Metabolic activation capacity by primary hepatocytes expands the applicability of the embryonic stem cell test as alternative to experimental animal testing

The Embryonic Stem Cell Test as Tool to Assess Structure-Dependent Teratogenicity: The Case of Valproic Acid

“…3A). The mean values for VPA were similar to previously reported ID 50 (D3) values obtained with the same cell line (Buesen et al , 2009; Hettwer et al , 2010; Peters et al , 2008). Consequently, the ID 50 (D3) values follow a ranking of SProP < SPenP < VPA << RPenP < RProP < 2-IP < 2-EMP (Table 1).…”

Alternative models in developmental toxicology

“…Attempts to use S9 liver fractions which contain both microsomal enzymes in addition to cytoplasmic enzymes, however, proved toxic in the WEC [Luijten et al 2008]. Primary hepatocyte cultures have also been used in conjunction with the EST to expand the applicability of the in vitro test system [Hettwer et al 2010]. The proteratogens cyclophosphamide and valpromide were selected for use and cultured with mouse embryonic stems directly as part of the EST or after preincubation with murine hepatocytes for 6 h. Preincubation of cyclophosphamide with hepatocytes resulted in a strong decrease in ID 50 concentration of differentiating cardiomyocytes.…”

Towardin vitrobiomarkers for developmental toxicity and their extrapolation to thein vivosituation

“…The addition of a bioactivation system might increase the in vitro developmental toxicity potency of a proteratogen. Hettwer et al (2010) showed that the proteratogen cyclophosphamide was 70-fold more potent in the ES-D3 cell differentiation assay of the EST when the medium with cylophosphamide was preincubated with murine hepatocytes (preconditioned medium) [70]. However, no increase in the in vitro potency was obtained for the proteratogen valpromide, using the same experimental procedure, because the levels of the active metabolite (valproic acid) in the culture medium were too low to increase the toxicity [70].…”

“…Hettwer et al (2010) showed that the proteratogen cyclophosphamide was 70-fold more potent in the ES-D3 cell differentiation assay of the EST when the medium with cylophosphamide was preincubated with murine hepatocytes (preconditioned medium) [70]. However, no increase in the in vitro potency was obtained for the proteratogen valpromide, using the same experimental procedure, because the levels of the active metabolite (valproic acid) in the culture medium were too low to increase the toxicity [70]. Furthermore, using preconditioned medium will not increase a proteratogen's potency in a test system, when the toxic bioactivation products are unstable [69].…”