Zebrafish on a chip: a novel platform for real-time monitoring of drug-induced developmental toxicity.

Yinbao Li,Jianbin Pan,Zuanguang Chen,Duanping Sun,Xinchun Li,Hongzhi Yang,Beibei Zhang,Fan Yang,Lijuan Shi,Baohong Zhang

doi:10.1371/journal.pone.0094792

Abstract

Pharmaceutical safety testing requires a cheap, fast and highly efficient platform for real-time evaluation of drug toxicity and secondary effects. In this study, we have developed a microfluidic system for phenotype-based evaluation of toxic and teratogenic effects of drugs using zebrafish (Danio rerio) embryos and larvae as the model organism. The microfluidic chip is composed of two independent functional units, enabling the assessment of zebrafish embryos and larvae. Each unit consists of a fluidic concentration gradient generator and a row of seven culture chambers to accommodate zebrafish. To test the accuracy of this new chip platform, we examined the toxicity and teratogenicity of an anti-asthmatic agent-aminophylline (Apl) on 210 embryos and 210 larvae (10 individuals per chamber). The effect of Apl on zebrafish embryonic development was quantitatively assessed by recording a series of physiological indicators such as heart rate, survival rate, body length and hatch rate. Most importantly, a new index called clonic convulsion rate, combined with mortality was used to evaluate the toxicities of Apl on zebrafish larvae. We found that Apl can induce deformity and cardiovascular toxicity in both zebrafish embryos and larvae. This microdevice is a multiplexed testing apparatus that allows for the examination of indexes beyond toxicity and teratogenicity at the sub-organ and cellular levels and provides a potentially cost-effective and rapid pharmaceutical safety assessment tool.

Highlights

Pharmaceuticals that may be administered to women of childbearing age must be tested for their teratogenic and embryotoxic effects in rodent and non-rodent models [1]
Unlike mammalian embryos that grow inside their mother’s uterus and make direct observation of embryonic development difficult [6], zebrafish embryos develop externally and their body is transparent enough to allow for characterization under a common optical microscope
It was found that developmental retardation at high concentrations of Apl, such as C5, C6 and C7 groups exhibited lower hatching rates at 72 hpf (Fig. 4A). These results indicate that when the concentration of Apl is higher than C2, it may inhibit the normal development of the zebrafish embryo

Summary

Introduction

Pharmaceuticals that may be administered to women of childbearing age must be tested for their teratogenic and embryotoxic effects in rodent and non-rodent models [1]. Khoshmanesh established a microfluidic chip that docks zebrafish embryos automatically and cultures them under flow conditions [12] This chip does not allow for the timely removal of the dead embryos which can theoretically influence the development of the remaining live embryos. A multi-channel microfluidic perfusion platform was developed for culturing single zebrafish embryos and capturing live images of various tissues and organs inside the embryo [14,15,16]. Another approach for single embryo observation was developed by immobilizing individual eggs in microfabricated holes with culture solution flowing underneath [17]. We believe that the microfluidic chip can serve as an excellent pharmaceutical evaluation platform for studying aminophylline-induced toxicity and teratogenicity during early embryonic and larvae development

Materials and Methods

Results and Discussion

Conclusions