Regulatory Genotoxicity Testing: A Critical Appraisal

1995 ◽  
Vol 23 (3) ◽  
pp. 352-379
Author(s):  
Robert D. Combes
Keyword(s):  
Data Interpretation ◽  
In Vitro Tests ◽  
Negative Results ◽  
Testing Strategies ◽  
Current Controversy ◽  
In Vivo Testing ◽  
Development And Validation ◽  
Genotoxicity Testing

This review considers current approaches to regulatory genotoxicity testing, focusing on how the use of animals can be further replaced, reduced and refined. The complementary roles of in vitro and in vivo testing, and the justification for using animals, are discussed in detail. Recommendations are made for improvements and further work, in the light of the considerable current controversy surrounding the composition and deployment of testing strategies, and the interpretation of the data generated, particularly for carcinogenicity prediction. The major problems are the oversensitivity of in vitro tests and the insensitivity of in vivo assays. On the basis of an analysis of some published databases, it is concluded that there is insufficient support for using in vivo genotoxicity assays for screening. Also, it is questionable whether the scientific benefits of using such assays always outweigh the costs to the animals involved. The considerable efforts being made to harmonise in vivo protocols and to develop improved methods for detecting genotoxicity are discussed. It is recommended that more emphasis be placed on characterising genotoxins in vitro, especially for mechanisms of activity, to optimise the benefits of any confirmatory animal tests.. Also, regulatory agencies are urged to require better-designed and more-scientifically sound protocols, in which animal numbers are minimised and data interpretation, particularly that of negative results, is facilitated. Lastly, in the development and validation of transgenic rodent systems, emphasis should be placed on developing protocols in which other acute toxicity and metabolism endpoints can be measured simultaneously with in vivo mutagenesis, while minimising animal numbers.

Aspiration Revisited: Prospective Evaluation of a Physiologically Pressurized Model With Animal Correlation and Broader Applicability to Filler Complications

2021 ◽  
Author(s):  
Hyoung-Jin Moon ◽  
Won Lee ◽  
Ji-Soo Kim ◽  
Eun-Jung Yang ◽  
Hema Sundaram
Keyword(s):  
Normal Saline ◽  
False Negative ◽  
Vascular Complications ◽  
Filler Injection ◽  
Negative Results ◽  
Needle Position ◽  
False Negative Results ◽  
In Vivo Testing

Abstract Background Aspiration testing before filler injection is controversial. Some believe that aspiration can help prevent inadvertent intravascular injection, while others cite false-negative results and question its value given that the needle position always changes somewhat during injection procedures. Objectives To test the relation of false-negative results to the viscosity of the material within the needle lumen and determine whether a less viscous material within the needle lumen could decrease the incidence of false-negative results. Methods In vitro aspiration tests were performed using 30-G and 27-G needle gauges, two cross-linked hyaluronic acid fillers, normal saline bags pressurized at 140 and 10 mmHg to mimic human arterial and venous pressures, and three needle lumen conditions (normal saline, air, and filler). Testing was repeated three times under each study condition (72 tests in total). For in vivo correlation, aspiration tests were performed on femoral arteries and central auricular veins in three rabbits (4–5 aspirations per site, 48 tests in total). Results In vitro and in vivo testing using 30-G needles containing filler both showed false-negative results on aspiration testing. In vitro and in vivo testing using needles containing saline or air showed positive findings. Conclusions False-negative results from aspiration testing may be reduced by pre-filling the needle lumen with saline rather than a filler. The pressurized system may help overcome challenges of animal models with intravascular pressures significantly different from those of humans. The adaptability of this system to mimic various vessel pressures may facilitate physiologically relevant studies of vascular complications.

scholarly journals A fast and reliable method for monitoring genomic instability in the model organism Caenorhabditis elegans

2021 ◽  
Author(s):  
Merle Marie Nicolai ◽  
Barbara Witt ◽  
Andrea Hartwig ◽  
Tanja Schwerdtle ◽  
Julia Bornhorst
Keyword(s):  
Caenorhabditis Elegans ◽  
Animal Experiments ◽  
Model Organism ◽  
Animal Testing ◽  
C Elegans ◽  
Testing Strategies ◽  
Genotoxic Agents ◽  
Genotoxicity Testing

AbstractThe identification of genotoxic agents and their potential for genotoxic alterations in an organism is crucial for risk assessment and approval procedures of the chemical and pharmaceutical industry. Classically, testing strategies for DNA or chromosomal damage focus on in vitro and in vivo (mainly rodent) investigations. In cell culture systems, the alkaline unwinding (AU) assay is one of the well-established methods for detecting the percentage of double-stranded DNA (dsDNA). By establishing a reliable lysis protocol, and further optimization of the AU assay for the model organism Caenorhabditis elegans (C. elegans), we provided a new tool for genotoxicity testing in the niche between in vitro and rodent experiments. The method is intended to complement existing testing strategies by a multicellular organism, which allows higher predictability of genotoxic potential compared to in vitro cell line or bacterial investigations, before utilizing in vivo (rodent) investigations. This also allows working within the 3R concept (reduction, refinement, and replacement of animal experiments), by reducing and possibly replacing animal testing. Validation with known genotoxic agents (bleomycin (BLM) and tert-butyl hydroperoxide (tBOOH)) proved the method to be meaningful, reproducible, and feasible for high-throughput genotoxicity testing, and especially preliminary screening.

scholarly journals Predicting and Testing Bioavailability of Magnesium Supplements

Nutrients ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 1663 ◽  
Author(s):  
Laura Blancquaert ◽  
Chris Vervaet ◽  
Wim Derave
Keyword(s):  
Serum Magnesium ◽  
Area Under The Curve ◽  
In Vitro Tests ◽  
In Vitro Test ◽  
Beneficial Effects ◽  
Dissolution Tests ◽  
Acute Ingestion ◽  
In Vivo Testing

Despite the presumption of the beneficial effects of magnesium supplementation, little is known about the pharmacokinetics of different magnesium formulations. We aimed to investigate the value of two in vitro approaches to predict bioavailability of magnesium and to validate this in subsequent in vivo testing. In vitro assessment of 15 commercially available magnesium formulations was performed by means of a Simulator of the Human Intestinal Microbial Ecosystem (SHIME®) and by dissolution tests. Two magnesium formulations with contrasting bioavailability prediction from both in vitro tests (best vs. worst) were selected for in vivo testing in 30 subjects. In vivo bioavailability was compared following one acute ingestion by monitoring blood magnesium concentrations up to 6 h following intake. The in vitro tests showed a very wide variation in absorption and dissolution of the 15 magnesium products. In the in vivo testing, a significant different serum magnesium absorption profile was found up to 4 h following supplement ingestion for the two supplements with opposing in vitro test results. Moreover, maximal serum magnesium increase and total area under the curve were significantly different for both supplements (+6.2% vs. +4.6% and 6.87 vs. 0.31 mM.min, respectively). Collectively, poor bioaccessibility and bioavailability in the SHIME model clearly translated into poor dissolution and poor bioavailability in vivo. This provides a valid methodology for the prediction of in vivo bioavailability and effectiveness of micronutrients by specific in vitro approaches.

scholarly journals Toxicological Evaluation of Piceatannol, Pterostilbene, and ε-Viniferin for Their Potential Use in the Food Industry: A Review

Foods ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 592
Author(s):  
Concepción Medrano-Padial ◽  
Ana Isabel Prieto ◽  
María Puerto ◽  
Silvia Pichardo
Keyword(s):  
Food Industry ◽  
Biological Activities ◽  
In Vitro Tests ◽  
Toxicological Evaluation ◽  
Cytotoxic Effects ◽  
Cancerous Cell ◽  
Toxicological Studies ◽  
Genotoxicity Testing

The application of stilbenes in the food industry is being considered because of their biological activities. Piceatannol, pterostilbene and ε-viniferin have awakened the industry’s interest. However, before they can be commercialized, we must first guarantee their safety for consumers. The present work reviews the toxicological studies performed with these stilbenes. A wide variety of studies has demonstrated their cytotoxic effects in both cancer and non-cancerous cell lines. In contrast, although DNA damage was detected by some authors, in vitro genotoxic studies on the effects of piceatannol, pterostilbene, and ε-viniferin remain scarce. None of the three reviewed substances have been evaluated using the in vitro tests required by the European Food Safety Authority (EFSA) as the first step in genotoxicity testing. We did not find any study on the toxic effects of these stilbenes in vivo. Thus, more studies are needed to confirm their safe use before they can be authorized as additive in the food industry.

scholarly journals The use of Bayesian methodology in the development and validation of a tiered assessment approach towards prediction of rat acute oral toxicity

2022 ◽  
Author(s):  
James W. Firman ◽  
Mark T. D. Cronin ◽  
Philip H. Rowe ◽  
Elizaveta Semenova ◽  
John E. Doe
Keyword(s):  
In Silico ◽  
Oral Toxicity ◽  
New Approach ◽  
Combining Data ◽  
Assessment Approach ◽  
In Vivo Testing ◽  
Degree Of Confidence ◽  
Development And Validation

AbstractThere exists consensus that the traditional means by which safety of chemicals is assessed—namely through reliance upon apical outcomes obtained following in vivo testing—is increasingly unfit for purpose. Whilst efforts in development of suitable alternatives continue, few have achieved levels of robustness required for regulatory acceptance. An array of “new approach methodologies” (NAM) for determining toxic effect, spanning in vitro and in silico spheres, have by now emerged. It has been suggested, intuitively, that combining data obtained from across these sources might serve to enhance overall confidence in derived judgment. This concept may be formalised in the “tiered assessment” approach, whereby evidence gathered through a sequential NAM testing strategy is exploited so to infer the properties of a compound of interest. Our intention has been to provide an illustration of how such a scheme might be developed and applied within a practical setting—adopting for this purpose the endpoint of rat acute oral lethality. Bayesian statistical inference is drawn upon to enable quantification of degree of confidence that a substance might ultimately belong to one of five LD50-associated toxicity categories. Informing this is evidence acquired both from existing in silico and in vitro resources, alongside a purposely-constructed random forest model and structural alert set. Results indicate that the combination of in silico methodologies provides moderately conservative estimations of hazard, conducive for application in safety assessment, and for which levels of certainty are defined. Accordingly, scope for potential extension of approach to further toxicological endpoints is demonstrated.

scholarly journals Use of Micropatterned Thin Film Nitinol in Carotid Stents to Augment Embolic Protection

2016 ◽  
Author(s):  
Mahdis Shayan ◽  
Brian T. Jankowitz ◽  
Puneeth Shridhar ◽  
Youngjae Chun
Keyword(s):  
Thin Film ◽  
Carotid Artery ◽  
Covered Stent ◽  
In Vitro Tests ◽  
Embolic Protection ◽  
Low Profile ◽  
Study Results ◽  
In Vivo Testing

Stenting is an alternative to endarterectomy for the treatment of carotid artery stenosis. However, stenting is associated with a higher risk of procedural stroke secondary to distal thromboembolism. Hybrid stents with a micromesh layer have been proposed to address this complication. We developed a micropatterned thin film nitinol (M-TFN) covered stent designed to prevent thromboembolism during carotid intervention. This innovation may obviate the need or work synergistically with embolic protection devices. The proposed double layered stent is low-profile, thromboresistant, and covered with a M-TFN that can be fabricated with fenestrations of varying geometries and sizes. The M-TFN was created in multiple geometries, dimensions, and porosities by sputter deposition. The efficiency of various M-TFN to capture embolic particles was evaluated in different atherosclerotic carotid stenotic conditions through in vitro tests. The covered stent prevented emboli dislodgement in the range of 70-96% during 30min duration tests. In vitro vascular cell growth study results showed that endothelial cell elongation, alignment and growth behaviour silhouettes significantly enhance specifically on the diamond-shape M-TFN with the dimensions of 145µm×20µm and a porosity of 32%. Future studies will require in-vivo testing. Our results demonstrate that M-TFN has a promising potential for carotid artery stenting.

A muscle-powered energy delivery system and means for chronic in vivo testing

1999 ◽  
Vol 86 (6) ◽  
pp. 2106-2114 ◽  
Author(s):  
Dennis R. Trumble ◽  
James A. Magovern
Keyword(s):  
In Vitro Tests ◽  
Access Port ◽  
Wide Range ◽  
Piston Displacement ◽  
In Vivo Testing ◽  
Implant Testing

Electrically stimulated skeletal muscle represents a potentially unlimited source of energy for the actuation of motor prostheses. Devices to harvest and deliver contractile power have proven mechanically feasible, but long-term efficacy has not been demonstrated. This report describes recent refinements in muscle energy converter (MEC) design and details the development of an implantable afterload chamber (IAC) designed to facilitate implant testing. The IAC comprises a fluid-filled bladder housed within a titanium cylinder that connects directly to the MEC. A vascular access port allows percutaneous measurement and adjustment of air pressure within the housing and provides a means both to monitor MEC function and to control hydraulic loading conditions. Data from in vitro tests show that IAC pressure mirrors changes in MEC-piston displacement over a wide range of actuation speeds and stroke lengths. Stroke lengths and actuation forces calculated from IAC pressure readings were typically found to be within 5% of measured values. This testing scheme may yield important information in regard to the ability to harness energy from in situ muscle over prolonged periods.

scholarly journals Characterization of Fibrous Wollastonite NYAD G in View of Its Use as Negative Standard for In Vitro Toxicity Tests

Minerals ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1378
Author(s):  
Dario Di Giuseppe ◽  
Valentina Scognamiglio ◽  
Daniele Malferrari ◽  
Luca Nodari ◽  
Luca Pasquali ◽  
...  
Keyword(s):  
Biological Response ◽  
Toxicity Tests ◽  
In Vitro Toxicity ◽  
In Vitro Tests ◽  
In Vivo Models ◽  
Mineral Fibre ◽  
In Vivo Testing ◽  
Positive Controls

Today, despite considerable efforts undertaken by the scientific community, the mechanisms of carcinogenesis of mineral fibres remain poorly understood. A crucial role in disclosing the mechanisms of action of mineral fibres is played by in vitro and in vivo models. Such models require experimental design based on negative and positive controls. Commonly used positive controls are amosite and crocidolite UICC standards, while negative controls have not been identified so far. The extensive characterisation and assessment of toxicity/pathogenicity potential carried out in this work indicate that the commercial fibrous wollastonite NYAD G may be considered as a negative standard control for biological and biomedical tests involving mineral fibres. Preliminary in vitro tests suggest that wollastonite NYAD G is not genotoxic. This material is nearly pure and is characterized by very long (46.6 µm), thick (3.74 µm) and non-biodurable fibres with a low content of metals. According to the fibre potential toxicity index (FPTI) model, wollastonite NYAD G is an inert mineral fibre that is expected to exert a low biological response during in vitro/in vivo testing.

Comparison of High Purity Factor IX Concentrates and a Prothrombin Complex Concentrate in a Canine Model of Thrombogenicity

1991 ◽  
Vol 66 (05) ◽  
pp. 609-613 ◽  
Author(s):  
I R MacGregor ◽  
J M Ferguson ◽  
L F McLaughlin ◽  
T Burnouf ◽  
C V Prowse
Keyword(s):  
High Purity ◽  
Time Course ◽  
Prothrombin Complex Concentrate ◽  
Degradation Products ◽  
Canine Model ◽  
Factor Ix ◽  
In Vitro Tests ◽  
Albumin Infusion

SummaryA non-stasis canine model of thrombogenicity has been used to evaluate batches of high purity factor IX concentrates from 4 manufacturers and a conventional prothrombin complex concentrate (PCC). Platelets, activated partial thromboplastin time (APTT), fibrinogen, fibrin(ogen) degradation products and fibrinopeptide A (FPA) were monitored before and after infusion of concentrate. Changes in FPA were found to be the most sensitive and reproducible indicator of thrombogenicity after infusion of batches of the PCC at doses of between 60 and 180 IU/kg, with a dose related delayed increase in FPA occurring. Total FPA generated after 100-120 IU/kg of 3 batches of PCC over the 3 h time course was 9-12 times that generated after albumin infusion. In contrast the amounts of FPA generated after 200 IU/kg of the 4 high purity factor IX products were in all cases similar to albumin infusion. It was noted that some batches of high purity concentrates had short NAPTTs indicating that current in vitro tests for potential thrombogenicity may be misleading in predicting the effects of these concentrates in vivo.

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