Hinther Ashley, Edwards Thea M, Guillette Louis J, Helbing Caren C
Department of Biochemistry and Microbiology, University of Victoria Victoria, BC, Canada.
Front Genet. 2012 Apr 4;3:51. doi: 10.3389/fgene.2012.00051. eCollection 2012.
Nitrate and nitrite are common aqueous pollutants that are known to disrupt the thyroid axis. In amphibians, thyroid hormone (TH)-dependent metamorphosis is affected, although whether the effect is acceleration or deceleration of this developmental process varies from study to study. One mechanism of action of these nitrogenous compounds is through alteration of TH synthesis. However, direct target tissue effects on TH signaling are hypothesized. The present study uses the recently developed cultured tail fin biopsy (C-fin) assay to study possible direct tissue effects of nitrate and nitrite. Tail biopsies obtained from premetamorphic Rana catesbeiana tadpoles were exposed to 5 and 50 mg/L nitrate (NO(3)-N) and 0.5 and 5 mg/L nitrite (NO(2)-N) in the absence and presence of 10 nM T(3). Thyroid hormone receptor β (TRβ) and Rana larval keratin type I (RLKI), both of which are TH-responsive gene transcripts, were measured using quantitative real time polymerase chain reaction. To assess cellular stress which could affect TH signaling and metamorphosis, heat shock protein 30, and catalase (CAT) transcript levels were also measured. We found that nitrate and nitrite did not significantly change the level of any of the four transcripts tested. However, nitrate exposure significantly increased the heteroscedasticity in response of TRβ and RLKI transcripts to T(3). Alteration in population variation in such a way could contribute to the previously observed alterations of metamorphosis in frog tadpoles, but may not represent a major mechanism of action.
硝酸盐和亚硝酸盐是常见的水体污染物，已知它们会扰乱甲状腺轴。在两栖动物中，甲状腺激素（TH）依赖的变态发育会受到影响，不过这种影响是加速还是减缓这个发育过程，不同研究的结果各不相同。这些含氮化合物的一种作用机制是通过改变TH的合成。然而，有人推测它们对TH信号传导存在直接的靶组织效应。本研究使用最近开发的培养尾鳍活检（C-fin）试验，来研究硝酸盐和亚硝酸盐可能的直接组织效应。从变态前的牛蛙蝌蚪获取尾组织活检样本，将其置于不存在和存在10 nM T3的情况下，分别暴露于5和50 mg/L硝酸盐（NO(3)-N）以及0.5和5 mg/L亚硝酸盐（NO(2)-N）中。使用定量实时聚合酶链反应测量甲状腺激素受体β（TRβ）和牛蛙幼虫I型角蛋白（RLKI），这两者都是TH反应性基因转录本。为了评估可能影响TH信号传导和变态发育的细胞应激，还测量了热休克蛋白30和过氧化氢酶（CAT）的转录水平。我们发现，硝酸盐和亚硝酸盐并未显著改变所测试的四种转录本中任何一种的水平。然而，硝酸盐暴露显著增加了TRβ和RLKI转录本对T3反应的异方差性。以这种方式改变群体变异可能导致先前观察到的青蛙蝌蚪变态发育的改变，但可能并不代表主要的作用机制。
