Rodent studies liver assays

The assessment of metabolites often requires complex preclinical studies and there are currently no cell-based assays that fully reproduce the in vivo metabolism of the human liver, or other tissues. The rodent S9 liver extracts used as an exogenous source of metabolism are incomplete metabolic surrogates, and perhaps more relevantly here, their properties are inappropriate for handling on an automated HTS platform. 

In vitro cytotoxicity assays using isolated cells have been applied intermittently to cyanobacterial toxicity testing over several years." Cells investigated for suitability in cyanobacterial toxin assays include primary liver cells (hepatocytes) isolated from rodents and fish, established permanent mammalian cell lines, including hepatocytes, fibroblasts and cancerous cells, and erythrocytes. Earlier work suggested that extracts from toxic cyanobacteria disrupted cells of established lines and erythrocytes," but studies with purified microcystins revealed no alterations in structure or ion transport in fibroblasts or erythrocytes,... 

Recent evidence confirms that species differences can involve more than one aspect of PPARa-mediated regulation of gene expression. The insensitivity of human liver to rodent peroxisome proliferators is associated with low levels of expression of PPARa in human liver. Marked species differences in the expression of PPARa mRNA have been demonstrated between rodent and human liver, with the latter expressing 1-10% of the levels found in mouse or rat liver (Palmer et al, 1994 Tugwood et al, 1996 Palmer etal, 1998). Using a sensitive and specific immuno/DNA binding assay. Palmer et al (1998) have shown that active PPARa protein is expressed at variable concentrations in human livers. The study compared 20 different human livers and found that those with the highest levels of PPARa protein expression contained less than 10% of the level in mice. Most of the samples (13/20) contained no detectable PPARa activity, but did... 

Supporting data from studies in isolated rodent tissues and the standard geno-toxicity assays in Salmonella and other species can help inform the mechanistic interpretations. However, a risk assessor must evaluate these data with caution. Are the results from liver (where regenerative hyperplasia is most often observed) relevant to tumors that may be observed in other organs If a chemical caused tumors in liver, bladder, and lung, for example, but correlative cytotoxicity was only documented in liver, what should be concluded from the liver cytotoxicity data The data may seem solid for a cytotoxic threshold for liver tumors in rats, while kidney tumors are observed without notable renal cytotoxicity in mice, and a genotoxic metabolite is detected. In this case, a cautious risk assessor would not likely conclude that a nonlinear extrapolation is appropriate for determination of a regulatory standard, based on the mouse data. 

Perhaps the most widely used assay for mutagenic potential is the Ames test (Ames etal., 1973, 1975 Ames, 1979). In this test the compound under investigation is studied for its ability to bring about a reverse mutation in bacteria. The bacterial tester strains are selected to detect two types of mutation base-pair substitution (TA 1535, TA 100) and frameshift (TA 1537, TA 1538, TA 98). Generally, the assay is carried out in the presence and absence of an enzyme preparation (S-9) isolated from rodent liver in order to study the mutagenicity of potential metabolites. 

In contrast to in vitro tests, most in vivo assays in rodents have failed to show evidence of antimony s genotoxicity. Antimony trioxide tested negative (no evidence of clastogenicity) in the mouse bone marrow micronucleus assay and in the rat liver DNA repair assay (Elliott et al. 1998), and failed to induce micronuclei or chromosomal aberrations in the bone marrow of rats in a subchronic dosing study (Kirkland et al. 2007). 

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