以短暂暴露于邻苯二甲酸二丙酯(Dipropyl Phthalate)之淡水多齿新米虾(Neocaridina denticulata)建立生态毒理基因组生物标志物

Development of Ecotoxicogenomic Biomarkers on the Freshwater Shrimp (Neocaridina denticulate) Following Short-Term Exposure to Dipropyl Phthalate

摘要: 邻苯二甲酸酯类化合物(phthalate esters; PAEs)是广泛用于塑料制品之塑化剂和造成环境污染甚为可观的內分泌干擾物質。本研究以多齿新米虾(Neocaridina denticulata)暴露于次致死剂量(50 mg/L)之一種PAE——邻苯二甲酸二丙酯(dipropyl phthalate; DPrP)后1天，利用抑制性扣减杂交法建立一套基因组指标，探讨其全面基因的表现变化。实验中共筛得71个独特的表达序列标签(expressed sequence tages; ESTs)，包括23段ESTs对应为已知功能基因及48段ESTs为未知功能。根据生理功能，已知功能基因分别与9类相关，包括代谢、呼吸、防御、核糖体、染色体、转译、传讯、视觉与结构。虾子暴露于高浓度DPrP (50 mg/L)时，3个免疫及代谢相关的基因和3个未知功能基因，共6个基因的mRNA表现量明显下降。暴露于非致死剂量1.0 mg/L时，共8个基因受到影响，包括正调节的4个已知功能和1个未知功能基因，以及3未知功能基因被负调控。虾子暴露于更低剂量0.5 mg/L后，仅4个基因受到影响，包括3个已知功能和1个未知功能基因之mRNA表现量明显上升。综合这些结果暗示，包括DPrP在內之非致死浓度PAEs污染的水域可能藉由影响多齿新米虾的生理功能而威胁其存活；此外，起源自多齿新米虾的ESTs也许可用在水生环境中其他污染物之生态毒性效应的研究。 Phthalate esters (PAEs), which are widely used in industrial chemicals that serve as important additives to impart flexibility to polyvinyl chloride resins and have become widely diffused in the environment, are considered to be endocrine disrupting chemicals (EDCs). In order to assess the toxicity of PAE to aquatic crustacean, after shrimps (Neocaridina denticulate) were short-term exposed to a sublethal concentration (50 mg/L) of dipropyl phthalate (DPrP), the differential expression genes were isolated and identified using suppression subtractive hybridization (SSH) on the samples prepared from the whole individual. There were 71 unique expressed sequence tags (ESTs) which were identified by homology with data-based sequences, including 23 ESTs corresponded to known genes and 48 ESTs with unknown function. The known genes could be divided into nine classes on the basis of physiological function: genes related to ribosomal, metabolism, immune and structural molecules, concerning the translation-related molecules, involved respiration and signaling, and responding to signaling and vision. By comparing the level of gene transcription in DPrP-treated group vs. non-treated group using semi-quantitative RT-PCR, we found that six of twelve selected genes were significantly up-regulated following exposure to DPrP at high concentration (50 mg/L), including two immune-related genes, one metabolism-related gene and 3 unknown genes. There were eight genes which are significantly responding to DPrP treatment at 1.0 mg/L, including five up-regulated genes (four known genes and one unknown gene) and three down-regulated unknown genes. After exposure to 0.5 mg/L of DPrP, only four genes (three known genes and one unknown gene) were affected and their mRNA levels increased significantly. These results suggest that N. denticulate may be harmed via the change of the globally physiological function following exposure in non-lethal PAE-polluted aquatic environment; in addition, the ESTs derived from N. denticulate can be used for studying the ecotoxicological effect of other pollutants in the aquatic environment.