三氯生和三氯卡班对斑马鱼幼鱼的免疫毒性比较

Comparison of triclosan and triclocarban in triggering immunotoxicity in larval zebrafish

摘要:

三氯生(triclosan,TCS)和三氯卡班(triclocarban,TCC)作为两种高效的广谱杀菌剂被广泛使用，尤其是在新冠肺炎疫情期间，然而其二次污染造成的健康风险备受关注。TCS和TCC具有相似的母核结构和高亲脂性特点，其对环境生物的毒性效应和致毒机制的差异尚不明晰，尤其是二者的免疫毒性研究较少。本研究以脊椎动物斑马鱼为模型，比较二者在相同浓度暴露下(0.6µmol/L)的免疫毒性及其作用机制。结果发现，TCS和TCC均能导致受精后72 h (hours post fertilization,hpf)的斑马鱼受精卵孵化率低于60%，对120 hpf幼鱼的致死率分别达40%和50%，并伴有抑制体节生长、游囊关闭、心包水肿、卵黄囊肿淤积与吸收障碍等畸形，且TCC致畸程度显著高于TCS。TCS暴露刺激固有免疫细胞增殖率为20%，T细胞减少了35%，而TCC对固有免疫细胞和T细胞分化均呈抑制作用，抑制率分别为25%和60%。实时荧光定量PCR (real-time quantitative PCR,RT-qPCR)和ELISA的结果显示，TCS和TCC分别致il-1β、il-6和tnf-α炎症因子编码基因显著上调，而二者对il-10和IgM的表达呈相反趋势，且均导致C3的表达下降。皮尔逊(Pearson)相关性分析表明，发现TCS与TCC诱导的发育毒性与免疫细胞分化之间分别呈正相关和负相关，但二者均与促炎因子表达量呈正相关。GO功能和KEGG通路富集分析发现，TCS与TCC作用靶分子富集在不同的信号通路上，关键网络hub基因和富集的调控通路也有所不同。本研究为揭示TCS和TCC引发免疫毒性的不同机制提供了证据，并为二者所致健康风险的识别、预警和管理提供了理论参考。

Abstract:

As two efficient broad-spectrum sterilizing agents, triclosan (TCS) and triclocarban (TCC) are widely used, especially during the COVID-19 pandemic. The health risks caused by secondary pollution of TCS and TCC have aroused wide concern. Because of the similar mother nucleus structure and high lipophilicity, it remains unknown about the differences in the effect and mechanism of the toxicity (especially immunotoxicity) between TCS and TCC in organisms in the environment. In this study, we used zebrafish as a model to compare the immunotoxicity and mechanisms between the two pollutants at the same exposure concentration (0.6 µmol/L). The results showed that both TCS and TCC led to a hatching rate below 60% at the time point of 72 hours post fertilization (hpf) and the mortality rates of 40% and 50% at 120 hpf in larval zebrafish, respectively. The zebrafish exposed to TCS and TCC displayed malformations, such as shortened body, swimming sac closure, pericardial edema, yolk cyst deposition, and absorption disorder. Moreover, the developmental abnormalities caused by TCC were significantly severer than those caused by TCS. TCS exposure increased the proliferation rate of innate immune cells to 20% and decreased the number of mature T cells by 35%, while TCC exposure inhibited the differentiation of both innate immune cells and T cells, with the inhibition rates of 25% and 60%, respectively. The results of real-time quantitative PCR (RT-qPCR) and ELISA showed that TCS and TCC exposure up-regulated the expression levels of il-1β, il-6, and tnf-α, while il-10 and IgM exhibited opposite expression patterns. Additionally, both compounds slightly decreased C3 expression. The Pearson correlation analysis showed that the developmental toxicity induced by TCS and TCC had positive and negative correlations with the differentiation of immune cells, respectively. However, the toxicity induced by either TCS or TCC was positively correlated with the expression of pro-inflammatory cytokines. GO function and KEGG pathway enrichment analyses demonstrated that the target molecules of TCS and TCC were enriched in different signaling pathways, and the key network hub genes and the enriched regulatory pathways differed between TCS and TCC. The findings provide compelling evidence that TCS and TCC adopt different mechanisms in triggering immunotoxicity and offer a theoretical reference for the recognition, warning, and management of TCS and TCC-induced health risks.