Presslauer Christopher, Nagasawa Kazue, Dahle Dalia, Babiak Joanna, Fernandes Jorge M O, Babiak Igor
Faculty of Biosciences and Aquaculture, University of Nordland, 8049 Bodø, Norway.
PLoS One. 2014 Dec 1;9(12):e114209. doi: 10.1371/journal.pone.0114209. eCollection 2014.
A method to mitigate or possibly eliminate reproduction in farmed fish is highly demanded. The existing approaches have certain applicative limitations. So far, no immunization strategies affecting gonadal development in juvenile animals have been developed. We hypothesized that autoimmune mechanisms, occurring spontaneously in a number of diseases, could be induced by targeted immunization. We have asked whether the immunization against specific targets in a juvenile zebrafish gonad will produce an autoimmune response, and, consequently, disturbance in gonadal development. Gonadal soma-derived factor (Gsdf), growth differentiation factor (Gdf9), and lymphocyte antigen 75 (Cd205/Ly75), all essential for early gonad development, were targeted with 5 immunization tests. Zebrafish (n = 329) were injected at 6 weeks post fertilization, a booster injection was applied 15 days later, and fish were sampled at 30 days. We localized transcripts encoding targeted proteins by in situ hybridization, quantified expression of immune-, apoptosis-, and gonad-related genes with quantitative real-time PCR, and performed gonadal histology and whole-mount immunohistochemistry for Bcl2-interacting-killer (Bik) pro-apoptotic protein. The treatments resulted in an autoimmune reaction, gonad developmental retardation, intensive apoptosis, cell atresia, and disturbed transcript production. Testes were remarkably underdeveloped after anti-Gsdf treatments. Anti-Gdf9 treatments promoted apoptosis in testes and abnormal development of ovaries. Anti-Cd205 treatment stimulated a strong immune response in both sexes, resulting in oocyte atresia and strong apoptosis in supporting somatic cells. The effect of immunization was FSH-independent. Furthermore, immunization against germ cell proteins disturbed somatic supporting cell development. This is the first report to demonstrate that targeted autoimmunity can disturb gonadal development in a juvenile fish. It shows a straightforward potential to develop auto-immunization-based technologies to mitigate fish reproduction before they reach maturation. However, the highly variable results between treatments and individuals suggest significant optimization should be performed to achieve the full potential of this technology.
人们迫切需要一种减轻或可能消除养殖鱼类繁殖的方法。现有的方法存在一定的应用局限性。到目前为止，尚未开发出影响幼年动物性腺发育的免疫策略。我们假设，在许多疾病中自发出现的自身免疫机制可以通过靶向免疫诱导产生。我们研究了针对幼年斑马鱼性腺中的特定靶点进行免疫是否会产生自身免疫反应，进而导致性腺发育紊乱。性腺体细胞衍生因子（Gsdf）、生长分化因子（Gdf9）和淋巴细胞抗原75（Cd205/Ly75）都是早期性腺发育所必需的，我们通过5项免疫试验对其进行了靶向研究。在受精后6周对斑马鱼（n = 329）进行注射，15天后进行加强注射，并在30天时对鱼进行取样。我们通过原位杂交定位编码靶向蛋白的转录本，用定量实时PCR定量免疫、凋亡和性腺相关基因的表达，并对Bcl2相互作用杀手（Bik）促凋亡蛋白进行性腺组织学和整体免疫组织化学分析。这些处理导致了自身免疫反应、性腺发育迟缓、强烈的细胞凋亡、细胞闭锁和转录产物紊乱。抗Gsdf处理后，睾丸明显发育不全。抗Gdf9处理促进了睾丸中的细胞凋亡和卵巢的异常发育。抗Cd205处理在两性中均刺激了强烈的免疫反应，导致卵母细胞闭锁和支持性体细胞中的强烈细胞凋亡。免疫的效果不依赖于促卵泡激素。此外，针对生殖细胞蛋白的免疫会干扰体细胞支持细胞的发育。这是第一份证明靶向自身免疫可干扰幼年鱼类性腺发育的报告。它显示出开发基于自身免疫的技术以在鱼类成熟前减轻其繁殖的直接潜力。然而，处理之间和个体之间结果的高度变异性表明，应进行重大优化以充分发挥该技术的潜力。
