CITAB-Centre for the Research and Technology of Agro-Environmental and Biological Sciences, ECVA-School of Life and Environmental Sciences, UTAD-University of Trás-os-Montes and Alto Douro, Quinta de Prados, Apartado 1013, 5001-801, Vila Real, Portugal.
Environ Sci Pollut Res Int. 2014 Feb;21(4):3116-25. doi: 10.1007/s11356-013-2248-y. Epub 2013 Nov 8.
Liver lesions in wild fish have been associated with xenobiotic exposure. Facing reports of pollution in the Douro River estuary (north of Portugal), we have been making field surveys using fishes and targeting histopathological biomarkers of exposure and effect. Herein, we intended to better characterize and report the rate of one poorly understood lesion-hepatocellular fibrillar inclusions (HFI)-found in European flounder (Platichthys flesus). With this report, we aimed to establish sound baseline data that could be viewed as a starting point for future biomonitoring, while offering the world's second only pool of field data on such a liver toxicopathic lesion, which could be compared with data available from the UK estuaries. Sampling was done in the Douro River estuary over 1 year. A total of 72 animals were fished with nets, in spring-summer (SS) and autumn-winter (AW) campaigns. Livers were processed for histopathology and both routine and special staining procedures (alcian blue, periodic acid Schiff (PAS), tetrazonium coupling reaction). Immunohistochemistry targeted AE1/AE3 (pan cytokeratins). The severity of the HFI extent was graded using a system with four levels, varying from 0 (absence of HFI) to 3 (high relative density of cells with HFI). Cells (isolated/groups) with HFI appeared in 35 % or more of the fish, in the total samples of each season, and over 40 % in more homogeneous sub-samples. There were no significant differences when comparing samples versus sub-samples or SS versus AW. When merging the data sets from the two seasons, the frequency of fish with HFI was ≈36 % for the total sample and ≈49 % for the sub-sample. The extreme group (biggest and smallest fish) revealed a HFI frequency of only 16 %, which differed significantly from the total and sub-sampled groups. Immunostaining and PAS were negative for the HFI, and alcian blue could, at times, faintly stain the inclusions. These were positive with the tetrazonium reaction. We showed the presence of HFI in European flounder from the Douro River estuary, proving that they are essentially protein in nature, that no seasonal changes existed in the HFI frequency, and that it was rarer in the smallest and biggest fish groups. Within the ranges of weight/size of our total sample, we estimate that the frequency of HFI in the local flounder is ≈35 %. That rate stands as a baseline value for future assessments, namely for biomonitoring purposes targeting correlations with the estuary pollution status.
野生鱼类的肝脏病变与外源生物暴露有关。由于有报道称在葡萄牙北部的杜罗河口存在污染，我们一直在使用鱼类进行实地调查，针对的是暴露和效应的组织病理学生物标志物。在此，我们旨在更好地描述和报告在欧洲比目鱼（Platichthys flesus）中发现的一种尚未完全了解的病变——肝细胞纤维状内含物（HFI）的发生率。通过本报告，我们旨在建立可靠的基线数据，可以将其视为未来生物监测的起点，同时提供世界上仅有的第二个有关这种肝毒性病变的实地数据池，该数据可与来自英国河口的数据进行比较。该采样是在杜罗河口进行的，持续了一年。在春季-夏季（SS）和秋季-冬季（AW）两个季节中，使用网捕鱼，共采集了 72 条鱼。对肝脏进行了组织病理学处理，并进行了常规和特殊染色程序（阿利新蓝，过碘酸雪夫（PAS），四唑偶氮反应）。免疫组织化学针对 AE1/AE3（细胞角蛋白）。使用一个有四个等级的系统对 HFI 程度的严重程度进行分级，从 0（无 HFI）到 3（有 HFI 的细胞相对密度高）。在每个季节的总样本中，有 35%或更多的鱼以及在更均匀的亚样本中，有 HFI 的细胞（单个/组）出现。当比较样本与亚样本或 SS 与 AW 时，没有发现差异。当将两个季节的数据合并时，总样本中 HFI 鱼的频率约为 36%，亚样本中 HFI 鱼的频率约为 49%。最大和最小的鱼的极端组显示 HFI 的频率仅为 16%，与总样本和亚样本组有显著差异。免疫染色和 PAS 对 HFI 呈阴性，阿利新蓝有时可以淡染内含物。这些在四唑偶氮反应中呈阳性。我们证明了杜罗河口的欧洲比目鱼中存在 HFI，证明它们本质上是蛋白质，HFI 的频率在季节之间没有变化，而且在最小和最大的鱼群中更为罕见。在我们总样本的体重/大小范围内，我们估计当地比目鱼中 HFI 的频率约为 35%。该比率是未来评估的基准值，即针对与河口污染状况相关的生物监测目的。
