Nozari Amin, Do Selena, Trudeau Vance L
Department of Biology, University of Ottawa, Ottawa, ON, Canada.
Front Endocrinol (Lausanne). 2021 Nov 17;12:727777. doi: 10.3389/fendo.2021.727777. eCollection 2021.
Transgenic zebrafish models have been successfully used in biomonitoring and risk assessment studies of environmental pollutants, including xenoestrogens, pesticides, and heavy metals. We employed zebrafish larva (transgenic SR4G line) with a cortisol-inducible green fluorescence protein reporter (eGFP) as a model to detect stress responses upon exposure to compounds with environmental impact, including bisphenol A (BPA), vinclozolin (VIN), and fluoxetine (FLX). Cortisol, fluorescence signal, and mRNA levels of eGFP and 11 targeted genes were measured in a homogenized pool of zebrafish larvae, with six experimental replicates for each endpoint. Eleven targeted genes were selected according to their association with stress-axis and immediate early response class of genes. Hydrocortisone (CORT)and dexamethasone (DEX) were used as positive and negative controls, respectively. All measurements were done in two unstressed and stressed condition using standardized net handling as the stressor. A significant positive linear correlation between cortisol levels and eGFP mRNA levels was observed (r> 0.9). Based on eGFP mRNA levels in unstressed and stressed larvae two predictive models were trained (Random Forest and Logistic Regression). Both these models could correctly predict the blunted stress response upon exposure to BPA, VIN, FLX and the negative control, DEX. The negative predictive value (NPV) of these models were 100%. Similar NPV was observed when the predictive models trained based on the mRNA levels of the eleven assessed genes. Measurement of whole-body fluorescence intensity signal was not significant to detect blunted stress response. Our findings support the use of SR4G transgenic larvae as an biomonitoring model to screen chemicals for their stress-disrupting potentials. This is important because there is increasing evidence that brief exposures to environmental pollutants modify the stress response and critical coping behaviors for several generations.
转基因斑马鱼模型已成功应用于环境污染物的生物监测和风险评估研究，这些污染物包括外源性雌激素、农药和重金属。我们使用带有皮质醇诱导型绿色荧光蛋白报告基因（eGFP）的斑马鱼幼虫（转基因SR4G品系）作为模型，来检测暴露于具有环境影响的化合物（包括双酚A（BPA）、乙烯菌核利（VIN）和氟西汀（FLX））时的应激反应。在一组匀浆的斑马鱼幼虫中测量皮质醇、荧光信号以及eGFP和11个目标基因的mRNA水平，每个终点进行6次实验重复。根据11个目标基因与应激轴和即时早期反应基因类别的关联进行选择。氢化可的松（CORT）和地塞米松（DEX）分别用作阳性和阴性对照。所有测量均在两种无应激和应激条件下进行，使用标准化的网捞操作作为应激源。观察到皮质醇水平与eGFP mRNA水平之间存在显著的正线性相关性（r>0.9）。基于无应激和应激幼虫中的eGFP mRNA水平训练了两个预测模型（随机森林和逻辑回归）。这两个模型都能正确预测暴露于BPA、VIN、FLX和阴性对照DEX时的应激反应减弱情况。这些模型的阴性预测值（NPV）为100%。当基于11个评估基因的mRNA水平训练预测模型时，观察到类似的NPV。测量全身荧光强度信号对于检测应激反应减弱并不显著。我们的研究结果支持使用SR4G转基因幼虫作为生物监测模型，以筛选具有应激干扰潜力的化学物质。这很重要，因为越来越多的证据表明，短期暴露于环境污染物会改变应激反应和几代人的关键应对行为。
