Status of animal experimentation in nutrition and dietetic research: Policies of 100 leading journals and new approach methodologies.

Given animal research is challenged with inadequacies, e.g., animal-to-human knowledge translation, ethical considerations, and cost:benefit, new approach methodologies (NAMs) have been proposed as a replacement. With reference to the field of nutrition and dietetics, our aim was to examine the policies of its leading journals regarding human-based vs. traditional animal-based research; and to explore emerging NAMs that provide alternatives to animal experimentation. We reviewed 100 leading journals from an established database (SCImago Journal Rankings) in the nutrition and dietetics category for the year 2022. Eighty-three journals met the inclusion criteria. NAMs were extracted from a range of established sources. 9.6% (n = 8) of journals state they do not publish animal-based studies; 4.8% (n = 4) consider animal studies with qualifications, whereas the remaining 85.5% (n = 71) publish animal studies without qualification. Across sources, NAMs commonalities were identified including in vitro, in chemico, and in silico methods; and individual and population-based studies. Of leading nutrition/dietetic journals, relatively few have shifted to strictly non-animal methods. Greater attention to the increasing range of NAMs may not only reduce the need for animal research in the field, but may provide superior human-relevant outcomes. Studies are needed to establish their potential superiority.

Why Not Phase Out Animal Experimentation? Considering Objections from Freedom of Inquiry and Cross-Border Displacement.

Animal experimentation raises value conflicts between animal protection and other goods, such as freedom of inquiry or health and safety. If governments can phase out the practice by non-prohibitive incentive-setting, the pro tanto moral rationale for doing so is obvious. So why should they not? This article first sketches a fictional scenario in which a government adopts a phase-out plan for animal experimentation. It then considers two moral objections to this plan: First, the plan unduly restricts freedom of inquiry, and second, it merely displaces animal experimentation across borders and thus fails to reduce animal suffering. Both arguments are refined premise by premise to articulate their strongest versions. The two objections can help to narrow down desiderata for good phase-out plans. However, they do not provide a compelling case against phase-out planning as such because they miss its incremental and constructive nature. Unless better arguments can be provided, it appears that government inaction on phasing out animal experimentation lacks moral justification.

Beyond Anthropocentrism: The Moral and Strategic Philosophy behind Russell and Burch's 3Rs in Animal Experimentation.

The 3Rs framework in animal experimentation- "replace, reduce, refine" - has been alleged to be expressive of anthropocentrism, the view that only humans are directly morally relevant. After all, the 3Rs safeguard animal welfare only as far as given human research objectives permit, effectively prioritizing human use interests over animal interests. This article acknowledges this prioritization, but argues that the characterization as anthropocentric is inaccurate. In fact, the 3Rs prioritize research purposes even more strongly than an ethical anthropocentrist would. Drawing on the writings of Universities Federation for Animal Welfare (UFAW) founder Charles W. Hume, who employed Russell and Burch, it is argued that the 3Rs originally arose from an animal-centered ethic which was however restricted by an organizational strategy aiming at the voluntary cooperation of animal researchers. Research purposes thus had to be accepted as given. While this explains why the 3Rs focus narrowly on humane method selection, not on encouraging animal-free question selection in the first place, it suggests that governments should (also) focus on the latter if they recognize animals as deserving protection for their own sake.

Conceptual foundations for a clarified meaning of the 3Rs principles in animal experimentation.

Russell and Burch's 1959 original definitions of the 3Rs (replacement, reduction and refinement) are widely used today as standards for the ethical use of non-human animals in research, although they have a number of limitations. Authors and institutions around the world have addressed some of these, coming up in certain cases with more accurate, functional, and up-to-date definitions. However, not only do there still remain limitations needing to be addressed, but some that have been addressed resulted in discrepancies, contradictions, and general confusion as to how best apply the 3Rs in practice. In order to clarify the meaning of the 3Rs and enable more optimal implementation of these principles in animal experimentation, this article provides a theoretical discussion for revised definitions of the original 3Rs via examination of some of their main limitations and inconsistencies. First, we offer up the original definitions as presented in the context of Russell and Burch's book The Principles of Humane Experimental Technique. Then, we examine the main limitations and present clear specifications and requirements for such revised definitions. After presenting our revised definitions, we conclude with various implications for animal welfare within the context of experimentation.

Exploring University Students' Perspectives on Animal Experimentation: Insights From a Cross-Sectional Study.

Introduction Attitudes toward animal experimentation are rapidly evolving with time. This cross-sectional study intends to assess the attitudes of university students at Eastern Mediterranean University toward animal research based on different factors and lifestyle choices. Materials and methods Stratified random sampling was used. A total of 215 participants were chosen from the Faculties of Medicine, Pharmacy, Law, and the Department of Psychology (Faculty of Arts and Sciences). An open-access, self-administered, 14-question questionnaire was used. Mann-Whitney U tests were used for score comparisons. Results The response rate was 213 (99.07%). Eighty-eight (41.31%) of the participants were male. The mean age was 21.72 ± 2.23. Mann-Whitney U tests revealed significant differences for Q4, Q10, and Q14 (p = 0.012, 0.020, and 0.016, respectively) with respect to gender. Being a pet owner significantly affected the mean scores of Q7 and Q10 (p = 0.046 and p = 0.000, respectively). Conclusion The present study reveals nuanced student attitudes toward animal experimentation, balancing concern for animal welfare with the necessity of research. Gender and pet ownership significantly influence these views. The findings underscore the need for continued education on humane and ethical research practices. Future studies should broaden the demographic scope to better understand and track these attitudes over time.

Influence of polystyrene nanoplastics on the toxicity of haloperidol to amphibians: An in vivo and in vitro approach.

Chemical pollution is a major driver for the current worldwide crisis of amphibian decline. The present study aimed to assess the influence of polystyrene nanoplastics (PS-NPLs) on the toxicity of haloperidol to aquatic life stages of amphibians, by using in vivo (tadpoles of Xenopus laevis and Pelophylax perezi) and in vitro (A6 and XTC-2 cell lines of X. laevis) biological models. Tadpoles of both species were exposed, for 96 h, to haloperidol: 0.404 to 2.05 mg l-1 (X. laevis) or 0.404 to 3.07 mg L-1 (P. perezi). The most sensitive species to haloperidol (X. laevis) was exposed to haloperidol's LC50,96h combined with two PS-NPLs concentrations (0.01 mg L-1 or 10 mg L-1); the following endpoints were monitored: mortality, malformations, body lengths and weight. In vitro cytotoxicity was assessed by exposing the two cell lines, for 72 h, to: haloperidol (0.195 to 100 mg L-1) alone and combined with 0.01 mg L-1 or 10 mg L-1 of PS-NPLs. Xenopus laevis tadpoles revealed a higher lethal and sublethal sensitivity to haloperidol than those of P. perezi, with LC50,96h of 1.45 and 2.20 mg L-1. In vitro assays revealed that A6 cell line is more sensitive haloperidol than XTC-2: LC50,72h of 13.2 mg L-1 and 5.92 mg L-1, respectively. Results also suggested a higher sensitivity of in vivo models when compared to in vitro biological. Overall, PS-NPLs did not influence haloperidol's toxicity for in vivo and in vitro biological models, except for a reduction on the incidence of malformations while increasing the lethal toxicity (at the lowest concentration) in tadpoles. These opposite interaction patterns highlight the need for a deeper comprehension of NPLs and pharmaceuticals interactions. Results suggest a low risk of haloperidol for anuran tadpoles, though in the presence of PS-NPLs the risk may be increased.

Toward Transparency on Animal Experimentation in Switzerland: Seven Recommendations for the Provision of Public Information in Swiss Law.

In Switzerland, the importance of transparency in animal experimentation is emphasized by the Swiss Federal Council, recognizing the public's great interest in this matter. Federal reporting on animal experimentation indicates a total of 585,991 animals used in experiments in Switzerland in 2022. By Swiss law, the report enables the public to learn about many aspects such as the species and degree of suffering experienced by the animals, but some information of interest to the public is missing, such as the fate of the animals at the end of the experiment (e.g., euthanized, rehomed in a private home, reused in another experiment). When it comes to animals bred in facilities but not used in experiments, further information of interest is not required to be made public according to Swiss law, for example, the number and fate of "surplus" animals (i.e., animals bred but not used in experiments for a variety of reasons such as not carrying the phenotypical properties needed). Considering that the Swiss government has a duty to provide a full accounting of animal experimentation conducted on the public's behalf, further relevant information should be disclosed. While efforts toward transparency, such as the STAAR Agreement, have been made in the scientific community, these mostly reflect the legal requirements already in force. If Switzerland is to move toward more transparency in public information on animal experimentation, an update of the legal requirements is needed. In this article, we give recommendations for Swiss law to move toward more transparency in public information on seven aspects: (1) the fate of the animals at the end of the experiment; (2) the sources of funding for animal experimentation; (3) the harm-benefit analysis performed by researchers and ethics committees to justify an experiment using animals; (4) the number of breeding/surplus animals; (5) the fate of breeding/surplus animals; (6) the harms experienced by animals in facilities; and (7) the funding of animal facilities.

Use of animals for scientific and educational purposes by Mexican institutions.

In Mexico, it is not known which institutions use animals for scientific purposes. This work reports, based on data requested from the National Institute of Transparency, Access to Information and Protection of Personal Data (INAI), the types of institutions that use animals for research and how many of these have an ethics committee. Research centres, colleges, hospitals, national institutes, technical colleges, and public universities are the types of institutions that report using animals for experimentation. Only 54% of institutions have ethics committees. Mexican institutions from 2015 to 2021 used a total of 2,112,786 animals. Mammals are the most widely used animal group. The scientific purposes for using animals depend on the type of institution that uses them. In Mexico, it is necessary to update the regulations in order to regulate the use, protection and the care of laboratory animals.

En México se desconoce cuáles son las instituciones que utilizan animales con fines científicos. Se reporta, a partir de datos solicitados al Instituto Nacional de Transparencia, Acceso a la Información y Protección de Datos Personales (INAI), los tipos de instituciones que usan animales y cuántas poseen un comité interno para el cuidado y uso de los animales de laboratorio. Los centros de investigación, colegios, hospitales, institutos nacionales, tecnológicos y universidades públicas son los tipos de instituciones que reportaron usar animales. El 54% de las instituciones poseen comités de ética. Un total de 2,112,786 animales fueron usados por instituciones del 2015 al 2021. Los mamíferos es el grupo animal más utilizado. El uso de los animales se encuentra en función del tipo de institución que los utiliza. En México, es necesario actualizar la normatividad, con el fin de regular el uso, la protección y el cuidado de los animales de laboratorio.

Prediction of the minimum amount of anti-adhesive agent required for entire intra-abdominal cavity using fluorescent dye.

PURPOSE: Studies on the appropriate amount of anti-adhesive agents for preventing postoperative adhesion are lacking. This animal study aimed to investigate the distribution of an anti-adhesive agent in the abdominal cavity and estimate the necessary amount to cover the entire cavity. METHODS: Fluorescent dye Flamma-552 was conjugated to Guardix-sol to create Guardix-Flamma, which was laparoscopically applied to the abdominal cavity of two 10-kg pigs in different amounts: 15 mL for G1 and 35 mL for G2. After 24 hours, the distribution of Guardix-Flamma was examined under the near-infrared mode of the laparoscope, and the thickness was measured in tissues from the omentum, small, and large intestine by immunohistochemistry. RESULTS: The average area of the abdominal cavity in 10 kg pigs was 2,755 cm2. Guardix-Flamma fluorescence was detected in the greater omentum, ascites in the pelvis, and right quadrant area in G1, whereas in G2, it was detected everywhere. On average, the total thickness of G1 and G2 were 12.68 ± 9.80 µm and 18.16 ± 15.57 µm, respectively. Guardix-Flamma thickness applied to the omentum, small, and large intestines of G2 were 1.31-, 1.45-, and 1.49-times thicker than those of G1, respectively, and were all statistically significant (P < 0.05). CONCLUSION: The entire abdominal cavity of the 10 kg pig was not evenly covered with 15 mL of Guardix. Although 35 mL of Guardix is sufficient to cover the same area with an average thickness of 18 µm, further studies should evaluate the minimum thickness required for an effective anti-adhesive function.

Investigation through Animal-Computer Interaction: A Proof-of-Concept Study for the Behavioural Experimentation of Colour Vision in Zoo-Housed Primates.

Zoos are an important repository of animals, which have a wide range of visual systems, providing excellent opportunities to investigate many comparative questions in sensory ecology. However, behavioural testing must be carried out in an animal welfare-friendly manner, which is practical for zoo staff. Here, we present a proof-of-concept study to facilitate behavioural research on the sensory ecology of captive primates. A system consisting of a tablet computer and an automated feeder connected wirelessly was developed and presented to captive primate species to evaluate interactions with and without previous training. A colour stimulus, analogous to the Ishihara test, was used to check the level of interaction with the device, supporting future studies on sensory ecology with zoo animals. Animals were able to use the system successfully and displayed signs of learning to discriminate between the visual stimuli presented. We identified no risk for small primates in their interactions with the experimental setup without the presence of keepers. The use of electronic devices should be approached with caution to prevent accidents, as a standard practice for environmental enrichment for larger animals (e.g., spider monkeys). In the long term, the system developed here will allow us to address complex comparative questions about the functions of different visual systems in captive animals (i.e., dichromatic, trichromatic, etc.).

Efficacy of robot arm-assisted endoscopic submucosal dissection in live porcine stomach (with video).

Endoscopic submucosal dissection (ESD) is technically challenging and requires a high level of skill. However, there is no effective method of exposing the submucosal plane during dissection. In this study, the efficacy of robot arm-assisted tissue traction for gastric ESD was evaluated using an in vivo porcine model. The stomach of each pig was divided into eight locations. In the conventional ESD (C-ESD) group, one ESD was performed at each location (N = 8). In the robot arm-assisted ESD (R-ESD) group, two ESDs were performed at each location (N = 16). The primary endpoint was the submucosal dissection speed (mm2/s). The robot arm could apply tissue traction in the desired direction and successfully expose the submucosal plane during submucosal dissection in all lesion locations. The submucosal dissection speed was significantly faster in the R-ESD group than in the C-ESD group (p = 0.005). The blind dissection rate was significantly lower in the R-ESD group (P = 0.000). The robotic arm-assisted traction in ESD enabled a significant improvement in submucosal dissection speed, blind dissection rate which suggests the potential for making ESD easier and enhancing procedural efficiency and safety.

The role of light emitting diode in wound healing: A systematic review of experimental studies.

Wounds represent a growing global issue demanding increased attention. To expedite wound healing, technologies are under development, and light emitting diode (LED) devices of varying wavelengths are being explored for their stimulating influence on the healing process. This article presents a systematic literature review aiming to compile, organize, and analyze the impacts of LED devices on wound healing. This review is registered on the PROSPERO platform [CRD42023403870]. Two blinded authors conducted searches in the Pubmed, Web of Science, Scopus, Embase, and ScienceDirect databases. In vitro and in vivo experimental studies assessing LED utilization in the wound healing process were included. The search yielded 1010 studies, of which 27 were included in the review. It was identified that LED stimulates different healing pathways, promoting enhanced cell proliferation and migration, angiogenesis stimulation, increased collagen deposition, and modulation of the inflammatory response. Thus, it can be concluded that the LED stimulates cellular and molecular processes contingent on the utilized parameters. The effects depend on the standards used. Cell migration and proliferation were better influenced by green and red LED. The extracellular matrix components and angiogenesis were regulated by all wavelengths and the modulation of inflammation was mediated by green, red, and infrared LEDs.

Peri-implantitis induction and resolution around zirconia versus titanium implants.

BACKGROUND: This study compared titanium and zirconia implant ligature-induced peri-implant defect progression and response to regenerative surgical intervention. METHODS: Eight tissue-level endosseous implants were placed in 6 mixed-breed foxhounds, with 2 zirconia and 2 titanium alternating in each hemimandible. Cotton ligatures were placed subgingivally for 16 weeks followed by 8 weeks of spontaneous progression. Standardized radiographs were captured every 2 weeks to evaluate the rate of bone loss. Regenerative surgery was performed utilizing water-jet decontamination, enamel matrix derivative, and locally harvested autogenous bone. After 16 weeks of healing, final radiographic bone levels as well as probing depths, recession, and clinical attachment levels were assessed. RESULTS: All 48 implants integrated successfully. The final average post-ligature radiographic defects were 2.88 and 3.05 mm for titanium and zirconia implants, respectively. There was no significant difference between materials in the rate of radiographic bone loss (p = 0.09). Following regenerative surgery, the total average amount of radiographic bone gain was 1.41 and 1.20 mm for titanium and zirconia, respectively. The percentage of defect fill was 51.56% and 37.98% (p = 0.03) for titanium and zirconia, respectively. Inter-group differences were minimal for clinical parameters at the time of sacrifice including periodontal pocket depths (p = 0.81), recession (p = 0.98), or clinical attachment levels (p = 0.51). CONCLUSIONS: No significant difference was found in the rate of peri-implant defect development between titanium and zirconia implants. Both materials gained significant radiographic bone following regenerative surgery with significantly greater defect percentage fill in titanium implants. The final clinical parameters were similar in both groups.

A Novel Minimally Invasive Surgically Induced Skeletal Muscle Injury Model in Sheep.

Sports-related muscle injuries account for 10-55% of all injuries, which is a growing concern, especially given the aging world population. To evaluate the process of skeletal muscle injury and compare it with muscle lesions observed in humans, we developed a novel in vivo model in sheep. In this model, muscle injury was induced by an ultrasound-guided transverse biopsy at the myotendinous junction of the medial gastrocnemius muscle. Twelve male sheep were examined at 3, 7, 14, and 28 days post-injury. Histological, immunofluorescence, and MRI analyses indicate that our sheep model could resemble key human clinicopathological features. Statistically significant differences (p < 0.05) were observed in collagen I, dMHC, α-SMA, and CD68 immunohistochemical detection when comparing injured and healthy muscles. The injured gastrocnemius muscle exhibited elevated levels of type I collagen, infiltration of CD68(+) macrophages, angiogenesis, and the emergence of newly regenerated dMHC(+) myofibers, which persisted for up to 4 weeks post-injury. Similarly, the progression of muscle injury in the sheep model was assessed using advanced clinical 3 T MRI and compared with MRI scans from human patients. The data indicate that the sheep muscle injury model presents features similar to those observed in human skeletal muscle injuries. This makes it a valuable large animal model for studying muscle injuries and developing novel therapeutic strategies.

Utility of pemafibrate in nonalcoholic steatohepatitis model mice induced by a choline-deficient, high-fat diet and dextran sulfate sodium.

Aim: The purpose of this study was to examine the effect of pemafibrate in a murine model of non-alcoholic steatohepatitis (NASH). Methods: Forty-two, 19-week-old, male, C57BL/6J mice were divided into three groups: a Control group (n = 14), a dextran sulfate sodium (DSS) group (n = 14), and a DSS + PEM group (n = 14). All mice were given a standard rodent diet for the first week, followed by a choline-deficient, high-fat diet (CDHF) for the next 12 weeks. The 22nd day after the animals arrived was taken as Day 1 of the experiment. The Control group continued the CDHF diet and MilliQ water. The DSS group continued the CDHF diet, but starting on Day 1, the group received 0.8 % DSS to drink for 7 consecutive days, followed by MilliQ water for 10 days; this was taken as one course, and it was repeated on the same schedule until autopsy. The DSS + PEM group received the CDHF diet with PEM 0.1 mg/kg/day. Their drinking water was the same as that of the DSS group. On Seven animals from each group were autopsied on each of Day 50 and Day 120, and histopathological and immunohistochemical examinations, as well as quantitative RNA and cytokine measurements, of autopsied mice were performed. Results: Pemafibrate improved hepatic steatosis (decreased steatosis area), improved liver inflammation enhanced by DSS (decreased aspartate transaminase and alanine aminotransferase), improved hepatic fibrosis promoted by DSS (decreased fibrotic areas and a marker of fibrosis), inhibited tumorigenesis, and decreased intestinal inflammation in the NASH model mice. Conclusions: In a murine model of NASH, mixing PEM 0.1 mg/kg/day into the diet inhibited disease progression and tumor formation.

The activin/BMP-2 chimera AB204 promotes periodontal tissue regeneration in a buccal dehiscence model: a pilot study.

PURPOSE: A combination of activin and bone morphogenetic protein-2 (BMP-2), termed AB204, has been shown to improve osteogenic potential with fewer side effects than BMP-2 alone. This study was performed to evaluate the effect of AB204 on periodontal tissue regeneration in a dog buccal dehiscence model. METHODS: Buccal dehiscence defects were created on the maxillary premolars (P1, P2, and P3) of 6 mongrel dogs. After 5 weeks, the dogs were randomly assigned to 1 of 3 groups: the control, collagen matrix (CM), and CM/AB204 groups. Grafting procedures were then performed. The dogs were sacrificed 8 weeks after the grafting procedure, and volumetric and histological analyses were conducted. RESULTS: The thickness of the buccal gingiva in the CM/AB204 group was greater than those in the other groups at 2 weeks (P<0.05). The ridge width in the AB204/CM group exceeded the width in the other groups at 4 and 8 weeks; however, the difference was not statistically significant. Histological analysis revealed that the CM/AB204 group demonstrated the formation of new bone surrounded by newly formed periodontal ligament and cementum (P=0.035). CONCLUSIONS: The combined application of CM and AB204 shows promise in facilitating the regeneration of periodontal attachment, including the formation of new bone, cementum, and periodontal ligament.

Enterocolitis necrotizante experimental con lipopolisacárido oral y función protectora de la lechematerna / Experimental necrotizing enterocolitis using oral lipopolysaccharide and protective role of breastmilk

Introducción: La enterocolitis necrotizante (ECN) es una enfermedad potencialmente mortal que afecta a los neonatos, y frente a laque la leche materna ha demostrado tener un papel protector. Administrando lipopolisacáridos (LPS) por vía oral en ratas recién nacidas(RRN), hemos desarrollado un modelo experimental para inducir undaño intestinal similar al que provoca la ECN con objeto de evaluarel aspecto macroscópico y microscópico del intestino, y de ese modo,analizar la presencia de ECN y estudiar el papel que desempeña laleche materna (LM). Material y métodos: Las RRN se dividieron en tres grupos: el grupoA (control, n= 10) permaneció con su madre; el grupo B (LPS, n=25)fue aislado tras el nacimiento, alimentado por sonda con una fórmulaespecial para ratas y LPS oral, y sometido a estrés (hipoxia tras sonda);y el grupo C (LM, n= 12) fue alimentado con leche materna tras elnacimiento y posteriormente aislado y sometido a estrés al igual que elgrupo B. El día 4 se sacrificó a las RRN y se recuperaron sus intestinospara su posterior evaluación. Resultados: En el grupo de control, no se observó ECN ni macroscópica ni histológicamente, mientras que los dos grupos sometidos aestrés (B y C) presentaron una incidencia global de la ECN del 73%.La mayoría de los sujetos del grupo B desarrollaron signos histológi-cos de ECN (85%), y los del grupo C registraron una incidencia de laECN estadísticamente menor (50%, p= 0,04), lo que significa que laLM desempeña una función protectora frente a la ECN (OR= 0,19; IC95%: 0,40-0,904). Conclusión: Nuestro modelo reveló una incidencia significativa dela ECN en RRN (73%), desempeñando la LM la misma función protectora que en el caso de los humanos recién nacidos, lo que significa que estemodelo experimental de ECN es fiable y reproducible. Gracias a dichologro, podremos investigar nuevos y potenciales objetivos terapéuticospara una peligrosa enfermedad que, a día de hoy, carece de tratamiento.(AU)

Introduction: Necrotizing enterocolitis (NEC) is a life-threateningcondition that afflicts neonates. Breastfeeding has demonstrated to playa protective role against it. By administering lipopolysaccharides (LPS)orally in newborn rats (NBR), we have developed an experimental modelto induce NEC-like gut damage. Our aim was to assess the macroscopicand microscopic appearance of the gut, to evaluate the presence of NECand study the role of breast milk (BM). Material and methods: NBR were divided into 3 groups: GroupA (control, n= 10) remained with the mother, group B (LPS, n= 25)was isolated after birth, gavage-fed with special rat formula and oralLPS, then submitted to stress (hypoxia after gavage) and group c (BM,n= 12) was breastfed once after birth, then isolated, and submitted tostress like group B. On day 4, NBR were sacrificed, and intestine washarvested and assessed. Results: In the control group NEC was not present either macroscopically or histologically. Both groups submitted to stress (B and C)presented a global incidence of NEC of 73%. Most of group B developedhistologic signs of NEC (85%) and group C showed a statistically lowerincidence of NEC (50%, p= 0.04), playing the BM a protective roleagainst NEC (OR= 0.19; 95% CI: 0.40- 0.904)Conclusion: Our model showed a significant incidence of NEC inNBR (73%) with the same protective role of BM as in newborn humans,achieving a reliable and reproducible experimental NEC model. This willallow us to investigate new potential therapeutic targets for a devastatingdisease that currently lacks treatment.(AU)

60 Years of the 3Rs symposium: Lessons learned and the road ahead.

When The Principles of Humane Experimental Technique was published in 1959, authors William Russell and Rex Burch had a modest goal: to make researchers think about what they were doing in the laboratory ­ and to do it more humanely. Sixty years later, their groundbreaking book was celebrated for inspiring a revolution in science and launching a new field: The 3Rs of alternatives to animal experimentation. On November 22, 2019, some pioneering and leading scientists and researchers in the field gathered at the Johns Hopkins Bloomberg School of Public Health in Bal­timore for the 60 Years of the 3Rs Symposium: Lessons Learned and the Road Ahead. The event was sponsored by the Johns Hopkins Center for Alternatives to Animal Testing (CAAT), the Foundation for Chemistry Research and Initiatives, the Alternative Research & Development Foundation (ARDF), the American Cleaning Institute (ACI), the International Fragrance Association (IFRA), the Institute for In Vitro Sciences (IIVS), John "Jack" R. Fowle III, and the Society of Toxicology (SoT). Fourteen pres­entations shared the history behind the groundbreaking publication, international efforts to achieve its aims, stumbling blocks to progress, as well as remarkable achievements. The day was a tribute to Russell and Burch, and a testament to what is possible when people from many walks of life ­ science, government, and industry ­ work toward a common goal.

William Russell and Rex Burch published their book The Principles of Humane Experimental Technique in 1959. The book encouraged researchers to replace animal experiments where it was possible, to refine experiments with animals in order to reduce their suffering, and to reduce the number of animals that had to be used for experiments to the minimum. Sixty years later, a group of pioneering and leading scientists and researchers in the field gathered to share how the publi­cation came about and how the vision inspired international collaborations and successes on many different levels including new laws. The paper includes an overview of important milestones in the history of alternatives to animal experimentation.