Establishment of knockdown of superoxide dismutase 2 and expression of CYP3A4 cell system to evaluate drug-induced cytotoxicity

Yoshikawa, Yukitaka; Hosomi, Hiroko; Fukami, Tatsuki; Nakajima, Miki; Yokoi, Tsuyoshi

doi:10.1016/j.tiv.2009.05.024

Cited by 23 publications

(13 citation statements)

References 25 publications

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“…Dantrolene was also implicated in the generation of reactive oxygen species via cytochrome P450 reductase. However, we previously reported that CYP3A4-mediated cytotoxicity was not involved in the effects of the nitroaromatic drugs, flutamide and dantrolene, which we investigated using CYP3A4-expressing rat BRL3A cells (Yoshikawa et al, 2009), supporting our present data in Fig. 2.…”

Section: Discussionsupporting

confidence: 91%

Development of a Highly Sensitive Cytotoxicity Assay System for CYP3A4-Mediated Metabolic Activation

Hosomi

Fukami²,

Iwamura³

et al. 2011

Drug Metab Dispos

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ABSTRACT:Drug-induced hepatotoxicity, which is a rare but serious adverse reaction to a large number of pharmaceutical drugs, is sometimes associated with reactive metabolites produced by drug-metabolizing enzymes. In the present study, we constructed a cell-based system to evaluate the cytotoxicity of reactive metabolites produced by CYP3A4 using human hepatoma cells infected with an adenovirus vector expressing human CYP3A4 (AdCYP3A4). When seven hepatoma cell lines (HepG2, Hep3B, HLE, HLF, Huh6, Huh7, and Fa2N4 cells) were infected with AdCYP3A4, HepG2 cells showed the highest CYP3A4 protein expression and testosterone 6␤-hydroxylase activity (670 pmol ⅐ min ؊1 ⅐ mg ؊1 ). With the use of AdCYP3A4-infected HepG2 cells, the cytotoxicities of 23 drugs were evaluated by the 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium monosodium salt assay, and the cell viability when treated with 11 drugs (amiodarone, desipramine, felbamate, isoniazid, labetalol, leflunomide, nefazodone, nitrofurantoin, tacrine, terbinafine, and tolcapone) was significantly decreased. Moreover, the transfection of siRNA for nuclear factor erythroid 2-related factor 2 (Nrf2) to decrease the cellular expression level of Nrf2 exacerbated the cytotoxicity of some drugs (troglitazone, flutamide, acetaminophen, clozapine, terbinafine, and desipramine), suggesting that the genes regulated by Nrf2 are associated with the detoxification of the cytotoxicities mediated by CYP3A4. We constructed a highly sensitive cell-based system to detect the drug-induced cytotoxicity mediated by CYP3A4. This system would be beneficial in preclinical screening in drug development and increase our understanding of the druginduced cytotoxicity associated with CYP3A4.

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Section: Discussionsupporting

confidence: 91%

Development of a Highly Sensitive Cytotoxicity Assay System for CYP3A4-Mediated Metabolic Activation

Hosomi

Fukami²,

Iwamura³

et al. 2011

Drug Metab Dispos

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“…Recombinant adenovirus vectors provide versatile systems for efficient transfection of more than one gene. Using this technology, CYP3A4-expressing and superoxide dismutase (Yoshikawa et al 2009) or γ-glutamylcysteine synthetase (Hosomi et al 2010) knockdown rat hepatoma cells have been generated. These cell systems were proposed as useful screening tools for the prediction of metabolism-mediated hepatotoxicity produced by oxidative stress and/or generation of active metabolites by CYP enzymes.…”

Section: Alternative Models To Primary Human Hepatocytesmentioning

confidence: 99%

Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME

Hewitt²,

et al. 2013

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This review encompasses the most important advances in liver functions and hepatotoxicity and analyzes which mechanisms can be studied in vitro. In a complex architecture of nested, zonated lobules, the liver consists of approximately 80 % hepatocytes and 20 % non-parenchymal cells, the latter being involved in a secondary phase that may dramatically aggravate the initial damage. Hepatotoxicity, as well as hepatic metabolism, is controlled by a set of nuclear receptors (including PXR, CAR, HNF-4α, FXR, LXR, SHP, VDR and PPAR) and signaling pathways. When isolating liver cells, some pathways are activated, e.g., the RAS/MEK/ERK pathway, whereas others are silenced (e.g. HNF-4α), resulting in up- and downregulation of hundreds of genes. An understanding of these changes is crucial for a correct interpretation of in vitro data. The possibilities and limitations of the most useful liver in vitro systems are summarized, including three-dimensional culture techniques, co-cultures with non-parenchymal cells, hepatospheres, precision cut liver slices and the isolated perfused liver. Also discussed is how closely hepatoma, stem cell and iPS cell–derived hepatocyte-like-cells resemble real hepatocytes. Finally, a summary is given of the state of the art of liver in vitro and mathematical modeling systems that are currently used in the pharmaceutical industry with an emphasis on drug metabolism, prediction of clearance, drug interaction, transporter studies and hepatotoxicity. One key message is that despite our enthusiasm for in vitro systems, we must never lose sight of the in vivo situation. Although hepatocytes have been isolated for decades, the hunt for relevant alternative systems has only just begun.Electronic supplementary materialThe online version of this article (doi:10.1007/s00204-013-1078-5) contains supplementary material, which is available to authorized users.

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“…Ademais, segundo Lopez-Novoa et al (31) e Quiros et al (32) , a gentamicina pode interagir com a mitocôndria, ativando a via intrínseca da apoptose e interferindo com o funcionamento da cadeia respiratória, o que compromete a produção de ATP e contribui para a geração de ERs. O sulfametoxazol, outro importante fármaco utilizado no tratamento da mastite, também é conhecido por induzir a formação ERs (33)(34)(35) .…”

Section: Resultsunclassified

Avaliação De Estresse Oxidativo No Plasma De Bovinos Leiteiros Com Mastite

et al. 2018

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ResumoA mastite bovina está associada a uma resposta antibacteriana endógena mediada pela produção de espécies reativas. Contudo, o excesso de reações oxidativas pode desencadear apoptose celular agravando o quadro clínico dos animais. Neste contexto, o objetivo deste estudo foi avaliar a resposta redox no plasma de vacas leiteiras com e sem mastite submetidas ou não ao tratamento com antibioticoterapia. As vacas foram divididas em Grupo Controle (G1), vacas sem mastite; grupo G2, vacas com mastite sem tratamento com antimicrobianos; grupo G3, vacas com mastite tratadas com antibiótico. As amostras sanguíneas foram coletadas após a primeira ordenha da manhã. Foram analisados a existência de lipoperoxidação (LPO) e os níveis de proteínas carboniladas (PCs), de glutationa reduzida (GSH), de ácido ascórbico (ASA) e de ácido úrico (AU). Os animais do G3 apresentaram aumento na LPO e das PCs. Em todos os grupos, os níveis de GSH permaneceram inalterados. Os valores plasmáticos de ASA e de AU mostraram-se diminuídos nos animais dos grupos G2 e G3. Os resultados demonstraram que o tratamento com antimicrobianos parece agravar os danos oxidativos presentes na mastite bovina, reforçando a importância da busca por alternativas terapêuticas a fim de minimizar esse efeito. Palavras-chave: antibióticos; oxidação celular; vacas. AbstractBovine mastitis is associated with an endogenous antibacterial response mediated by the production of reactive species. However, excess oxidative reactions can trigger cellular apoptosis, aggravating the clinical state of the animals. In this context, the objective of this study was to evaluate the plasma redox response of dairy cows with and without mastitis submitted or not to antibiotic therapy. The cows were divided into control group (G1), cows without mastitis; group G2, cows with mastitis without antimicrobial treatment; group G3, cows with mastitis treated with antibiotics. Blood samples

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Establishment of knockdown of superoxide dismutase 2 and expression of CYP3A4 cell system to evaluate drug-induced cytotoxicity

Cited by 23 publications

References 25 publications

Development of a Highly Sensitive Cytotoxicity Assay System for CYP3A4-Mediated Metabolic Activation

Development of a Highly Sensitive Cytotoxicity Assay System for CYP3A4-Mediated Metabolic Activation

Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME

Avaliação De Estresse Oxidativo No Plasma De Bovinos Leiteiros Com Mastite

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