The Optimization Of In Vitro Genotoxicity Test

Genotoxicity tests can detect carcinogenicity or mutagenicity ofchemicals, and play an important role in the drug safety evaluation andprediction of environmental carcinogens. To establish and optimize ashort-term in vitro test system with good sensitivity and specificity canprovide important reference basis and reduce the potential failure in theearly-stage of drug discovery. Furthermore, the metabolic activationcondition is an important factor in the test system, which will affect the testresults in the in vitro genotoxicity test.We optimized genotoxicity battery test using human lymphoblastoidcell (TK6cell). The test system combined three tests including in vitrocomet assay, micronucleus test and thymidine kinase gene mutation assay,which can detect the genotoxicity of test articles with one treatment atdifferent time points. Compared with the commonly-used rodent cells, TK6cell is human cell with a normal p53function. Moreover, the test systemshowed good sensitivity, specificity and covered three genetic endpointsincluding DNA damage, chromosome abnormality and gene mutation. Inthis study, we evaluated the genotoxicity of two Chinese herbs components-Tetrandrine (Tet) and Triptolide (TL) by the developed test system. Theresults of comet assay indicated that Tet may induce DNA damage; theresults of micronucleus test showed that TL may cause chromosomedamage, suggested both chemicals have the potential genotoxicity.Cytochrome P450subtype CYP1A1and CYP2E1are involved inmetabolic activation of many carcinogens. AHH-1cell expresses CYP1A1and h2E1V2cell expresses both CYP1A1and CYP2E1. In the present study,we employed the two cell lines to assess the genotoxicity of carcinogen –Acrylamide (AA) by performing TK mutation assay. The resultsshowed that AA exhibited stronger mutagenic effect in h2E1V2cell,suggested that CYP2E1play an important role in its metabolism.In summary, we established two optimized in vitro genotoxicity testsystem with good sensitivity, specificity and easy to operate. We evaluatedthe genotoxicity of test articles and studied the toxic mechanism.