Parng Chuenlei, Seng Wen Lin, Semino Carlos, McGrath Patricia
Phylonix, Cambridge, MA 02139, USA.
Assay Drug Dev Technol. 2002 Nov;1(1 Pt 1):41-8. doi: 10.1089/154065802761001293.
The zebrafish embryo has become an important vertebrate model for assessing drug effects. It is well suited for studies in genetics, embryology, development, and cell biology. Zebrafish embryos exhibit unique characteristics, including ease of maintenance and drug administration, short reproductive cycle, and transparency that permits visual assessment of developing cells and organs. Because of these advantages, zebrafish bioassays are cheaper and faster than mouse assays, and are suitable for large-scale drug screening. Here we describe the use of zebrafish bioassays for assessing toxicity, angiogenesis, and apoptosis. Using 18 chemicals, we demonstrated that toxic response, teratogenic effects, and LC(50) in zebrafish are comparable to results in mice. The effects of compounds on various organs, including the heart, brain, intestine, pancreas, cartilage, liver, and kidney, were observed in the transparent animals without complicated processing, demonstrating the efficiency of toxicity assays using zebrafish embryos. Using endogenous alkaline phosphatase staining and a whole-animal enzyme assay, we demonstrated that SU5416 and flavopiridol, compounds shown to have antiangiogenic effects in mammals, inhibit blood vessel growth in zebrafish, and this bioassay is suitable for high-throughput screening using a 96-well microplate reader. We also demonstrated that in vivo acridine orange staining can be used to visualize apoptotic events in embryos treated with brefeldin A, neomycin, or caspase inhibitors. After in vivo staining, acridine orange can be extracted and quantitated using a fluorescence microplate reader, providing a screening system for agents that modulate apoptosis.
斑马鱼胚胎已成为评估药物作用的重要脊椎动物模型。它非常适合用于遗传学、胚胎学、发育学和细胞生物学研究。斑马鱼胚胎具有独特的特征，包括易于饲养和给药、繁殖周期短以及透明度高，这使得能够对发育中的细胞和器官进行视觉评估。由于这些优势，斑马鱼生物测定比小鼠测定更便宜、更快速，并且适合大规模药物筛选。在这里，我们描述了使用斑马鱼生物测定来评估毒性、血管生成和细胞凋亡。使用18种化学物质，我们证明斑马鱼的毒性反应、致畸作用和半数致死浓度（LC50）与小鼠的结果相当。在透明动物中无需复杂处理即可观察到化合物对包括心脏、大脑、肠道、胰腺、软骨、肝脏和肾脏在内的各种器官的影响，这证明了使用斑马鱼胚胎进行毒性测定的有效性。使用内源性碱性磷酸酶染色和全动物酶测定，我们证明在哺乳动物中显示具有抗血管生成作用的化合物SU5416和黄酮哌啶醇可抑制斑马鱼的血管生长，并且这种生物测定适合使用96孔微孔板读数器进行高通量筛选。我们还证明，体内吖啶橙染色可用于可视化用布雷菲德菌素A、新霉素或半胱天冬酶抑制剂处理的胚胎中的凋亡事件。体内染色后，吖啶橙可以用荧光微孔板读数器提取和定量，从而提供一种用于筛选调节细胞凋亡的试剂的系统。
