scholarly journals In Vitro Mutagenic and Genotoxic Assessment of a Mixture of the Cyanotoxins Microcystin-LR and Cylindrospermopsin

Toxins ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 318 ◽  
Author(s):  
Leticia Díez-Quijada ◽  
Ana I. Prieto ◽  
María Puerto ◽  
Ángeles Jos ◽  
Ana M. Cameán
Keyword(s):  
Risk Evaluation ◽  
Evaluation Framework ◽  
Kinase Assay ◽  
In Vitro Tests ◽  
Food Contaminants ◽  
The Individual ◽  
Mutation Assay ◽  
Mouse Lymphoma

The co-occurrence of various cyanobacterial toxins can potentially induce toxic effects different than those observed for single cyanotoxins, as interaction phenomena cannot be discarded. Moreover, mixtures are a more probable exposure scenario. However, toxicological information on the topic is still scarce. Taking into account the important role of mutagenicity and genotoxicity in the risk evaluation framework, the objective of this study was to assess the mutagenic and genotoxic potential of mixtures of two of the most relevant cyanotoxins, Microcystin-LR (MC-LR) and Cylindrospermopsin (CYN), using the battery of in vitro tests recommended by the European Food Safety Authority (EFSA) for food contaminants. Mixtures of 1:10 CYN/MC-LR (CYN concentration in the range 0.04–2.5 µg/mL) were used to perform the bacterial reverse-mutation assay (Ames test) in Salmonella typhimurium, the mammalian cell micronucleus (MN) test and the mouse lymphoma thymidine-kinase assay (MLA) on L5178YTk± cells, while Caco-2 cells were used for the standard and enzyme-modified comet assays. The exposure periods ranged between 4 and 72 h depending on the assay. The genotoxicity of the mixture was observed only in the MN test with S9 metabolic fraction, similar to the results previously reported for CYN individually. These results indicate that cyanobacterial mixtures require a specific (geno)toxicity evaluation as their effects cannot be extrapolated from those of the individual cyanotoxins.

In Vitro Methods as a Tool in Neurotoxicity Studies

1995 ◽  
Vol 23 (4) ◽  
pp. 491-496
Author(s):  
Hanna Tähti ◽  
Leila Vaalavirta ◽  
Tarja Toimela
Keyword(s):  
Risk Evaluation ◽  
Toxicity Testing ◽  
In Vitro Models ◽  
In Vitro Tests ◽  
Industrial Chemicals ◽  
Mercury Compounds ◽  
Toxicological Data ◽  
In Vivo Tests

— There are several hundred industrial chemicals with neurotoxic potential. The neurotoxic risks of most of these chemicals are unknown. Additional methods are needed to assess the risks more effectively and to elucidate the mechanisms of neurotoxicity more accurately than is possible with the conventional methods. This paper deals with general tasks concerning the use of in vitro models in the evaluation of neurotoxic risks. It is based on our previous studies with various in vitro models and on recent literature. The induction of glial fibrillary acidic protein in astrocyte cultures after treatment with known neurotoxicants (mercury compounds and aluminium) is discussed in more detail as an important response which can be detected in vitro. When used appropriately with in vivo tests and with previous toxicological data, in vitro neurotoxicity testing considerably improves risk assessment. The incorporation of in vitro tests into the early stages of risk evaluation can reduce the number of animals used in routine toxicity testing, by identifying chemicals with high neurotoxic potential.

scholarly journals Polyphenols: From Theory to Practice

Foods ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2595
Author(s):  
Alberto Bertelli ◽  
Marco Biagi ◽  
Maddalena Corsini ◽  
Giulia Baini ◽  
Giorgio Cappellucci ◽  
...  
Keyword(s):  
Matrix Effect ◽  
Virgin Olive Oil ◽  
Extra Virgin Olive Oil ◽  
Biological Properties ◽  
Health Claims ◽  
In Vitro Tests ◽  
The Matrix ◽  
Gender Characteristics

Background: The importance of polyphenols in human health is well known; these compounds are common in foods, such as fruits, vegetables, spices, extra virgin olive oil and wine. On the other hand, the different factors that modulate the biological activity of these compounds are less well known. Conceptualization of the work: In this review we took into account about 200 relevant and recent papers on the following topics: “polyphenols bioavailability”, “polyphenols matrix effect”, “food matrix effect”, “polyphenols-cytochromes interaction”, after having reviewed and updated information on chemical classification and main biological properties of polyphenols, such as the antioxidant, anti-radical and anti-inflammatory activity, together with the tricky link between in vitro tests and clinical trials. Key findings: the issue of polyphenols bioavailability and matrix effect should be better taken into account when health claims are referred to polyphenols, thus considering the matrix effect, enzymatic interactions, reactions with other foods or genetic or gender characteristics that could interfere. We also discovered that in vitro studies often underrate the role of phytocomplexes and thus we provided practical hints to describe a clearer way to approach an investigation on polyphenols for a more resounding transfer to their use in medicine.

Effect of Ginkgo biloba extract on rat hepatic microsomal CYP1A activity: role of ginkgolides, bilobalide, and flavonols

2004 ◽  
Vol 82 (1) ◽  
pp. 57-64 ◽  
Author(s):  
I fan Kuo ◽  
Jie Chen ◽  
Thomas K.H Chang
Keyword(s):  
Ginkgo Biloba ◽  
Ginkgo Biloba Extract ◽  
Inhibitory Potency ◽  
Hepatic Microsomes ◽  
Cyp1a Activity ◽  
Ginkgo Biloba Extracts ◽  
The Individual ◽  
Vitro Effect

The present study investigated the in vitro effect of Ginkgo biloba extracts and some of the individual constituents (ginkgolides, bilobalide, and flavonols such as kaempferol, quercetin, isorhamnetin, and their glycosides) on CYP1A-mediated 7-ethoxyresorufin O-dealkylation in hepatic microsomes isolated from rats induced with β-naphthoflavone. G. biloba extract competitively inhibited CYP1A activity, with an apparent Ki value of 1.6 ± 0.4 µg/mL (mean ± SE). At the concentrations present in the G. biloba extracts, ginkgolides A, B, C, and J and bilobalide did not affect CYP1A activity, whereas kaempferol (IC50 = 0.006 ± 0.001 µg/mL, mean ± SE), isorhamnetin (0.007 ± 0.001 µg/mL), and quercetin (0.050 ± 0.003 µg/mL) decreased this activity. The monoglycosides (1 and 10 µg/mL) and diglycosides (10 µg/mL) of kaempferol and quercetin but not those of isorhamnetin also inhibited CYP1A activity. The order of inhibitory potency was kaempferol ~ isorhamnetin > quercetin, and for each of these flavonols the order of potency was aglycone >> monoglycoside > diglycoside. In summary, G. biloba extract competitively inhibited rat hepatic microsomal CYP1A activity, but the effect was not due to ginkgolides A, B, C, or J, bilobalide, kaempferol, quercetin, isorhamnetin, or the respective flavonol monoglycosides or diglycosides.Key words: bilobalide, CYP1A, cytochrome P450, Ginkgo biloba, ginkgolide, flavonol.

Comparative Effects of Platelet Suppressant Drugs on Platelet Survival Time

1975 ◽  
Author(s):  
E. Genton ◽  
J. Ellis ◽  
P. Steele
Keyword(s):  
Heart Disease ◽  
Venous Thromboembolism ◽  
Valvular Heart Disease ◽  
Laboratory Tests ◽  
Drug Effect ◽  
Platelet Reactivity ◽  
In Vitro Tests ◽  
Platelet Survival

The important role of platelets in thrombosis makes inhibitors of their reactivity potentially useful therapeutically. A number of laboratory tests have been identified which measure platelet reactivity, but it is not clear which test and which drug effect will correlate with thrombosis and thrombosis prevention. Platelet survival (SURV) correlates with thromboembolism in patients with valvular heart disease and is shortened in several other diseases. Therefore, it is of interest to identify drugs which prolong shortened SURV. Patients with arterial and venous thromboembolism and shortened SURV (51Chromium) were treated with platelet suppressants and restudied after 12 weeks. Sulfinpyrazone prolonged SURV(2.4±.04 to 3.1 ±.06 days; p < 0.001; n = 94; average ± SEM; normal, 3.7±.04 days) and 68 (72%) had some prolongation and 39 (42%) had normalization (> 3.3 days). Dipyridamole (100 mg qd) combined with aspirin (1200 mg qd) prolonged SURV (2.6±.11 to 3.2±0.12 days; p < 0.001; n = 13) and 9 of 13 (69%) had prolongation and 6 (46%) had normalization. Clofibrate altered SURV (2.6±.09 to 3.4±.14days;p < 0.001; n = 12) and 10 of 12 (83% ) had prolongation and normalization occurred in 6 (50%). Aspirin (1200 mg qd), cyproheptadine (32 mg qd) and propranolol (160 mg qd) failed to alter SURV.Thus, of drugs which alter in vitro tests of platelet reactivity, only sulfinpyrazone, dipyridamole and clofibrate improve shortened SURV.

Plk3 phosphorylates topoisomerase IIα at Thr1342, a site that is not recognized by Plk1

2008 ◽  
Vol 411 (1) ◽  
pp. 27-32 ◽  
Author(s):  
Masato Iida ◽  
Masao Matsuda ◽  
Hideya Komatani
Keyword(s):  
Cell Cycle ◽  
Kinase Assay ◽  
Cell Cycle Distribution ◽  
Physical Interaction ◽  
Topoisomerase Iiα ◽  
In Vitro Kinase Assay ◽  
A Site ◽  
Cycle Regulation

The Plk (polo-like kinase) family is involved in cell-cycle machinery. Despite the possible overlapping involvement of Plk1 and Plk3 in cell-cycle distribution, the precise role of each Plk might be different. To investigate mechanisms that may differentiate their physiological roles, we compared the substrate specificities of Plk1 and Plk3 using synthetic peptides. Among these substrate peptides, topoisomerase IIα EKT1342DDE-containing synthetic peptide was strongly phosphorylated by Plk3 but not by Plk1. By modulating the topoisomerase IIα peptide, we identified residues at positions +1, +2 and +4 as determinants of differential substrate recognition between Plk1 and Plk3. Acidic residues at positions +2 and +4 appear to be a positive determinant for Plk3 but not Plk1. Variation at position +1 appears to be tolerated by Plk3, while a hydrophobic residue at +1 is critical for Plk1 activity. The direct phosphorylation of Thr1342 of topoisomerase IIα by Plk3 was demonstrated with an in vitro kinase assay, and overexpression of Plk3 induced the phosphorylation of Thr1342 in cellular topoisomerase IIα. Furthermore, the physical interaction between Plk3 and topoisomerase IIα was also demonstrated in cells in addition to phosphorylation. These data suggest that topoisomerase IIα is a novel physiological substrate for Plk3 and that Plk1 and Plk3 play different roles in cell-cycle regulation.

Differential Roles of NMDA Receptor Subtypes NR2A and NR2B in Dendritic Branch Development and Requirement of RasGRF1

2010 ◽  
Vol 103 (4) ◽  
pp. 1758-1770 ◽  
Author(s):  
Fernando J. Sepulveda ◽  
Fernando J. Bustos ◽  
Eveling Inostroza ◽  
Felipe A. Zúñiga ◽  
Rachael L. Neve ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Receptor Subtypes ◽  
Dendritic Branch ◽  
Spinal Cord Neurons ◽  
Whole Cell Patch Clamp ◽  
Electrophysiological Recordings ◽  
Branch Formation ◽  
The Individual

N-methyl-d-aspartate receptors (NMDARs) are known to regulate axonal refinement and dendritic branching. However, because NMDARs are abundantly present as tri-heteromers (e.g., NR1/NR2A/NR2B) during development, the precise role of the individual subunits NR2A and NR2B in these processes has not been elucidated. Ventral spinal cord neurons (VSCNs) provide a unique opportunity to address this problem, because the expression of both NR2A and NR2B (but not NR1) is downregulated in culture. Exogenous NR2A or NR2B were introduced into these naturally NR2-null neurons at 4 DIV, and electrophysiological recordings at 11 DIV confirmed that synaptic NR1NR2A receptors and NR1NR2B receptors were formed, respectively. Analysis of the dendritic architecture showed that introduction of NR2B, but not NR2A, dramatically increased the number of secondary and tertiary dendritic branches of VSCNs. Whole cell patch-clamp recordings further indicated that the newly formed branches in NR2B-expressing neurons were able to establish functional synapses because the frequency of miniature AMPA-receptor synaptic currents was increased. Using previously described mutants, we also found that disruption of the interaction between NR2B and RasGRF1 dramatically impaired dendritic branch formation in VSCNs. The differential role of the NR2A and NR2B subunits and the requirement for RasGRF1 in regulating branch formation was corroborated in hippocampal cultures. We conclude that the association between NR1NR2B-receptors and RasGRF1 is needed for dendritic branch formation in VSCNs and hippocampal neurons in vitro. The dominated NR2A expression and the limited interactions of this subunit with the signaling protein RasGRF1 may contribute to the restricted dendritic arbor development in the adult CNS.

COMPUTERIZED PLATELET AGGREGOMETRY

1987 ◽  
Author(s):  
T Szabados ◽  
F Hermán ◽  
G Kollányi ◽  
P Hadházy ◽  
K Magyar
Keyword(s):  
Dose Response ◽  
In Vitro Tests ◽  
Response Curves ◽  
Dual Channel ◽  
Large Numbers ◽  
Platelet Aggregometry ◽  
Enormous Amount ◽  
Dose Response Curves ◽  
The Individual

One of the commonly used in vitro tests for assessing platelet function is a photodensitometric assay first established by Born. However the collection and analysis of aggregation data are tedious and time consuming. The single- and dual-channel aggregometers have limited utility for the analysis of large numbers of plasma samples in clinical laboratories or in studies on the mode of action of drugs.We now report on a multichannel platelet aggregome-ter system consisting of an IBM XT personal computer and three microprocessor-controlled 4 channel aggregometers. The system collects, displays and analyzes 12 different aggregation curves simultaneously, and - like other computerized systems - (1) significantly increa ses the efficiency and ease in performing the experiment, analyzing and presenting the data; (2) provides systematic storage and rapid retrieval of the data;(3) saves an enormous amount of time; (4) because of multichannel capability eliminates the effects of time-related changes in PRP on the dose-response curves.However our system has some advantages over the computerized aggregometer systems used so far: (1) since the aggregometers contain built in microprocessors they can be utilized to measure and analyse platelet aggregation without being coupled to a computer; (2) a special program helps to check the validity of the calculated parameters under visual control; (3) the individual points of the dose-response curves can be checked at any time during the experiment.

Modulation of factor VII levels by intron 7 polymorphisms: population and in vitro studies

Blood ◽  
2000 ◽  
Vol 95 (11) ◽  
pp. 3423-3428 ◽  
Author(s):  
Mirko Pinotti ◽  
Raffaella Toso ◽  
Domenico Girelli ◽  
Debora Bindini ◽  
Paolo Ferraresi ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Fold Increase ◽  
Factor Vii ◽  
The Novel ◽  
Expression Studies ◽  
Sequence Variations ◽  
Fvii Deficiency ◽  
The Individual

Abstract Previous studies have established that factor VII gene (F7) polymorphisms (5′F7 and R353Q) contribute about one-third of factor VII (FVII) level variation in plasma. However, F7 genotyping in patients with cardiovascular disease has produced conflicting results. Population and expression studies were used to investigate the role of intron 7 (IVS7 ) polymorphisms, including repeat and sequence variations, in controlling activated FVII (FVIIa) and antigen (FVIIag) levels. Genotype–phenotype studies performed in 438 Italian subjects suggested a positive relation between the IVS7 repeat number and FVII levels. The lowest values were associated with theIVS7 + 7G allele. The screening of 52 patients with mild FVII deficiency showed an 8-fold increase in frequency (8%) of this allele, and among heterozygotes for identical mutations, lower FVII levels were observed in the IVS7 + 7G carriers. This frequent genetic component participates in the phenotypic heterogeneity of FVII deficiency. The evaluation of the individual contribution of polymorphisms was assisted by the expression of each IVS7variant, as a minigene, in eukaryotic cells. The novel quantitative analysis revealed that higher numbers of repeats were associated with higher mRNA expression levels and that the IVS7 + 7Gallele, previously defined as a functionally silent polymorphism, was responsible for the lowest relative mRNA expression. Taken together, these findings indicate that the IVS7 polymorphisms contribute to the plasmatic variance of FVII levels via differential efficiency of mRNA splicing. These studies provide further elements to understand the control of FVII levels, which could be of importance to ensure the hemostatic balance under pathologic conditions.

In vitro evaluation of a novel ketolide antimicrobial agent, RU-64004.

1997 ◽  
Vol 41 (2) ◽  
pp. 454-459 ◽  
Author(s):  
C Jamjian ◽  
D J Biedenbach ◽  
R N Jones
Keyword(s):  
Moraxella Catarrhalis ◽  
In Vitro Tests ◽  
In Vitro Activity ◽  
Gram Positive ◽  
Beta Hemolytic Streptococcus ◽  
Serious Infections ◽  
Resistant Strains ◽  
Vancomycin Resistant

Ketolides, a novel macrolide subclass, possess a mode of action that is similar to that of structurally related macrolide-lincosamide-streptogramin (MLS) compounds. By using reference in vitro tests, the in vitro activity of RU-64004 was compared to those of six other MLS compounds against more than 800 clinical pathogens, including 356 gram-positive organisms. The spectrum of activity of the ketolide was most similar to that of clindamycin versus staphylococci and streptococci and superior to those of all macrolides tested against oxacillin-resistant staphylococci and vancomycin-resistant (vanA, vanB, and vanC) enterococcal isolates. The activity of the ketolide was greater than those of the macrolides, azalides, or clindamycin tested against vancomycin-susceptible enterococci (MICs at which 90% of isolates are inhibited [MIC90S], 0.25 to 4 micrograms/ml), penicillin-resistant pneumococci (MIC90, 0.25 micrograms/ml), and most beta-hemolytic streptococci. All Streptococcus pneumoniae and beta-hemolytic streptococcus strain were inhibited by ketolide concentrations of < or = 0.25 micrograms/ml. Against 165 erythromycin-resistant strains, RU-64004 inhibited (MICs, < or = 0.5 micrograms/ml) approximately one-third of staphylococci, all streptococci, and slightly more than one-half of the enterococci. Quinupristin-dalfopristin (a streptogramin combination) was active against all tested isolates with the exception of non-Enterococcus faecium enterococci, against which the ketolide exhibited greater potency (MIC50S, 0.03 to 2 micrograms/ml). The ketolide was also active against Haemophilus influenzae (MIC90, 2 micrograms/ml), Moraxella catarrhalis (MIC90, 0.12 micrograms/ml), pathogenic Neisseria spp. (MIC90, 0.5 micrograms/ml), and many gram-positive anaerobes (MIC90, 0.5 micrograms/ml). RU-64004 may enhance the role of macrolide drugs in the treatment of some serious infections caused by MLS-resistant gram-positive organisms.

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