is covered by the outmost layer, the ectoderm that will form the epidermis and the nervous system. Improper cell movements during gastrulation results in severe cases in abortive development and in less severe cases form the basis of many congenital defects in animals and humans. The signaling mechanisms and the cellular processes underlying gastrulation have been studied in a variety of experimental model systems. The study of gastrulation in higher vertebrates such as amniotes (birds reptiles and mammals) has focused on the development of the chick and mouse embryo. The chick embryo has the advantage that development takes place outside the mother and is therefore easily accessible to experimental manipulation. The chick embryo is flat and translucent which makes observation of cell movements during gastrulation possible. To understand complex processes such as gastrulation it is essential to be able to follow the movements of all cells in the embryo. This requires very powerful microscopic techniques and one of the aims of this research is to build and develop a microscope with which this will be possible. This will require close collaboration between physicists, computer scientist and life scientists and we have assembled such a consortium. Once the instrument is build we will use it to map out cell division and movement during early chick development and generate a blueprint of this process. In a second phase we will start to investigate the signaling systems that control these movements by experimental perturbation, we will up and down regulate critical signaling molecules and study their quantitative effects on early development. From this we will build up a picture of the most critical processes that control gastrulation which will help in understanding many congenital defects and diseases in later life and knowledge obtained in these studies will be essential to be able to prevent and cure some of these cases in the future.

了解胚胎发育是生命科学研究的一大挑战所有高等生物胚胎发育的第一阶段是精子与卵细胞融合形成受精卵，随后通常会进行一系列快速细胞分裂，其中产生许多细胞。由于卵子细胞质中存在的不对称性，或母体提供的外部不对称性，不同的细胞启动不同的基因表达程序，从而使细胞增殖、经历程序性细胞死亡、分化和细胞移动。为了发生适当的发育，这些过程必须在 3 维空间和时间中精确协调，这是通过广泛的细胞间通信来实现的。细胞间通讯可以涉及通过直接细胞间接触（短程）或通过信号分子的分泌来进行信号传导，信号分子可以在细胞之间的空间中扩散并传播相对较长的距离。这些过程只能产生有限数量的形式和形状，并且在许多情况下，细胞需要从它们产生的地方移动到需要它们的地方。这在原肠胚形成过程以及神经系统的形成和连接过程中尤其重要。原肠胚形成是胚胎发生的关键阶段，胚胎的主体规划在此阶段确定并出现对称轴。它涉及大规模的长距离细胞运动，在此过程中三个胚层（外胚层、中胚层和内胚层）的细胞在胚胎中占据正确的拓扑位置。内胚层位于胚胎的最里面，成体和内胚层细胞形成消化道和相关腺体（肝胰腺等）的内壁。内胚层被中胚层包围，中胚层将产生肌肉和骨骼。中胚层被最外层覆盖，外胚层将形成表皮和神经系统。原肠胚形成过程中细胞运动不当会导致严重的发育不良，而在不太严重的情况下，则会形成动物和人类许多先天性缺陷的基础。原肠胚形成的信号传导机制和细胞过程已在各种实验模型系统中进行了研究。对羊膜动物（鸟类、爬行动物和哺乳动物）等高等脊椎动物的原肠胚形成的研究主要集中在小鸡和小鼠胚胎的发育上。鸡胚胎的优点是发育发生在母体外，因此易于进行实验操作。鸡胚是扁平且半透明的，这使得观察原肠胚形成过程中的细胞运动成为可能。为了理解原肠胚形成等复杂过程，必须能够跟踪胚胎中所有细胞的运动。这需要非常强大的显微技术，这项研究的目的之一是建造和开发一种可以实现这一目标的显微镜。这需要物理学家、计算机科学家和生命科学家之间的密切合作，我们已经组建了这样一个联盟。一旦仪器建成，我们将用它来绘制早期雏鸡发育过程中的细胞分裂和运动，并生成该过程的蓝图。在第二阶段，我们将开始研究通过实验扰动控制这些运动的信号系统，我们将上调和下调关键信号分子并研究它们对早期发育的定量影响。由此，我们将了解控制原肠胚形成的最关键过程，这将有助于了解许多先天性缺陷和晚年疾病，并且在这些研究中获得的知识对于将来能够预防和治愈其中一些病例至关重要。

项目成果

Myosin-II-mediated cell shape changes and cell intercalation contribute to primitive streak formation.

• DOI: 10.1038/ncb3138
• 发表时间: 2015-04
• 期刊: Nature cell biology
• 影响因子: 21.3
• 作者: Rozbicki E;Chuai M;Karjalainen AI;Song F;Sang HM;Martin R;Knölker HJ;MacDonald MP;Weijer CJ
• 通讯作者: Weijer CJ

Dynamic morphoskeletons in development.

发育中的动态形态骨架。

• DOI: 10.1073/pnas.1908803117
• 发表时间: 2020
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Serra M

Gaussian vs. Bessel light-sheets: performance analysis in live large sample imaging

高斯光片与贝塞尔光片：实时大样本成像中的性能分析

• DOI: 10.1117/12.2277324
• 发表时间: 2017
• 作者: Donnachie M

A 'chemotactic dipole' mechanism for large-scale vortex motion during primitive streak formation in the chick embryo

鸡胚原条形成过程中大规模涡旋运动的“趋化偶极子”机制

• DOI: 10.1088/1478-3975/8/4/045008
• 发表时间: 2011
• 期刊: Physical Biology
• 影响因子: 2
• 作者: Sandersius S