On the three-dimensional structure of the proteoglycans in the cornea and how it controls corneal transparency

限制眼睛的外部外壳必须坚固，以防止异物和细菌渗入内部。它必须很轻，这样眼睛才不会不必要地沉重，当我们想要环顾四周时，微小的肌肉可以高精度地移动它。但最重要的是，至少在前面，它必须是透明的，这样光线才能到达视网膜，我们可以看到周围。但是，大自然制定了什么策略来建立一个坚固、轻便、透明的结构呢？正如工程师们所知，使用正确的材料是很重要的。眼睛外壳的主要分子成分，特别是它的前窗(称为角膜)的分子成分是胶原蛋白。胶原蛋白是一种链状蛋白质，它聚集在一起形成被称为纤维的长分子绳。胶原蛋白很轻，因为它能组装成纤维，所以也很坚韧。如果我们用电子显微镜观察角膜中的胶原蛋白，我们可以看到胶原蛋白纤维并排排列，形成片状结构。一张纸上单个相邻纤维之间的距离非常精确，约为50 nm。胶原蛋白纤维之间距离的精确度是透明的关键。要理解为什么，首先考虑一下当我们把一块石头扔到池塘的平坦表面上会发生什么，这是很有用的。当石头击中池塘表面时，会产生从中心点向外移动的波形圈。如果我们扔两块石头，一块石头产生的波与另一块石头产生的波相遇，就会产生干涉图案；在两个波的波峰相遇的地方，我们就会有一个两倍高的波峰。在穿透的地方，我们将有一个低一倍的穿透，而在一个波峰与穿透的相遇之处，波浪将消失。如果我们设法落下一排规则间隔的石头，其中一块离另一块很近，我们会发现圆形波将在除向前方向外的所有方向上消失。这是因为任何波的波峰都会在除向前方向以外的任何地方与另一波的穿透相遇。光本身就是一种波，当它击中胶原纤维时，它产生的波与池塘表面的石头产生的波没有什么不同。由于胶原蛋白纤维的间距相等，光在除正向外的所有方向上都会被抵消，光可以穿过角膜到达视网膜。然而，有一件关键的事情是未知的，那就是角膜中的胶原蛋白是如何保持均匀分布的。我们知道，在胶原纤维周围还有其他称为蛋白多糖的蛋白质。在电子显微镜中可以看到蛋白多糖伸向邻近的胶原纤维，以这种方式建立一个网络，可能有助于保持胶原纤维的均匀分布。在这个项目中，我们打算找出蛋白多糖在胶原纤维周围和之间的确切位置和三维空间排列；因此，我们想要确定保持胶原纤维在适当位置的确切机制。我们还建议通过照射已知强度的光线和测量发出的光线的强度来测量角膜不同区域的角膜透明度。最后，我们希望将不同区域角膜透明度的测量与显微镜下测量的胶原纤维间距、纤维直径、蛋白多糖位置和三维结构相关联。通过这种方式，我们将了解蛋白多糖是如何在角膜中维持三维有序的，蛋白多糖对胶原纤维直径和纤维间距有什么影响，以及这些参数对角膜透明度有多重要。这些信息将对研究眼部疾病的医生非常有用，因为在这种疾病中，角膜失去了透明度，对于希望为失去角膜功能的人制造人工角膜的生物工程师也是如此。

The external case limiting the eye must be tough to prevent foreign objects and germs from penetrating inside. It has to be light so that the eye is not unnecessarily heavy and tiny muscles can move it with high precision when we want to look around. But above all, at least at the front, it has to be transparent, so that light can reach the retina and we can see around us. But what strategy has nature developed to build a structure which is tough, light and also transparent? As engineers know, it is important to use the right materials. The main molecular component of the external case of the eye, and in particular that of its front window (called the cornea) is collagen. Collagen is a chain-like protein that assembles together to form long molecular ropes called fibrils. Collagen is light and because it assembles into fibrils, it is very tough as well. If we look at the collagen in the cornea using an electron microscope, we can see that the collagen fibrils run side by side to form sheet-like structures. The distance between individual neighbouring fibrils in a sheet is very precise and about 50 nm. The precision of the distance between the collagen fibrils is the key to transparency. To understand why, it is useful to think first of what happens when we throw a stone on the flat surface of a pond. When the stone hits the pond surface, wave circles are produced that move outwards from a central point. If we throw two stones, the waves produced by one stone meet those produced by the other stone and an interference pattern is produced; where the crests of two waves meet, we will have a wave crest that is twice as high. Where the throughs meet we will have a through which is twice as low, and where a crest meets a through, the wave will disappear. If we managed to drop a row of regularly spaced stones, one quite close to another, we would find that the circular waves would disappear in all directions, except the forward direction. This is because the crest of any wave would meet the through of another wave everywhere except in the forward direction. Light is itself a wave, and when it hits a collagen fibril, it produces waves not dissimilar from those produced by a stone on a pond surface. Because the collagen fibres are equally spaced, light cancels out in all directions except the forward direction and the light can cross the cornea to reach the retina. One crucial thing which is not known though, is how the collagen in the cornea is maintained equally spaced. We know that around the collagen fibrils there are other proteins called proteoglycans. Proteoglycans are seen in the electron microscope to reach out for adjacent collagen fibrils, building in this way a network that may help to keep the collagen fibrils equally spaced. In this project, we propose to find out the exact location and 3-dimensional spatial arrangement of the proteoglycans around and between the collagen fibrils; thus, we want to determine the exact mechanisms that keep the collagen fibrils in place. We also propose to measure corneal transparency in various areas of the cornea by shining light of known intensity through it and by measuring the intensity of the light coming out. Finally we want to correlate the measurement of the corneal transparency in different areas with the measurements taken in the microscope about the collagen fibril spacing, fibril diameters and proteoglycan location and 3-dimensional structure. In this way we will have an idea of how the 3-dimensional order is maintained in the cornea by the proteoglycans, what effects proteoglycans have on collagen fibril diameter and interfibrillar distance, and how important these parameters are for corneal transparency. This information will be very useful to doctors studying the illnesses of the eye in which the cornea loses its transparency, and also for bioengineers who want to build artificial corneas for people that have lost their corneal functionality.