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

限制眼睛的外壳必须坚硬，以防止异物和细菌进入内部。它必须很轻，这样眼睛就不会那么沉重，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，至少在前部，它必须是透明的，这样光线才能到达视网膜，我们才能看到周围。但是，大自然是如何发展出一种策略来建造一个坚固、轻盈又透明的结构呢？工程师们都知道，使用合适的材料是很重要的。眼睛的外部外壳，特别是它的前窗（称为角膜）的主要分子成分是胶原蛋白。胶原蛋白是一种链状蛋白质，聚集在一起形成长长的分子绳，称为原纤维。胶原蛋白很轻，因为它能聚集成原纤维，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，我们可以看到胶原原纤维并排排列形成片状结构。薄片中单个相邻原纤维之间的距离非常精确，约为50纳米。胶原原纤维之间距离的精确度是透明度的关键。为了理解其中的原因，我们可以先想想当我们把一块石头扔到池塘的平坦表面上会发生什么。当石头撞到池塘表面时，产生了从中心点向外移动的波浪圈。如果我们投掷两块石头，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；在两个波峰相交的地方，我们会得到一个两倍高的波峰。在穿过的地方，我们会有一个低两倍的穿过，在波峰和穿过的地方，波浪会消失。如果我们设法把一排排列整齐的石头扔下去，一个挨一个，我们就会发现，除了向前的方向外，圆形的波浪在各个方向上都消失了。这是因为任何波浪的波峰都会在任何地方遇到另一个波浪的穿过，除了前进方向。光本身就是一种波，当它碰到胶原蛋白纤维时，产生的波与池塘表面的石头产生的波没有什么不同。因为胶原纤维的间距是相等的，所以除了向前方向外，所有方向的光都被抵消了，光可以穿过角膜到达视网膜。但有一件重要的事情我们还不知道，那就是角膜中的胶原蛋白是如何保持等距分布的。我们知道在胶原原纤维周围有其他的蛋白质叫做蛋白聚糖。在电子显微镜下可以看到蛋白聚糖伸向邻近的胶原原纤维，以这种方式构建一个网络，可能有助于保持胶原原纤维的均匀间隔。在这个项目中，我们打算找出胶原原纤维周围和之间的蛋白聚糖的确切位置和三维空间排列；因此，我们想要确定保持胶原原纤维的确切机制。我们还建议通过将已知强度的光照射在角膜的各个区域并测量出光的强度来测量角膜的透明度。最后，我们希望将不同区域角膜透明度的测量与显微镜下对胶原原纤维间距、原纤维直径、蛋白多糖位置和三维结构的测量联系起来。通过这种方式，我们将了解蛋白聚糖是如何在角膜中维持三维秩序的，蛋白聚糖对胶原原纤维直径和纤维间距离有什么影响，以及这些参数对角膜透明度有多重要。这一信息对研究角膜失去透明度的眼部疾病的医生非常有用，对那些想要为失去角膜功能的人制造人工角膜的生物工程师也非常有用。

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.