In aquaculture, fish species may experience stressful episodes caused by poor farming conditions. The exponential increase of global aquaculture has raised the number of research studies aimed at demonstrating the sensitivity of aquatic animals in confined environments. The development of a real-time PCR and immunohistochemistry methods were investigated to evaluate the presence, localization, and quantity of biomarkers of oxidative stress in European sea bass (Dicentrarchus labrax). In particular, stress tests such as manipulation and temperature changes were conducted through molecular methods to identify the expression level of heat shock protein 70 (HSP70) in stressed animals compared with a control group. The immunohistochemical technique was also applied to locate and study the trends-levels of nitrotyrosine (NT), heat shock protein 70 (HSP70), malondialdehyde (MDA), and 4-hydroxy-2-nonenal (HNE) in different tissues from stressed animals and control group. The presence of the rodlet cell (RCs) was evaluated by histology in both a control and stressed group. Our results show that the real-time PCR method developed is specific for the evaluated target gene and that manipulation and temperature increase are strong stressors for animals. Relative quantification data revealed a gene expression increase of HSP70 in the stressed group of animals compared to the control group. The antibodies used for the immunohistochemical staining were efficient, and it was possible to appreciate the increase of immunoprecipitates in European sea bass either manipulated or stressed by temperature increase. The present study can be a starting point to allow the quantification of HSP70 and the identification of other stress biomarkers in D. labrax.

中文翻译：

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开发分子和组织学方法以评估鲈鱼（Dicentrarchus labrax）中的应力氧化生物标记。

在水产养殖中，鱼类品种可能因恶劣的耕作条件而承受压力。全球水产养殖的呈指数增长已经增加了旨在证明水生动物在受限环境中的敏感性的研究数量。研究了实时PCR和免疫组织化学方法的发展，以评估欧洲鲈鱼（Dicentrarchus labrax）中氧化应激生物标记物的存在，定位和数量。尤其是，通过分子方法进行了诸如操作和温度变化之类的压力测试，以确定与对照组相比应激动物中热休克蛋白70（HSP70）的表达水平。免疫组化技术也被用于定位和研究硝基酪氨酸（NT）的趋势水平，应激动物和对照组不同组织中的热休克蛋白70（HSP70），丙二醛（MDA）和4-羟基-2-壬烯醛（HNE）。对照组和应激组均通过组织学评估了干细胞（RCs）的存在。我们的结果表明，开发的实时PCR方法对评估的目标基因具有特异性，并且操纵和温度升高是动物的强大压力源。相对定量数据显示，与对照组相比，应激组动物的HSP70基因表达增加。用于免疫组织化学染色的抗体是有效的，并且有可能欣赏到欧洲鲈鱼中免疫沉淀物的增加，无论是受温度升高操纵还是受到温度升高的影响。