Statement of problem. Denture stomatitis affects complete denture wearers and is frequently treated with antifungals drugs, as well as treating the denture with sodium hypochlorite. Whether the limitations of these treatments can be overcome with local hygiene protocols that do not damage the denture materials or adversely affect the patient is unclear. Purpose. The purpose of this randomized controlled trial was to evaluate the effect of denture hygiene protocols on complete denture wearers with denture stomatitis. Material and methods. For this randomized, double-blind controlled clinical trial, 108 participants were assigned to parallel groups: 0.25% sodium hypochlorite (positive control) 0.15% Triclosan, denture cleaning tablets, or denture cleaning tablets plus gingival cleaning tablets. The participants were instructed to brush the dentures and the palate and immerse the denture in the solutions. The outcomes of denture stomatitis remission, biofilm removal, decrease of microbial load (colony-forming units), and odor level of the mouth and denture were measured at baseline and after 10 days. Descriptive analyses were used for sociodemographic characterization of the participants; the Pearson chi-square test was used to compare participant frequency with different degrees of denture stomatitis. The data were not normally distributed (Shapiro-Wilks test) or homogeneous (Levene test). So, the Kruskal-Wallis and Dunn post hoc tests and Wilcoxon test were used to compare the effects of solutions and time on the variables (a=.05). Results. The frequency of the highest to lowest denture stomatitis scores was significantly different for the 0.15% Triclosan and denture cleaning tablets groups. No significant difference was found among the groups in terms of denture stomatitis scores, biofilm, or colony -forming unit count of Candida spp. or C. albicans and S. mutans; a significant reduction was found in these parameters. The 0.25% sodium hypochlorite and 0.15% Triclosan treatments caused a significant reduction in Gram-negative microorganisms; these 2 protocols, and the denture cleaning tablets showed a significant reduction in Staphylococcus spp.; all protocols had similar effects. Only the S. mutans count of the palate decreased after 10 days. The odor level of the mouth and the denture was not significantly different (P=.778). Conclusions. The evaluated protocols can be recommended for the hygiene of complete dentures, since they were effective for all the variables studied. (J Prosthet Dent 2022;128:664-73)

PURPOSE This randomized controlled trial compared four denture hygiene protocols in terms of patient satisfaction, oral health-related quality of life (OHRQoL), and salivary parameters in complete denture wearers with denture stomatitis. MATERIAL AND METHODS For this randomized, double-blind controlled clinical trial, 108 participants were assigned to soak their dentures in one of the following solutions: (1) 0.25% sodium hypochlorite (positive control), (2) 0.15% Triclosan, (3) denture disinfecting tablets, or (4) denture disinfecting tablets plus palatine mucosa brushing solution. The outcomes of patient satisfaction, OHRQoL, and salivary parameters (salivary flow rate and pH) were measured at baseline and after 10 days. Results were compared by Kruskal-Wallis, between-group by the Dunn test, and Wilcoxon tests between times (α = 0.05). RESULTS After the hygiene protocols, and when compared with baseline, the overall patient satisfaction, maxillary denture satisfaction, maxillary denture comfort, and maxillary denture retention were ameliorated. A significant improvement was noted in OHRQoL in 3 of 4 domains evaluated (orofacial pain and discomfort, masticatory discomfort and disability, and psychological disability and discomfort). The salivary flow rate (unstimulated and stimulated); and pH were not significantly affected at the times evaluated. CONCLUSIONS Complete denture wearers may feel more satisfied with their complete dentures when treated for denture stomatitis. The tested treatments lead to similar improvement in terms of patient satisfaction and OHRQoL. This article is protected by copyright. All rights reserved.

three-dimensional skin models have become an indispensable tool in current industrial and regulatory toxicity testing activities. Since the early 90’s, several models of the skin have been developed, characterized, validated and accepted as valid replacement method for animal experimentation. In addition current legislative dossiers such as REACH (Registration, Evaluation, Authorization and Restriction of Chemicals), the Cosmetics Directive and the CLP (Classification, Labeling and Packaging) directive continue to stimulate the implementation of skin models in general. Part 1 of this publication will provide an update of the different commercially available skin models in terms of their basic characteristics and testing applications. Part 2, to be published later, will focus in detail on their use for dermal toxicity testing applications including corrosion, skin irritation, phototoxicity, dermal absorption, skin sensitization and genotoxicity testing.

In the last decades significant regulatory attempts were made to replace, refine and reduce animal testing to assess the risk of consumer products for the human eye. As the original in vivo Draize eye test is criticized for limited predictivity, costs and ethical issues, several animal-free test methods have been developed to categorize substances according to the global harmonized system (GHS) for eye irritation. This review summarizes the progress of alternative test methods for the assessment of eye irritation. Based on the corneal anatomy and current knowledge of the mechanisms causing eye irritation, different ex vivo and in vitro methods will be presented and discussed with regard to possible limitations and status of regulatory acceptance. In addition to established in vitro models, this review will also highlight emerging, full thickness cornea models that might be suited to predict all GHS categories.

Today, engineered three-dimensional tissue models from human cells of the skin, cornea, mucosa, and respiratory tract are widely used in pharmaceutical, chemical, consumer, and cosmetic industries as alternatives to animal testing. The currently available scientifically valid 3D tissue-engineered models, however, are protected by patents and/or are being commercialized using proprietary tissue culture procedures owned by only a few private companies. As a consequence, their continued commercial availability might not be guaranteed in the long-term. In collaboration with its partners, the ALEXANDRA Association strives to ensure a long-term, sustainable, and global implementation of alternative methods based on nonprofit, patent-free, and proprietary-free development of “open source” (OS) human tissue engineering technologies. By doing so, the association proposes a new standard in 3D tissue reconstruction as a basis for animal-free testing method implementation by supporting the development, production, validation, and technology transfer based on a global OS concept.

of SLS in marketed toothpastes would alter the morphology and biochemical properties (viability and inflammatory cytokine release) of human 3D cell cultures.Since tissue cell cultures are generally recognised in the literature as being a sensitive method of assaying the cytotoxic potential of test materials [16][17][18], the objective of this experiment was to measure the toxicity of 5 marketed toothpastes on a reconstituted human oral and gingival epithelium model.The working hypothesis was that toothpastes containing SLS showed more negative effects on cell cultures. Materials and MethodsA complete synopsis of the experimental protocol is shown as flowchart in Figure 1. Test materialsToothpastes were blinded and labelled as A, B, C, D and E (Table 1).Toothpastes A, D and E contained SLS while toothpastes B and C did not.They all did contain fluoride.A detailed listing of the ingredients in each toothpaste is listed in Table 1.In order to simulate the oral condition (dilution by saliva), toothpastes were mixed with water in 1:2 ratios to produce a 30% toothpaste slurry.As positive control, SLS

Natalie Alépée 1, Anthony Bahinski 2, Mardas Daneshian , Bart De Wever , Ellen Fritsche 5, Alan Goldberg 6, Jan Hansmann 7, Thomas Hartung 3,6, John Haycock 8, Helena T. Hogberg 6, Lisa Hoelting 9, Jens M. Kelm 10, Suzanne Kadereit 9, Emily McVey 11, Robert Landsiedel 12, Marcel Leist 3,9, Marc Lübberstedt 13, Fozia Noor 14, Christian Pellevoisin 1, Dirk Petersohn 15, Uwe Pfannenbecker 16, Kerstin Reisinger 15, Tzutzuy Ramirez 12, Barbara Rothen-Rutishauser 17, Monika Schäfer-Korting 18, Katrin Zeilinger 13 and Marie-Gabriele Zurich 19,20 l’Oréal R&I, Aulnay, France; Wyss Institute for Biologically Inspired engineering, Harvard University, Boston, USA; Center for Alternatives to Animal testing – europe (CAAt-europe), University of Konstanz, Konstanz, Germany; the Alexandra Association, Monaco; leibniz Research Institute for environmental Medicine, Duesseldorf, Germany; Center for Alternatives to Animal testing (CAAt), Johns Hopkins University, Bloomberg School of Public Health, Baltimore, USA; Department of tissue engineering and Regenerative Medicine, University Hospital Wuerzburg, Wuerzburg, Germany; Department of Materials Science of Engineering, University of Sheffield, Sheffield, UK; Doerenkamp-Zbinden Chair of in vitro toxicology and Biomedicine, University of Konstanz, Konstanz, Germany; InSphero AG, Schlieren, Switzerland; Board for the Authorization of Plant Protection Products and Biocides, Wageningen, the Netherlands; BASF Se, experimental toxicology and ecology, ludwigshafen, Germany; Bioreactor Group, Berlin-Brandenburg Center for Regenerative therapies (BCRt), Charité Campus Virchow-Klinikum, Berlin, Germany; Biochemical engineering, Saarland University, Saarbruecken, Germany; Henkel AG & Co KGaA, Duesseldorf, Germany; Beiersdorf AG, Hamburg, Germany; Adolphe Merkle Institute, University of Fribourg, Switzerland; Institute for Pharmacy (Pharmacology and toxicology), Freie Universität Berlin, Berlin, Germany; Department of Physiology, Faculty of Biology and Medicine, University of lausanne, lausanne, Switzerland; Swiss Center for Applied Human toxicology (SCAHt), University of lausanne, lausanne, Switzerland

For more than two decades, scientists have been trying to replace the regulatory in vivo Draize eye test by in vitro methods, but so far only partial replacement has been achieved. In order to better understand the reasons for this, historical in vivo rabbit data were analysed in detail and resampled with the purpose of (1) revealing which of the in vivo endpoints are most important in driving United Nations Globally Harmonized System/European Union Regulation on Classification, Labelling and Packaging (UN GHS/EU CLP) classification for serious eye damage/eye irritation and (2) evaluating the method’s within-test variability for proposing acceptable and justifiable target values of sensitivity and specificity for alternative methods and their combinations in testing strategies. Among the Cat 1 chemicals evaluated, 36–65 % (depending on the database) were classified based only on persistence of effects, with the remaining being classified mostly based on severe corneal effects. Iritis was found to rarely drive the classification (<4 % of both Cat 1 and Cat 2 chemicals). The two most important endpoints driving Cat 2 classification are conjunctiva redness (75–81 %) and corneal opacity (54–75 %). The resampling analyses demonstrated an overall probability of at least 11 % that chemicals classified as Cat 1 by the Draize eye test could be equally identified as Cat 2 and of about 12 % for Cat 2 chemicals to be equally identified as No Cat. On the other hand, the over-classification error for No Cat and Cat 2 was negligible (<1 %), which strongly suggests a high over-predictive power of the Draize eye test. Moreover, our analyses of the classification drivers suggest a critical revision of the UN GHS/EU CLP decision criteria for the classification of chemicals based on Draize eye test data, in particular Cat 1 based only on persistence of conjunctiva effects or corneal opacity scores of 4. In order to successfully replace the regulatory in vivo Draize eye test, it will be important to recognise these uncertainties and to have in vitro tools to address the most important in vivo endpoints identified in this paper.

Integrated approaches using different in vitro methods in combination with bioinformatics can (i) increase the success rate and speed of drug development; (ii) improve the accuracy of toxicological risk assessment; and (iii) increase our understanding of disease. Three-dimensional (3D) cell culture models are important building blocks of this strategy which has emerged during the last years. The majority of these models are organotypic, i.e., they aim to reproduce major functions of an organ or organ system. This implies in many cases that more than one cell type forms the 3D structure, and often matrix elements play an important role. This review summarizes the state of the art concerning commonalities of the different models. For instance, the theory of mass transport/metabolite exchange in 3D systems and the special analytical requirements for test endpoints in organotypic cultures are discussed in detail. In the next part, 3D model systems for selected organs--liver, lung, skin, brain--are presented and characterized in dedicated chapters. Also, 3D approaches to the modeling of tumors are presented and discussed. All chapters give a historical background, illustrate the large variety of approaches, and highlight up- and downsides as well as specific requirements. Moreover, they refer to the application in disease modeling, drug discovery and safety assessment. Finally, consensus recommendations indicate a roadmap for the successful implementation of 3D models in routine screening. It is expected that the use of such models will accelerate progress by reducing error rates and wrong predictions from compound testing.

Measurement of ocular irritancy is a necessary step in the safety evaluation of both industrial and consumer products. Assessment of the acute eye irritation potential is therefore part of the international regulatory requirements for testing of chemicals. The Bovine Corneal Opacity and Permeability (BCOP) assay is generally accepted as a valid in vitro alternative method to the Draize eye irritation test to detect corrosive and severe eye irritants (category 1), but has not proven sensitive enough to discriminate accurately moderate (category 2A/2B) to mild and non-irritating compounds. In the currently accepted BCOP assay, opacity is determined by the amount of light transmission through the cornea, and permeability is determined by the amount of sodium fluorescein dye that passes through all corneal cell layers. Both measurements are used to assign an In Vitro Irritancy Score (IVIS) for prediction of the in vivo ocular irritation potential of a test substance. Nowadays, opacity is measured by an OP-KIT opacitometer providing a center-weighted reading of light transmission by measuring changes in voltage when the transmission of white light passes through the cornea alters. As a consequence, this may underestimate opacity that develops as spots or heterogeneous opaque areas on the periphery of an isolated cornea. A prototype of a laser light-based opacitometer (PLLBO) allowing better measurement of opacities was developed by Van Goethem et al. (2010). This new device showed improved sensitivity to detect subtle changes in corneal transparency. Furthermore, the new opacitometer allowed the analysis of the complete corneal surface and was able to detect more efficiently opaque spots located along the sides of the excised corneas. A further improved prototype of the PLLBO was constructed in combination with a camera and a speckle noise reducer. Treatment conditions of the corneas in the cornea holders were optimized in order to mimic more the real in vivo situation. A set of test compounds with irritancy potencies especially in the mild and moderate range was tested. The improved LLBO showed some promising features which potentially could improve the usefulness of the BCOP test. Adaptation of cornea holders showed to be of limited value and only restricted to concentrations up to 15% which mimics more test conditions in industry. This 3-year research project was sponsored by the Stavros Niarchos Foundation (Greece).

At the IVTIP (in vitro testing industrial platform) meeting of November 26th 2009 entitled 'Toxicology in the 21st century ('21C')--working our way towards a visionary reality' all delegates endorsed the emerging concept of the '21C' vision as the way forward to enable a thorough, reliable and systematic approach to future toxicity testing without the use of animals. One of the emerging concepts focused on integrating a defined number of tests modelling in vivo-relevant and well-characterised toxicity pathways representing mechanistic endpoints. At this meeting the importance of Integrated Testing Strategies (ITS) as tools towards reduction and eventually replacement of the animals currently used for hazard identification and risk assessment was recognised. A follow-up IVTIP Spring 2010 meeting entitled 'Integrated In Vitro Testing Strategies (ITS)--Implementation Challenges' was organised to address pending questions about ITS. This report is not a review of the ITS literature, but a summary of the discussions triggered by presented examples on how to develop and implement ITS. Contrasts between pharmaceutical and chemical industry, as well as a list of general but practical aspects to be considered while developing an ITS emerged from the discussions. In addition, current recommendations on the validation of ITS were discussed. In conclusion, the outcome of this workshop improved the understanding of the participants of some important factors that may impact the design of an ITS in function of its purpose (e.g., screening, or early decision making versus regulatory), the context in which they need to be applied (e.g., ICH guidelines, REACH) and the status and quality of the available tools. A set of recommendations of best practices was established and the importance of the applicability of the individual tests as well as the testing strategy itself was highlighted.

OBJECTIVE:Candida colonization is a consequence of orthodontic treatment and can lead to oral candidosis as a complication of maxillary removable appliance treatment. During orthodontic treatment, it is important to minimize colonization to prevent active infection that could consequently interfere with treatment. Hygiene is the most important factor in managing colonization; in this study, the efficacy of NitrAdine to reduce Candida was tested.METHOD AND MATERIALS:A randomized, double-blind, placebo-controlled trial was performed. Ninety-two patients 11 to 14 years of age were recruited at the Children's and the University Dental Clinics at Mater Dei Hospital, Tal-Qroqq, Msida, Malta. Forty-four patients used the product with NitrAdine, while 48 patients used a placebo. Sampling employing the imprint technique was performed before and after the product was used. Brilliance Candida agar was used for cultures and identification. Further identification was performed using Auxacolor 2 when required.RESULTS:The control group had a statistically significant increase in Candida during treatment, while the experimental group had a nonstatistically significant decrease.CONCLUSION:It was concluded that NitrAdine may reduce the Candida burden in maxillary removable appliances. Larger sample sizes are needed to achieve statistical significance.

In November 2009 the In Vitro Testing Industrial Platform (IVTIP) organized a meeting entitled 'Toxicology in the 21st century - working our way towards a visionary reality'. Participating delegates included scientists, key opinion leaders, developers and users of 3Rs-related tests and testing strategies. This paper summarizes the discussions with respect to the conditions required to move the vision towards an applicable reality. It should not be considered as a comprehensive review of technologies that could be relevant for moving the in vitro testing and risk assessment field forward.Overall, the US National Research Council (NRC) vision and strategy for toxicity testing in the 21st century was unanimously considered as the right approach to enable future toxicity testing without animal experimentation. Many elements of this vision were identified in the European initiatives aimed at the development of non-animal based methods. However, the need for concerted actions moving the current state-of-the-art towards a thorough, reliable and systematic approach to future toxicity testing was made evident by the discussions.Among the difficulties and hurdles on the way forward, the lack of physiologically relevant, metabolic competent and robust in vivo, ex vivo and in vitro models of both healthy and diseased people was frequently mentioned. In addition, there was a call for immediate implementation of emerging technologies and paradigms considered to be essential for transferring the vision into the reality of a toxicity-testing system assessing biologically significant perturbations in key pathways which are relevant for human biology. While the unique strengths of each of the available and emerging technologies was recognized, integration of available data and emerging technologies to integrated testing strategies (ITS) was highlighted as the preferred way forward. Method harmonization and standardization, as well as procedures and guidelines for putting together ITS, were urgently requested in order to facilitate proper implementation and acceptance.There was an urgent call for better coordination of the efforts that are ongoing or initiated in the 3Rs arena at national and international level. Education, training, communication and dissemination were addressed. It was recognised that the EPAA, through its 'Platform for Communication and Dissemination', has a very important and central role in this area. (C) 2011 Elsevier Ltd. All rights reserved.

OBJECTIVE: This study evaluated the efficacy of NitrAdineTM-based disinfecting cleaning tablets for complete denture, in terms of denture biofilm removal and antimicrobial action. MATERIAL AND METHODS: Forty complete denture wearers (14 men and 26 women) with a mean age of 62.3±9.0 years were randomly assigned to two groups and were instructed to clean their dentures according to two methods: brushing (control) - 3 times a day with denture brush and tap water following meals; brushing and immersion (Experimental) - brushing the denture 3 times a day with denture brush and tap water following meals and immersion of the denture in NitrAdineTM-based denture tablets (Medical InterporousTM). Each method was used for 21 days. Denture biofilm was disclosed by a 1% neutral red solution and quantified by means of digital photos taken from the internal surface before and after the use of the product. Microbiological assessment was conducted to quantify Candida sp. RESULTS: An independent t-test revealed a significant lower biofilm percentage for the experimental group (4.7, 95% CI 2.4 to 7.9) in comparison with the control group (mean 37.5, 95% CI 28.2 to 48.1) (t38=7.996, p<0.001). A significant reduction of yeast colony forming units could be found after treatment with Medical InterporousTM denture tablets as compared to the control group (Mann-Whitney test, Z=1.90; p<0.05). CONCLUSION: The present findings suggest that NitrAdineTM-based disinfecting cleaning tablets are efficient in removal of denture biofilm. In addition, a clear antimicrobial action was demonstrated. Therefore, they should be recommended as a routine denture maintenance method for the prevention of the development of microbial biofilm induced denture stomatitis.

In order to analyze the metabolism of the extracellular matrix in vitro, we have developed a multilayered full-thickness skin equivalent. Using immunohistological and PCR techniques we investigated the expression of elastin, fibrillin-1 and fibulin-5 (DANCE), the main components of the elastic system, as well as of MMPs in the dermal tissue of the skin models.

Irritation and other forms of local toxicity following contact with eyes is a potentially serious problem arising from occupational exposure to chemicals. Traditionally, evaluation of the irritant potential of novel chemicals has relied on the use of in vivo studies with rabbits. Concerns about the predictive potential of in vivo methods for human hazard and demand for economical and rapid screening of chemicals has stimulated a great deal of work to investigate in vitro alternatives for evaluating ocular irritation potential. This publication describes a screening study to assess a reconstituted corneal epithelial culture system, as an alternative for testing for ocular irritation with pharmaceutical process materials, extending the chemical domain with which this system has been tested. A total of 21 test chemicals were applied to commercially supplied reconstituted human corneal epithelial (HCE) cultures and effects on tissue viability (MTT reduction assay), tissue histology and IL-alpha expression were assessed. Positive controls (0.5% and 1% SDS) showed dose- and time-related adverse effects on tissues, consistent with known irritant effects. Negative controls showed no histological changes and retained high viability throughout the time-course of the experiment. Concordance was excellent with accuracy at each sampling time point of over 80% when viability (MTT reduction) was compared with existing EU classification of the test articles for ocular irritation (classification based on results of in vivo evaluation). Tissue viability as estimated by MTT reduction appears most useful as the primary means of assessing the irritation potential of the chemicals. Histopathological examination generally agreed with the results of the MTT assay. However, the use of cytokine analysis will need further consideration as results for this parameter showed no relationship with known irritation potential. These results infer that HCE cultures, alone or as a part of a tiered hazard screening programme, have promise for use in reducing reliance on live subject tests and contribute to generation of an appropriate hazard classification and label advice.

The desire for a tanned appearance along with increasing awareness of the hazards of ultraviolet (UV) light exposure has generated a renewed interest in artificial tanning products. Better formulations of sunless or self-tanners with improved aesthetics are more widely available. As consumer experience with the newer products has grown, this category has become more popular, and there has been an increasing proportion of overall sun care sales. Over 20% of young adults in both United States and Australia reported using these products in the preceding year (1,2). Individual users were also more likely to have sunburned consistent with higher use of these products in fairer Caucasians. In other studies, exclusive users of sunless tanners were more likely to practice overall sun protection (3,4) and decrease their use of tanning beds (5).