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1.
Artigo em Inglês | MEDLINE | ID: mdl-38821675

RESUMO

Currently, there is no test system, whether in vitro or in vivo, capable of examining all endpoints required for genotoxicity evaluation used in pre-clinical drug safety assessment. The objective of this study was to develop a model which could assess all the required endpoints and possesses robust human metabolic activity, that could be used in a streamlined, animal-free manner. Liver-on-chip (LOC) models have intrinsic human metabolic activity that mimics the in vivo environment, making it a preferred test system. For our assay, the LOC was assembled using primary human hepatocytes or HepaRG cells, in a MPS-T12 plate, maintained under microfluidic flow conditions using the PhysioMimix® Microphysiological System (MPS), and co-cultured with human lymphoblastoid (TK6) cells in transwells. This system allows for interaction between two compartments and for the analysis of three different genotoxic endpoints, i.e. DNA strand breaks (comet assay) in hepatocytes, chromosome loss or damage (micronucleus assay) and mutation (Duplex Sequencing) in TK6 cells. Both compartments were treated at 0, 24 and 45 h with two direct genotoxicants: methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS), and two genotoxicants requiring metabolic activation: benzo[a]pyrene (B[a]P) and cyclophosphamide (CP). Assessment of cytochrome activity, RNA expression, albumin, urea and lactate dehydrogenase production, demonstrated functional metabolic capacities. Genotoxicity responses were observed for all endpoints with MMS and EMS. Increases in the micronucleus and mutations (MF) frequencies were also observed with CP, and %Tail DNA with B[a]P, indicating the metabolic competency of the test system. CP did not exhibit an increase in the %Tail DNA, which is in line with in vivo data. However, B[a]P did not exhibit an increase in the % micronucleus and MF, which might require an optimization of the test system. In conclusion, this proof-of-principle experiment suggests that LOC-MPS technology is a promising tool for in vitro hazard identification genotoxicants.


Assuntos
Hepatócitos , Testes para Micronúcleos , Testes de Mutagenicidade , Mutagênicos , Humanos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Mutagênicos/toxicidade , Testes para Micronúcleos/métodos , Testes de Mutagenicidade/métodos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Dispositivos Lab-On-A-Chip , Dano ao DNA/efeitos dos fármacos , Ensaio Cometa/métodos , Ciclofosfamida/toxicidade , Metanossulfonato de Metila/toxicidade , Linhagem Celular , Benzo(a)pireno/toxicidade , Técnicas de Cocultura , Metanossulfonato de Etila/toxicidade , Mutação/efeitos dos fármacos
2.
Biotechnol Lett ; 25(12): 955-8, 2003 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12889830

RESUMO

Crude rapeseed oil and post-refining fatty acids were used as substrates for oxalic acid production by a mutant of Aspergillus niger. Both the final concentration and the yield of the product were highest at pH 4 to 5. With a medium containing 50 g lipids l(-1), production reached a maximum of 68 g oxalic acid l(-1) after 7 d. A high yield of the product (up to 1.4 g oxalic acid g(-1) lipids consumed) was achieved with oil and fatty acids combined.


Assuntos
Aspergillus niger/genética , Aspergillus niger/metabolismo , Ácidos Graxos/metabolismo , Ácido Oxálico/metabolismo , Óleos de Plantas/metabolismo , Aspergillus niger/química , Aspergillus niger/crescimento & desenvolvimento , Ácidos Graxos Monoinsaturados , Engenharia Genética/métodos , Concentração de Íons de Hidrogênio , Metabolismo dos Lipídeos , Mutação , Micélio/crescimento & desenvolvimento , Óleo de Brassica napus , Especificidade da Espécie , Especificidade por Substrato
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