Novel approaches to immortalise salmon fish cells.

动物通常用于生物科学的基础和应用研究。随着养鱼业的发展，人们开展了越来越多的研究，以了解疾病如何影响鱼类，以便找到对抗疾病和保护鱼类福利的方法。为了了解一种病毒如何传播疾病，或者它对鱼的致病性如何，通常在一个特殊的密闭水族馆设施中对一组鱼进行实验感染，并监测其影响。为了回答某些问题，有时有必要用鱼进行实验（体内实验），但通常可以通过在不朽的鱼细胞上进行实验（体外实验）来获得有价值的信息。这些细胞在含有营养物质的合成培养基中的塑料培养皿中分裂和无限生长，并产生细胞系，这是一种无限的生物材料来源。细胞从最初的几个细胞向外分裂，在培养皿表面形成一层。细胞系通常由一种细胞类型组成，例如肾细胞、心脏细胞或肾细胞。细胞系可以用来测试从死鱼身上提取的组织样本是否含有病毒（诊断疾病），或者鱼细胞是否能产生对抗病毒的产物（抗病毒反应）。当病毒被添加到鱼细胞中时，它会感染细胞并杀死它们，从而在细胞层上形成很容易看到的空隙（细胞病变效应）。不幸的是，有许多导致严重疾病的病毒株在目前可用的鱼类细胞系中不能很好地生长。此外，在鱼类中，不可能控制哪种类型的细胞会产生细胞系。这是因为即使你从心脏、肾脏或鳃组织的样本开始，这些组织都是由几种细胞类型组成的，迄今为止，在鱼类中，细胞系是“靠运气”产生的，也就是说，组织中的一种细胞类型可能开始分裂并继续分裂，但它可能不是病毒通常感染的细胞类型。事实上，很难将目前可用的一些细胞系分配给特定的鱼类组织。本项目旨在寻找新的方法，从目标细胞类型生产鱼细胞系。从鱼类器官分离出来的细胞可以通过基因改造进行永久性细胞分裂，并在合成培养基中存活。这些细胞可以无限繁殖，可以用体外实验代替动物实验。将单独或联合测试几种策略：通过已发现的诱导癌细胞分裂和生长的因素、通过生长因素、通过抑制细胞死亡的因素、通过已知可触发血细胞增殖的寄生虫产生的因素来实现细胞的永生。可以产生这些因子的遗传系统将被引入从不同组织类型中分离出来的初始细胞中，这些因子将作用于细胞以保持其分裂和/或防止其死亡。最终目标是减少调查和诊断鱼类疾病所需的活鱼数量，并提供用于改善鱼类健康和福利的工具。随着野生鱼类种群面临的压力越来越大，世界人口不断增加，水产养殖已成为许多社区重要的蛋白质来源，并极有可能继续增长。

Animals are commonly used in biological sciences for both fundamental and applied research. With the growth of the fish farming industry, more and more studies are carried out to understand how disease affects fish in order to find ways to combat them and protect fish welfare.To gain understanding on how a virus can transmit disease, or how pathogenic it is for the fish experiments are often carried out where a group of fish is experimentally infected in a special containment aquarium facility and the effects monitored. It is sometimes necessary to carry out experiments with fish (in vivo experiments) to answer certain questions but often valuable information can be obtained by performing the experiments instead on immortal fish cells (in vitro experiments). These are cells which divide and grow indefinitely on plastic dishes in a synthetic culture medium containing nutrients and give rise to a cell line, an indefinite source of biological material. The cells divide outwards from a few initial cells to form a layer on the surface of the dish. The cell lines are usually composed on one cell type, e.g. a kidney cell, heart cell or kidney cell. Cell lines can be used to test whether a tissue sample taken from a dead fish contains a virus or not (diagnosing disease), or whether the fish cell can make products to combat the virus (anti-viral response). When the virus is added to the fish cells, it infects the cells and kills them and this forms gaps in the cell layer which are easily seen (cytopathic effect). Unfortunately, there are many strains of viruses responsible for serious diseases that do not grow very well in the fish cell lines currently available. In addition in fish there is no possibility to control what type of cell will generate a cell line. This is because even if you start with a sample of heart or kidney or gill tissue, these tissues are made up of several cell types and to date, in fish, cell lines are generated "by luck", that is, one of the cell types in the tissue may start to divide and continue to do so, but it may not be the cell type which the virus normally infects. In fact, it is difficult to allocate some of the currently available cell lines to a certain fish tissue.The present project aims to find new methods to produce fish cell lines from targeted cell types. Cells dissociated from fish organs can be genetically modified to undergo a permanent cell division and survive in synthetic culture medium. These cells would propagate indefinitely and can be used to replace animal experiments by in vitro ones. Several strategies will be tested alone or in combination: immortalisation of cells by factors which have been found to induce cancer cells to divide and grow, by growth factor, by factors which inhibit cell death, by factors produced by parasites known to trigger the proliferation of blood cells. Genetic systems which can produce these factors will be introduced into the initial cells isolated from different tissue types, and the factors will act on the cell to keep it dividing and/or prevent it from dying. The ultimate aim is to reduce the number of live fish needed to investigate and diagnose fish disease and to provide tools for use in improving fish health and welfare. With increasing pressure on wild fish stocks and an increasing world population, aquaculture has become an important provider of protein for many communities and will most likely continue to grow.