Development of an in vitro toxicological test system based on zebrafish (Danio rerio) sperm analysis

Fish egg quality can be markedly influenced by the oocyte age after ovulation. In this study, we examined the duration of oocyte ageing in the zebrafish (Danio rerio) and whether prolonged ageing is associated with the incidence of ploidy anomalies in the resulting embryos. Oocytes were incubated in vitro for 6 h post-stripping (HPS) at 26 °C and fertilized at 2-h intervals. Meanwhile, for eggs fertilized immediately after stripping, the fertilization, embryo survival, and hatching rates started at ~80%; these rates decreased to 39%, 24%, and 16%, respectively, for oocytes that had been stored for 4 h (p ˂ 0.05), and there was an almost complete loss of egg viability at 6 HPS. Furthermore, almost 90% of the embryos derived from 6-h aged oocytes died prior to hatching, and all larvae originating from 4- and 6-h aged oocytes showed malformations. The proportion of ploidy abnormal embryos was significantly greater at 4 HPS (18.5%) than at either 0 or 2 HPS (4.7% and 8.8%, respectively). The results revealed that zebrafish oocytes retained their fertilization potential for up to 2 h after stripping at 26 °C and indicated the contribution of post-ovulatory oocyte ageing in the occurrence of ploidy anomalies in the resulting embryos.

Download Full-text

Role of Phase II Enzymes in the Bioactivation of 1,4-Dichlorobenzene and 1,4-Dibromobenzene

Alternatives to Laboratory Animals ◽

10.1177/026119299602400422 ◽

1996 ◽

Vol 24 (4) ◽

pp. 603-608

Author(s):

Moreno Paolini ◽

Laura Pozzetti ◽

Renata Mesirca ◽

Andrea Sapone ◽

Paola Silingardi ◽

...

Keyword(s):

Dna Binding ◽

Phase I ◽

Phase Ii ◽

Covalent Binding ◽

Fold Increase ◽

Test System ◽

Oxidative Enzymes ◽

Transferase Activity ◽

Genetic Toxicology

The use of sodium phenobarbital (PB, CYP2B1 inducer) combined with β-naphthoflavone (β-NF, 1A1) to induce certain Phase I reactions in S9 liver fractions is a standard method for conducting short-term bioassays for genotoxicity. However, because post-oxidative enzymes are also able to activate many precarcinogens, we tested the possibility of adapting S9 liver fractions derived from Phase II-induced rodents to the field of genetic toxicology. In this study, S9 liver fractions derived from Swiss albino CD1 mice fed 7.5g/kg 2-(3)-tert-butyl-4-hydroxyanisole (BHA; a monofunctional Phase II-inducer) for 3 weeks, show a clear pattern of induction with an approximately 3.5–9.5-fold increase in glutathione S-transferase activity. In vitro DNA binding of the promutagenic agents, [14C]-l,4-dichlorobenzene (DCB) and [14C]-1,4-dibromobenzene (DBB), is mediated by such metabolic liver preparations and showed a significant increase in covalent binding capability. In some instances, enzyme activity was more elevated when compared to that obtained with traditional (Phase I-induced) S9. Together with DNA binding, the genetic response of these chemicals in the diploid D7 strain of Saccharomyces cerevisiae used as a biological test system, revealed the ability of the BHA-derived preparations to activate the promutagenic agents, as exemplified by the significant enhancement of mitotic gene-conversion (up to 5.2-fold for DCB and 3.4-fold for DBB) and reverse point mutation (up to 3.6-fold for DCB and 2.5-fold for DBB) at a 4mM concentration. This novel metabolising biosystem, with enhanced Phase II activity, is recommended together with a traditional S9, for detecting unknown promutagens in genotoxicity studies. The routine use of either oxidative or post-oxidative S9 increases the responsiveness of the test and can contribute to the identification of promutagens not detected when using traditional protocols.

Download Full-text

Intravesical Chemotherapy: Primary in Vitro Culture of MGH-UI Bladder Tumor Spheroids as a Three Dimensional Chemosensitivity Test System

The Journal of Urology ◽

10.1016/s0022-5347(17)75821-1 ◽

1987 ◽

Vol 137 (6) ◽

Author(s):

R. Knüchel ◽

F. Hofstädter ◽

D. McCarthy ◽

W. Jenkins ◽

J.R.W. Masters

Keyword(s):

In Vitro Culture ◽

Bladder Tumor ◽

Three Dimensional ◽

Test System ◽

Intravesical Chemotherapy ◽

Chemosensitivity Test ◽

Tumor Spheroids

Download Full-text

Development of an in-vitro test system for the evaluation of cyclooxygenase-2 inhibitors

Inflammation Research ◽

10.1007/s000110050436 ◽

1999 ◽

Vol 48 (3) ◽

pp. 133-138 ◽

Cited By ~ 12

Author(s):

S. Laufer ◽

P. Zechmeister ◽

T. Klein

Keyword(s):

Test System ◽

Cyclooxygenase 2 ◽

In Vitro Test ◽

Vitro Test ◽

Cyclooxygenase 2 Inhibitors

Download Full-text

Assessment of the Mutagenic Activity of Extracts of Brazilian Propolis in Topical Pharmaceutical Formulations on Mammalian CellsIn VitroandIn Vivo

Evidence-based Complementary and Alternative Medicine ◽

10.1093/ecam/nen049 ◽

2011 ◽

Vol 2011 ◽

pp. 1-7 ◽

Cited By ~ 9

Author(s):

Juliana Marques Senedese ◽

Aline Rafaela Rodrigues ◽

Michelle Andrade Furtado ◽

Viviane Dias Faustino ◽

Andresa A. Berretta ◽

...

Keyword(s):

Chromosomal Aberrations ◽

Mutagenic Activity ◽

Biological Activities ◽

Antioxidant Properties ◽

Test System ◽

Negative Control ◽

Continuous Treatment ◽

Mutagenic Potential ◽

Topical Formulations

Propolis possesses various biological activities such as antibacterial, antifungal, anti-inflammatory, anesthetic and antioxidant properties. A topically applied product based on Brazilian green propolis was developed for the treatment of burns. For such substance to be used more safely in future clinical applications, the present study evaluated the mutagenic potential of topical formulations supplemented with green propolis extract (1.2, 2.4 and 3.6%) based on the analysis of chromosomal aberrations and of micronuclei. In thein vitrostudies, 3-h pulse (G1phase of the cell cycle) and continuous (20 h) treatments were performed. In thein vivoassessment, the animals were injured on the back and then submitted to acute (24 h), subacute (7 days) and subchronic (30 days) treatments consisting of daily dermal applications of gels containing different concentrations of propolis. Similar frequencies of chromosomal aberrations were observed for cultures submitted to 3-h pulse and continuous treatment with gels containing different propolis concentrations and cultures not submitted to any treatment. However, in the continuous treatment cultures treated with the 3.6% propolis gel presented significantly lower mitotic indices than the negative control. No statistically significant differences in the frequencies of micronuclei were observed between animals treated with gels containing different concentrations of propolis and the negative control for the three treatment times. Under the present conditions, topical formulations containing different concentrations of green propolis used for the treatment of burns showed no mutagenic effect in either test system, but 3.6% propolis gel was found to be cytotoxic in thein vitrotest.

Download Full-text

A Test System for in vitro Screening Antimycobacterial Drug Candidates for MmpS5-MmpL5 Mediated Resistance

Russian Journal of Genetics ◽

10.1134/s1022795421010154 ◽

2021 ◽

Vol 57 (1) ◽

pp. 114-116

Author(s):

K. V. Shur ◽

S. G. Frolova ◽

N. I. Akimova ◽

V. N. Danilenko ◽

D. A. Maslov

Keyword(s):

Test System ◽

In Vitro Screening ◽

Drug Candidates

Download Full-text

Genotoxic and Chemopreventive Effects of Vochysia divergens Leaves (Pantanal, Brazil)

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2018/6596142 ◽

2018 ◽

Vol 2018 ◽

pp. 1-7

Author(s):

Pollyanna Francielli de Oliveira ◽

Suzana Amorim Mendes ◽

Nathália Oliveira Acésio ◽

Luis Claudio Kellner Filho ◽

Leticia Pereira Pimenta ◽

...

Keyword(s):

Protective Effect ◽

Skin Diseases ◽

Chemical Constituents ◽

Test System ◽

Negative Control ◽

Control Group ◽

Xtt Assay ◽

Vochysia Divergens

The medicinal plant Vochysia divergens is a colonizing tree species of the Pantanal, a unique and little explored wetland region in Brazil. This species is used in folk medicine as syrups and teas to treat respiratory infections, digestive disorders, asthma, scarring, and skin diseases. The objectives of this study were to evaluate the antioxidant, cytotoxic, and genotoxic potential of the ethanolic extract of Vochysia divergens leaves (VdE), as well as the influence of VdE and its major component (the flavone 3′,5-dimethoxy luteolin-7-O-β-glucopyranoside; 3′5 DL) on MMS-induced genotoxicity. The extract significantly reduced the viability of V79 cells in the colorimetric XTT assay at concentrations ≥ 39 μg/mL. A significant increase in micronucleus frequencies was observed in V79 cell cultures treated with VdE concentrations of 160 and 320 μg/mL. However, animals treated with the tested doses of VdE (500, 1000, and 2000 mg/kg b.w.) exhibited frequencies that did not differ significantly from those of the negative control group, indicating the absence of genotoxicity. The results also showed that VdE was effective in reducing MMS-induced genotoxicity at concentrations of 20, 40, and 80 μg/mL in the in vitro test system and at a dose of 15 mg/kg b.w. in the in vivo test system. Its major component 3′5 DL exerted no protective effect, suggesting that it is not responsible for the effect of the extract. The results of the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay showed that VdE was able to scavenge 92.6% of free radicals. In conclusion, the results suggest that the protective effect of VdE may be related, at least in part, to the antioxidant activity of its chemical constituents.

Download Full-text