Direct visualisation of epithelial fluid transport at the subcellular scale

上皮细胞是一种特殊的细胞，它形成了身体组织与内部或外部世界之间的屏障。不同的器官具有不同的功能和不同的内环境，这些功能和内环境是由上皮细胞的运输活动建立和维持的。上皮细胞形成极化片，其顶端和底侧“端”在屏障上相反的方向。这种极性允许上皮片以定向方式泵送液体，以维持单个组织和整个身体的正常生理。这些流体泵送过程对正常健康至关重要，在各种疾病状态下经常被发现是错误的。例如，囊性纤维化患者的液体分泌紊乱会通过这一共同途径影响肺、消化系统和其他器官的功能。在眼睛中，紊乱的液体调节被认为是造成青光眼、白内障形成、黄斑变性和视网膜脱离的原因。这两种疾病导致了世界上大多数的失明。视网膜色素上皮位于视网膜神经元的后面，通过为神经细胞层做“家务”来执行一系列功能，以确保视力的保存。色素上皮是黑色的，以避免在眼球内反射，并伸出细小的手指包裹并保护重要的感光细胞，这些细胞负责实际的“观看”。当上皮被破坏或损伤时，液体会像水疱一样积聚在神经元和上皮之间，使上皮不能帮助神经元正常工作。我们仍然不清楚上皮如何正常工作以防止这种情况的发生，并且想知道如何通过正确的方式提高细胞的液体泵送能力来帮助细胞修复这种损伤。当上皮细胞在一个非常有限的空间内，在细胞之间泵送液体时，发生了很多事情。盐被泵入这些微小的空间，水随之产生液体分泌。科学家们仍然不确定这是如何做到的，以精确地匹配特定功能所需的盐和水的平衡——从眼泪和汗水到尿液和胆汁，这些分泌物的性质变化很大。唯一的问题是，这些如此重要的小空间也太小了，不容易往里看。该项目将开发一种新的工具，帮助我们了解上皮液体的运输。这实际上是一种工具的组合，所有的工具都被应用在一起，这将给我们关于整个组织的信息-从大到小。提出了两种新技术（一种是光学测量技术，另一种是光学刺激技术），它们将在我们认为对流体输送至关重要的微小空间中测量盐和水的运动。通过将它们与大规模测量相结合，我们可以进行实验，看看盐的分泌如何影响水的分泌，反之亦然。这有点像先有鸡还是先有蛋的问题，但通过一次进行许多测量来解决这个问题应该会给我们提供我们正在寻找的答案，从而了解发生了什么。如果我们能更好地了解液体的输送，我们就能在治疗由液体输送问题引起的疾病时做出更好的选择。特别是，这个项目将给那些患有视网膜疾病的人带来希望，这些疾病可能会通过我们在这个研究项目中开发的策略得到更好的治疗。我们希望将该系统也提供给其他研究其他组织中流体输送的研究人员，这样新技术的好处就能产生尽可能广泛的影响。

Epithelial cells are the specialized cells which form the barriers between tissues of the body and the inside or outside worlds. Different organs have different functions and different internal environments, which are set up and maintained by the transporting activities of epithelial cells. Epithelial cells form polarised sheets whose apical and basolateral "ends" face in opposite directions across the barrier. This polarity allows epithelial sheets to pump fluid in a directed fashion to maintain the normal physiology of individual tissues and the entire body. These fluid pumping processes are crucial to normal health and often found to be awry in various disease states. For example, disrupted fluid secretion in cystic fibrosis patients affects the function of their lungs, digestive system, and other organs all through this common pathway. In the eye, disturbed fluid regulation is believed to be responsible for glaucoma, cataract formation, macular degeneration, and retinal detachment. Between them, these diseases cause the majority of blindness worldwide. The retinal pigment epithelium lies behind the neurons of the retina, and performs a series of functions to make sure vision is preserved by doing the "housekeeping" for the layers of nerve cells. The pigment epithelium is black to avoid reflections within the eyeball, and reaches out tiny fingers to wrap around and protect the vital photoreceptor cells which do the actual "seeing". When the epithelium is disrupted or injured, fluid can accumulate between the neurons and the epithelium like a blister, keeping the epithelium from helping the neurons to work properly. We still don't understand exactly how the epithelium works normally to prevent this from happening, and would like to know how to help the cells repair such damage by boosting their fluid pumping abilities in the right way. A lot of what goes on when epithelia pump fluid happens between the cells, in a very confined space. Salt is pumped into these tiny spaces and water follows along to create secretion of fluid. Scientists are still not sure exactly how this is done to precisely match the balance of salts and water required for particular functions - from tears and sweat to urine and bile, these secretions can vary widely in their properties. The only trouble is, these tiny spaces which are so important are also too small to look inside very easily. This project will develop a new tool which will help us understand epithelial fluid transport. It's actually a combination of tools, all being applied together, which will give us information about what the whole tissue is doing - from the large scale right down to the very small. Two new techniques are proposed (one an optical measurement, and one an optical stimulation technique) which will make measurements of salt and water movements in the tiny spaces we believe are crucial to fluid transport. By applying them together with measurements at the large scale, we can then perform experiments to see how the secretion of salt affects secretion of water, and vice versa. It's a bit of a chicken and egg problem, but attacking it by making many measurements at once should give us the answers we are looking for to understand what's going on. If we understand fluid transport better we can make better choices in treating diseases resulting from fluid transport problems. In particular, this project will give hope to those with retinal disease which might be better treated by strategies we develop during this research project. We hope to make the system available to other researchers looking at fluid transport in other tissues too, so that the benefits of new technology can have the widest impact possible.