The Cellular Control of Corneal Development and Transparency and Generation of Biomimetic Corneal Tissue.

角膜发育和透明度的细胞控制以及仿生角膜组织的生成。

• 批准号: BB/M025349/1
• 负责人: Andrew Quantock
• 金额: $ 98.09万
• 依托单位: CARDIFF UNIVERSITY
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2015
• 资助国家: 英国
• 起止时间: 2015 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FM025349%2F1
• 关键词: Cellular; Control; Corneal; Development; Transparency

The cornea is the clear tissue at the front of the eye, which transmits light and focuses it sharply onto the retina. Accordingly, it is absolutely essential for vision. Composed mostly of collagen, the cornea is not unlike other collagen-rich tissues, such as tendons, cartilages, intervertebral discs, or even the sclera; the "white of the eye". But what makes the cornea optically clear is the exquisite way in which the collagen is structured.Collagen in the human body exists predominantly in the form of long fibres, which are very strong along their axes, and can be thought of as having mechanical properties rather like ropes on sailing ships or steel cables on suspension bridges. The organisation of collagen fibres in different tissues is contingent on the tissue's function, and the predominant requirement is usually mechanical. The cornea is unique in this regard because as well as the need to fulfil mechanical requirements, it needs to be transparent.The bulk of the cornea is made up of cells, known as keratocytes, and collagen. Most collagen exists in the form of very long and very thin fibres, or fibrils as they are called in the cornea because they are so thin. Remarkably, and unlike all the other collagen-rich tissues in the body, collagen fibrils in cornea are all of exactly the same diameter and are arranged into a near-perfect, hexagonal-type lattice. It is this precise structural arrangement of collagen which gives the cornea its transparency. But, how does it arise?Keratocytes in the cornea synthesise and deposit collagen. So, presumably these cells must be at least partly responsible for the way in which the collagen fibrils are laid down and arranged. We will use pioneering microscopic approaches across a wide range of magnifications, using laser and electron imaging technologies and working with scientists in the University of California, to study corneas from developing chicks in fertilised hen eggs. Our aim is to understand precisely how cells in the developing cornea interact with each other to make such a beautifully structured tissue as a transparent cornea (the basis of corneal transparency in the chick is the same as that in humans). Recently, we pioneered the use of three-dimensional volume electron microscopy for the study of cornea, and discovered that cells in the developing chick cornea all have highly extended, but thin, cell processes. This showed that the cells themselves occupy a volume of the cornea which is much larger than previously believed. Based on this discovery, our hypothesis is that cells in the developing cornea form an extended network in which they communicate with each other, and that, as a group, they have the innate ability to synthesise incredibly thin collagen fibrils and deposit them into a precise lattice-like arrangement to meet the needs of transparency. To test this hypothesis we will study corneas from the very earliest stages of development, which have never before been examined in three dimensions at such high magnifications. New methodologies for the mathematical modelling of corneal light transmission will be applied to this data to ascertain the key structural requirements for corneal transparency. We will also investigate the cellular contribution to corneal transparency, both by mathematical modelling and by direct measurement of light scattering.Finally, we will interfere with cell-communication pathways in corneal keratocytes, extracted from developing chick corneas and grown in the lab, to pinpoint what molecular mechanisms they use to communicate. These experiments and analyses, using a new way of growing cells in a three-dimensional environment, which we successfully developed to encourage tendon cells to synthesise aligned collagen fibres, will provide great insights for the field of corneal tissue engineering to help inform the intelligent design of the next generation of bioengineered corneal constructs.

角膜是眼睛前面的透明组织，它传输光线并将其清晰地聚焦到视网膜上。因此，它对于视力来说是绝对必要的。角膜主要由胶原蛋白组成，与其他富含胶原蛋白的组织如肌腱、软骨、椎间盘甚至巩膜没有什么不同。 “眼白”。但使角膜光学清晰的是胶原蛋白结构的精巧方式。人体内的胶原蛋白主要以长纤维的形式存在，这些纤维沿其轴非常坚固，可以被认为具有机械性能，就像帆船上的绳索或吊桥上的钢缆一样。不同组织中胶原纤维的组织取决于组织的功能，主要要求通常是机械性的。角膜在这方面是独特的，因为除了需要满足机械要求外，它还需要是透明的。角膜的大部分由细胞（称为角膜细胞）和胶原蛋白组成。大多数胶原蛋白以非常长且非常细的纤维或原纤维的形式存在，因为它们非常薄，因此在角膜中被称为原纤维。值得注意的是，与体内所有其他富含胶原蛋白的组织不同，角膜中的胶原纤维直径完全相同，并排列成近乎完美的六边形格子。正是胶原蛋白的这种精确的结构排列赋予了角膜透明度。但是，它是如何产生的呢？角膜中的角质细胞合成并沉积胶原蛋白。因此，推测这些细胞必须至少部分负责胶原原纤维的铺设和排列方式。我们将采用各种放大倍数的开创性显微方法，利用激光和电子成像技术，并与加州大学的科学家合作，研究受精鸡蛋中发育中的小鸡的角膜。我们的目标是准确了解发育中的角膜中的细胞如何相互作用，从而形成透明角膜这样结构精美的组织（小鸡角膜透明的基础与人类相同）。最近，我们率先使用三维体积电子显微镜来研究角膜，并发现发育中的小鸡角膜中的细胞都具有高度延伸但薄的细胞突起。这表明细胞本身占据的角膜体积比之前认为的要大得多。基于这一发现，我们的假设是，发育中的角膜中的细胞形成了一个扩展的网络，它们在其中相互交流，并且作为一个整体，它们具有天生的能力来合成极薄的胶原纤维，并将它们沉积成精确的格子状排列，以满足透明度的需要。为了检验这一假设，我们将从发育的最早阶段开始研究角膜，以前从未在如此高的放大倍数下进行过三维检查。角膜光传输数学建模的新方法将应用于这些数据，以确定角膜透明度的关键结构要求。我们还将通过数学建模和直接测量光散射来研究细胞对角膜透明度的贡献。最后，我们将干扰从发育中的小鸡角膜中提取并在实验室中生长的角膜角膜细胞的细胞通讯途径，以查明它们使用什么分子机制进行通讯。这些实验和分析使用了一种在三维环境中生长细胞的新方法，我们成功开发了这种方法来鼓励肌腱细胞合成对齐的胶原纤维，将为角膜组织工程领域提供深刻的见解，从而有助于为下一代生物工程角膜结构的智能设计提供信息。

项目成果

CD200 facilitates the isolation of corneal epithelial cells derived from human pluripotent stem cells.

• DOI: 10.1038/s41598-018-34845-2
• 发表时间: 2018-11-08
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Hayashi R;Ishikawa Y;Katayama T;Quantock AJ;Nishida K
• 通讯作者: Nishida K

A Surgical Cryoprobe for Targeted Transcorneal Freezing and Endothelial Cell Removal.

• DOI: 10.1155/2017/5614089
• 发表时间: 2017
• 期刊: Journal of ophthalmology
• 影响因子: 1.9
• 作者: Akhbanbetova A;Nakano S;Littlechild SL;Young RD;Zvirgzdina M;Fullwood NJ;Weston I;Weston P;Kinoshita S;Okumura N;Koizumi N;Quantock AJ
• 通讯作者: Quantock AJ

Fabrication of a Corneal Epithelial Cell Sheet from Human Pluripotent Stem Cells by a Method Based on Spontaneous Ocular Cell Differentiation

基于眼细胞自发分化的方法从人多能干细胞制备角膜上皮细胞片

• DOI: 10.1038/protex.2016.009
• 发表时间: 2016
• 期刊: Protocol Exchange
• 作者: Hayashi R

Chondroitin Sulphate/Dermatan Sulphate Proteoglycans: Potential Regulators of Corneal Stem/Progenitor Cell Phenotype In Vitro.

• DOI: 10.3390/ijms24032095
• 发表时间: 2023-01-20
• 期刊: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
• 影响因子: 5.6
• 作者: Bains, Kiranjit K.;Ashworth, Sean;Koudouna, Elena;Young, Robert D.;Hughes, Clare E.;Quantock, Andrew J.
• 通讯作者: Quantock, Andrew J.