Dong Wu, Teraoka Hiroki, Yamazaki Koji, Tsukiyama Shusaku, Imani Sumiko, Imagawa Tomohiro, Stegeman John J, Peterson Richard E, Hiraga T
Department of Toxicology, School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu 069-8501, Japan.
Toxicol Sci. 2002 Sep;69(1):191-201. doi: 10.1093/toxsci/69.1.191.
Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on local circulation and apoptosis in the midbrain were investigated in zebrafish (Danio rerio) embryos during early development. Embryos were exposed to TCDD from 24 h post fertilization (hpf) until observation, in water maintained at 28.5 degrees C. TCDD decreased blood flow in the mesencephalic vein, the only vessel perfusing the dorsal midbrain of the embryo. At 50 hpf, blood flow was maximally reduced in this vessel and gradually returned to the control level at 60 hpf. In contrast, blood flows in the trunk and in other vessels of the head of the embryo did not significantly change until 72 hpf. Furthermore, TCDD exposure caused apoptosis in the midbrain at 60 hpf, and the TCDD dose response relationship for this effect was similar to that for reduced blood flow in the mesencephalic vein at 50 hpf. The effects of TCDD on apoptosis in the midbrain, but not on blood flow, were abolished by Z-VAD-FMK, a general caspase inhibitor. TCDD effects on both endpoints were mimicked by beta-naphthoflavone (BNF), an aryl hydrocarbon receptor (AHR) agonist, and almost abolished by concomitant exposure to TCDD and alpha-naphthoflavone (ANF), an AHR antagonist. Concomitant exposure to TCDD and either an inhibitor of cytochrome P450 (CYP) (SKF525A or miconazole) or an antioxidant (N-acetylcysteine or ascorbic acid) inhibited these effects of TCDD. The incidence of apoptosis in the midbrain was inversely related to blood flow in this brain region following these various treatments and graded TCDD exposure concentrations (r = -0.91). The same range of TCDD exposure concentrations that reduced blood flow and increased apoptosis in the midbrain greatly enhanced CYP1A mRNA expression and immunoreactivity at 50 hpf in endothelial cells of blood vessels including the mesencephalic vein and the heart, but not the brain parenchyma. Taken together, these results suggest that TCDD induces apoptosis in the midbrain of the zebrafish embryo secondary to local circulation failure, which could be related to AHR activation, induction of CYP1A, and oxidative stress.
在斑马鱼（Danio rerio）胚胎发育早期，研究了2,3,7,8-四氯二苯并对二恶英（TCDD）对中脑局部循环和细胞凋亡的影响。从受精后24小时（hpf）至观察期，将胚胎置于28.5摄氏度的水中暴露于TCDD。TCDD减少了中脑静脉的血流量，中脑静脉是灌注胚胎背侧中脑的唯一血管。在50 hpf时，该血管中的血流量最大程度降低，并在60 hpf时逐渐恢复到对照水平。相比之下，直到72 hpf，胚胎躯干和头部其他血管中的血流量均未发生明显变化。此外，TCDD暴露在60 hpf时导致中脑细胞凋亡，这种效应的TCDD剂量反应关系与50 hpf时中脑静脉血流量减少的情况相似。一种通用的半胱天冬酶抑制剂Z-VAD-FMK消除了TCDD对中脑细胞凋亡的影响，但未消除对血流量的影响。TCDD对这两个终点的影响可被芳烃受体（AHR）激动剂β-萘黄酮（BNF）模拟，而同时暴露于TCDD和AHR拮抗剂α-萘黄酮（ANF）则几乎可消除这些影响。同时暴露于TCDD和细胞色素P450（CYP）抑制剂（SKF525A或咪康唑）或抗氧化剂（N-乙酰半胱氨酸或抗坏血酸）可抑制TCDD的这些作用。在这些不同处理以及分级TCDD暴露浓度后，中脑细胞凋亡的发生率与该脑区的血流量呈负相关（r = -0.91）。在50 hpf时，相同范围的TCDD暴露浓度降低了血流量并增加了中脑细胞凋亡，同时极大地增强了包括中脑静脉和心脏在内的血管内皮细胞中CYP1A mRNA的表达和免疫反应性，但未增强脑实质中的表达和反应性。综上所述，这些结果表明，TCDD继发于局部循环衰竭，诱导斑马鱼胚胎中脑的细胞凋亡，这可能与AHR激活、CYP1A诱导和氧化应激有关。
