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Preclinical biomedical research is limited by the predictiveness of in vivo and in vitro models. While in vivo models offer the most complex system for experimentation, they are also limited by ethical, financial, and experimental constraints. In vitro models are simplified models that do not offer the same complexity as living animals but do offer financial affordability and more experimental freedom; therefore, they are commonly used. Traditional 2D cell lines cannot fully simulate the complexity of the epithelium of healthy organs and limit scientific progress. The One Health Initiative was established to consolidate human, animal, and environmental health while also tackling complex and multifactorial medical problems. Reverse translational research allows for the sharing of knowledge between clinical research in veterinary and human medicine. Recently, organoid technology has been developed to mimic the original organs epithelial microstructure and function more reliably. While human and murine organoids are available, numerous other organoids have been derived from traditional veterinary animals and exotic species in the last decade. With these additional organoid models, species previously excluded from in vitro research are becoming accessible, therefore unlocking potential translational and reverse translational applications of animals with unique adaptations that overcome common problems in veterinary and human medicine.

Painted turtles are remarkable for their freeze tolerance and supercooling ability along with their associated resilience to hypoxia/anoxia and oxidative stress, rendering them an ideal biomedical model for hypoxia-induced injuries (including strokes), tissue cooling during surgeries, and organ cryopreservation. Yet, such research is hindered by their seasonal reproduction and slow maturation. Here we developed and characterized adult stem cell-derived turtle liver organoids (3D self-assembled in vitro structures) from painted, snapping, and spiny softshell turtles spanning ~175My of evolution, with a subset cryopreserved. This development is, to the best of our knowledge, a first for this vertebrate Order, and complements the only other non-avian reptile organoids from snake venom glands. Preliminary characterization, including morphological, transcriptomic, and proteomic analyses, revealed organoids enriched in cholangiocytes. Deriving organoids from distant turtles and life stages demonstrates that our techniques are broadly applicable to chelonians, permitting the development of functional genomic tools currently lacking in herpetological research. Such platform could potentially support studies including genome-to-phenome mapping, gene function, genome architecture, and adaptive responses to climate change, with implications for ecological, evolutionary, and biomedical research.Multi-omic profiling on 3D liver organoids derived from multiple species may help foster future research on unique turtle adaptations, along with their evolutionary and biomedical implications. [ABSTRACT FROM AUTHOR]

For decades, two-dimensional (2D) cell cultures have been used as the gold standard in vitro models for basic research, drug discovery and preclinical screening applications. However, since 2009, the field has quickly shifted towards the use of three-dimensional (3D) stem cell-derived organoids. Organoids offer a variety of advantages compared with conventional 2D models, including being selforganized, retaining their cellular polarity, and being simplified versions of organs in vitro. Adult stem cells can self-renew, differentiate into multiple cell types, and are genomically stable over multiple passage lines, providing a variety of technical benefits and offering the closest recapitulation of the in vivo environment. Our laboratory has focused on expanding, characterizing, and utilizing animal-derived 3D organoid models, specifically focusing on canines; but also including other animals such as swine, turtles, snakes, and squirrels to study their unique adaptations. Practically, we can use 3D canine colonoids for the screening of antiinflammatory candidates using our tumor necrosis factor (TNF)-α stimulation model. Additionally, we have grown and used swine lung organoids as a model for studying H1N1 viral uptake. Furthermore, we have grown and bio-banked tumor organoids/spheroids from a variety of canine cancers, including urothelial carcinoma (derived from urine and biopsies) and nasal carcinoma, among others, with future applications in precision veterinary medicine and comparative oncology. To accomplish this goal, we have carried out in vitro screening of a variety of chemotherapies and combination therapies at different concentrations across patients to predict in vivo responses and demonstrate the predictive ability of the organoid model. Finally, our laboratory has derived and characterized a variety of healthy organoid lines from canines including the pancreas, lung, liver, bladder, colon, and pituitary to act as healthy controls in mechanistic studies looking at disease pathogenesis and for uses in basic biology. In summary, in vitro organoid models from various animal species and tissues can be used in a variety of basic and applied research fields, including virology, toxicology, and oncology, with the potential to broaden our scientific understanding of disease and overall improve human and veterinary medicine [ABSTRACT FROM AUTHOR]