Alternative models in developmental toxicology

Abstract

In light of various pressures, toxicologists have been searching for alternative methods for safety testing of chemicals. According to a recent policy in the European Union (Regulation, Evaluation Authorisation and Restriction of Chemicals, REACH), it has been estimated that over the next twelve to fifteen years, approximately 30,000 chemicals may need to be tested for safety, and under current guidelines such testing would require the use of approximately 7.2 million laboratory animals [ Hofer et al. 2004 ]. It has also been estimated that over 80% of all animals used for safety testing under REACH legislation would be used for examining reproductive and developmental toxicity [Hofer et al., 2004]. In addition to REACH initiatives, it has been estimated that out of 5,000 to 10,000 new drug entities that a pharmaceutical company may start with, only one is finally approved by the Food and Drug Administration at a cost of over one billion dollars [ Garg et al. 2011 ]. A large portion of this cost is due to animal testing. Therefore, both the pharmaceutical and chemical industries are interested in using alternative models and in vitro tests for safety testing. This review will examine the current state of three alternative models - whole embryo culture (WEC), the mouse embryonic stem cell test (mEST), and zebrafish. Each of these alternatives will be reviewed, and advantages and disadvantages of each model will be discussed. These models were chosen because they are the models most commonly used and would appear to have the greatest potential for future applications in developmental toxicity screening and testing.

Figure 1

Differentiation of D3 mESCs to neural cells. A) D3 mESCs cultured according to Ying et al. [2003] early in the culture period (day 2) show little differentiation. B–C) As culture progresses, neural processes begin to appear. D) Numerous cells have neural processes by 14 days of culture.

Figure 2

Expression of the pluripotency markers Oct4, Nanog, and Sox2 in D3 mouse embryonic stem cells cultured for 14 days. Decreased expression over time indicates a decrease in pluripotency of the cells.

Figure 3

Expression of the neuronal lineage markers, Nestin, Sox1, Pax6, and Olig2 in differentiating mESCs. Increased expression of the marker genes indicate differentiation into cells specific for the neuronal lineage.

Figure 4

Differences between cell lines in their differentiation to osteoblasts. Cells from the E14TG2a and BK4 cell lines were cultured under identical conditions to drive differentiation toward osteoblasts. Mineralization was determined by staining with Alizarin Red S, a calcium specific stain. Top panel depicts cells grown in control media, middle panel depicts cells grown in media containing dexamethasone, and lower panel depicts cells grown in media containing vitamin D3.