Evaluation of in vitro toxicity information for zebrafish as a promising alternative for chemical hazard and risk assessment

In vitro assays are widely proposed as a test alternative to traditional in vivo standard acute and chronic toxicity tests. However, whether toxicity information derived from in vitro assays instead of in vivo tests could provide sufficient protection (e.g., 95 % of protection) for chemical risks remain evaluated. To investigate the feasibility of zebrafish (Danio rerio) cell-based in vitro test method as a test alternative, we comprehensively compared sensitivity differences among endpoints, among test methods (in vitro, FET and in vivo), and between zebrafish and rat (Rattus norvegicus), respectively using chemical toxicity distribution (CTD) approach. For each test method involved, sublethal endpoints were more sensitive than lethal endpoints for both zebrafish and rat, respectively. Biochemistry (zebrafish in vitro), development (zebrafish in vivo and FET), physiology (rat in vitro) and development (rat in vivo) were the most sensitive endpoints for each test method. Nonetheless, zebrafish FET test was the least sensitive one compared to its in vivo and in vitro tests for either lethal or sublethal responses. Comparatively, rat in vitro tests considering cell viability and physiology endpoints were more sensitive than rat in vivo test. Zebrafish was found to be more sensitive than rat regardless of in vivo or in vitro tests for each pairwise endpoint of concern. Those findings indicate that zebrafish in vitro test is a feasible test alternative to zebrafish in vivo and FET test and traditional mammalian test. It is suggesting that zebrafish in vitro test can be optimized by choosing more sensitive endpoints, such as biochemistry to provide sufficient protection for zebrafish in vivo test and to establish applications of zebrafish in vitro test in future risk assessment. Our findings are vital for evaluating and further application of in vitro toxicity toxicity information as an alternative for chemical hazard and risk assessment.