USE OF CANINE INTESTINAL ORGANOIDS AS AN ALTERNATIVE MODEL FOR THE STUDY OF GASTRINTESTINAL DISEASES
Paracellular barrier. Dogs. Clostridium. Intestinal epithelium.
Knowledge in the health sciences related to the physiology and morphology of the species, development of drugs, vaccines and techniques began with the use of animals in laboratory experiments. Despite the advances made, the search for alternative models to replace animal models has become a real necessity. In this context, the organoids consist of a three-dimensional system of cell culture that develops in a limited space, presents cell differentiation and morphology similar to the organ of origin, favoring its use in species-specific experiments. Currently, the host-pathogen interaction stands out as the main point of interest in research involving organoid technology. The present study aims to isolate and cultivate canine intestinal organoids and to evaluate the integrity of the paracellular barrier of the epithelium through exposure to purified toxins of bacteria of the genus Clostridium. Colony fragments from dogs submitted to euthanasia for causes unrelated to gastrointestinal diseases will be collected at the University of Veterinary Medicine in Utrecht, The Netherlands. These will be subjected to enzymatic digestion, seeded in Matrigel, expanded in suitable culture medium and evaluated for cell composition, functionality and morphology through real-time quantitative PCR and immunohistochemistry. The genetic stability of canine colonoids will be analyzed by karyotyping and the paracellular barrier function by fluorescence stereomicroscopy. Statistical analysis will be descriptive and the Mann-Whitney test for evaluation of gene expression and colonoid functionality, with a 95% confidence interval, will be applied.