Canine Intestinal Organoids as a Novel In Vitro Model of Intestinal Drug Permeability: A Proof-of-Concept Study.

A key component of efforts to identify the biological and drug-specific aspects contributing to therapeutic failure or unexpected exposure-associated toxicity is the study of drug-intestinal barrier interactions. While methods supporting such assessments are widely described for human therapeutics, relatively little information is available for similar evaluations in support of veterinary pharmaceuticals. There is, therefore, a critical need to develop novel approaches for evaluating drug-gut interactions in veterinary medicine. Three-dimensional (3D) organoids can address these difficulties in a reasonably affordable system that circumvents the need for more invasive in vivo assays in live animals. However, a first step in developing such systems is understanding organoid interactions in a 2D monolayer. Given the importance of orally administered medications for meeting the therapeutic need of companion animals, we demonstrate growth conditions under which canine-colonoid-derived intestinal epithelial cells survive, mature, and differentiate into confluent cell systems with high monolayer integrity. We further examine the applicability of this canine-colonoid-derived 2D model to assess the permeability of three structurally diverse, passively absorbed ß-blockers (e.g., propranolol, metoprolol, and atenolol). Both the absorptive and secretive apparent permeability (Papp) of these drugs at two different pH conditions were evaluated in canine-colonoid-derived monolayers and compared with that of Caco-2 cells. This proof-of-concept study provides promising preliminary results with regard to the utility of canine-derived organoid monolayers for species-specific assessments of therapeutic drug passive permeability.

Canine Intestinal Organoids in a Dual-Chamber Permeable Support System.

The permeable support system is typically used in conjunction with traditional two-dimensional (2D) cell lines as an in vitro tool for evaluating the oral permeability of new therapeutic drug candidates. However, the use of these conventional cell lines has limitations, such as altered expression of tight junctions, partial cell differentiation, and the absence of key nuclear receptors. Despite these shortcomings, the Caco-2 and MDCK models are widely accepted and validated for the prediction of human in vivo oral permeability. Dogs are a relevant translational model for biomedical research due to their similarities in gastrointestinal anatomy and intestinal microflora with humans. Accordingly, and in support of parallel drug development, the elaboration of an efficient and accurate in vitro tool for predicting in vivo drug permeability characteristics both in dogs and humans is highly desirable. Such a tool could be the canine intestinal organoid system, characterized by three-dimensional (3D), self-assembled epithelial structures derived from adult stem cells. The (1) Permeable Support Seeding Protocol describes the experimental methods for dissociating and seeding canine organoids in the inserts. Canine organoid isolation, culture, and harvest have been previously described in a separate set of protocols in this special issue. Methods for general upkeep of the canine intestinal organoid monolayer are discussed thoroughly in the (2) Monolayer Maintenance Protocol. Additionally, this protocol describes methods to assess the structural integrity of the monolayer via transepithelial electrical resistance (TEER) measurements and light microscopy. Finally, the (3) Permeability Experimental Protocol describes the tasks directly preceding an experiment, including in vitro validation of experimental results. Overall, the canine organoid model, combined with a dual-chamber cell culture technology, overcomes limitations associated with 2D experimental models, thereby improving the reliability of predictions of the apparent oral permeability of therapeutic drug candidates both in the canine and human patient.

Standardization and Maintenance of 3D Canine Hepatic and Intestinal Organoid Cultures for Use in Biomedical Research.

Dogs develop complex multifactorial diseases analogous to humans, including inflammatory diseases, metabolic diseases, and cancer. Therefore, they represent relevant large animal models with the translational potential to human medicine. Organoids are 3-dimensional (3D), self-assembled structures derived from stem cells that mimic the microanatomy and physiology of their organ of origin. These translational in vitro models can be used for drug permeability and discovery applications, toxicology assessment, and to provide a mechanistic understanding of the pathophysiology of multifactorial chronic diseases. Furthermore, canine organoids can enhance the lives of companion dogs, providing input in various areas of veterinary research and facilitating personalized treatment applications in veterinary medicine. A small group of donors can create a biobank of organoid samples, reducing the need for continuous tissue harvesting, as organoid cell lines can be sub-cultured indefinitely. Herein, three protocols that focus on the culture of intestinal and hepatic canine organoids derived from adult stem cells are presented. The Canine Organoid Isolation Protocol outlines methods to process tissue and embedding of the cell isolate in a supportive matrix (solubilized extracellular membrane matrix). The Canine Organoid Maintenance Protocol describes organoid growth and maintenance, including cleaning and passaging along with appropriate timing for expansion. The Organoid Harvesting and Biobanking Protocol describes ways to extract, freeze, and preserve organoids for further analysis.

Systemic diseases affecting the breast: Imaging, diagnosis, and management.

Various systemic diseases of benign or malignant etiologies can clinically manifest in the breast. Some imaging findings of breast lesions can be pathognomonic for a given condition, while others are non-specific, mimicking primary breast carcinoma and requiring tissue biopsy for definitive diagnosis. In addition to obtaining a detailed clinical history, radiologists should be familiar with the diverse clinical and imaging characteristics of these conditions to help exclude primary breast cancer and avoid unnecessary interventions. This review aims to discuss the clinical presentations, imaging features, pathologic findings, and management of systemic conditions that may affect the breast.

Qualitative evaluation of MRI features in aneurysmal bone cysts after percutaneous sclerotherapy.

OBJECTIVE: To report MRI findings of changes seen in aneurysmal bone cysts after percutaneous sclerotherapy treatment. MATERIALS AND METHODS: After applying exclusion criteria, a total of 36 patients who had aneurysmal bone cysts and undergone percutaneous sclerotherapy were included in this study. The pre-treatment and post-treatment MRIs were reviewed and multiple pre-determined MRI findings were evaluated. The presence of each post-treatment finding, as well as the time for each finding to develop, was recorded. RESULTS: Early post-sclerotherapy changes include increased perilesional edema and enhancement, which appear on MRI on average 5.1 months after the initial sclerotherapy. This is followed by decreased cystic areas, which can be seen on average 5.9 months after the initial treatment. The presence of fibrosis, improved cortical integrity, and improving mass effect are later post-treatment changes and appear on MRI on average 9.7 months, 10.6 months, and 16.1 months after the initial sclerotherapy, respectively. CONCLUSION: The early and late post-sclerotherapy MR findings of aneurysmal bone cysts were reported in this study.

Pediatric ovarian volumes measured at ultrasound after contralateral unilateral oophorectomy.

BACKGROUND: Changes that occur in the remaining ovary after contralateral oophorectomy are not well described. OBJECTIVE: To determine average ovarian volume in pediatric patients after contralateral oophorectomy compared to age-matched controls with two normal ovaries. MATERIALS AND METHODS: We performed a retrospective review of ultrasound examinations and electronic medical records of patients ages 0-18 years who had unilateral oophorectomy from 2000 to 2017 (n=64). We used 384 consecutive normal age-matched ovaries for comparison, analyzing mean ovarian volumes. RESULTS: Higher mean ovarian volume (mL) was observed in patients who had oophorectomy compared to controls in the first decade of life (P<0.003) and second decade of life (P<0.0003). Higher mean ovarian volume was seen in both premenarchal and menstruating patients with prior oophorectomy when compared to controls (P<0.05 and P<0.0001, respectively). When comparing volume during menstrual cycle, we saw higher mean ovarian volumes in the oophorectomy group compared to the control group for the follicular (P<0.0001), pre-ovulatory (P=0.0005) and luteal phases (P<0.0003). We provide an updated reference of normal ovarian volumes for pediatric patients, with values similar to those already reported in the literature. CONCLUSION: Ovarian volume is higher in pediatric patients with one normal ovary following contralateral oophorectomy. The provided normative volumes can be used in evaluating these patients.

Radiologic evaluation and management of postoperative spine paraspinal fluid collections.

Postoperative paraspinal fluid collections can present a management dilemma to both radiologists and surgeons. Although many of these collections present as incidental findings and are unrelated to the presenting signs and symptoms that led to the imaging study, certain collections in the context of the appropriate clinical scenario may require additional evaluation and even emergent intervention. This article reviews those collections that are most frequently encountered and suggests management strategies that may assist in the evaluation and management of the patient.

Current status of endoscopic vein harvest in cardiac and peripheral vascular surgery.

Endoscopic harvesting of the saphenous vein (EVH) has been shown to minimize the morbidity associated with saphenous vein harvest for either coronary artery bypass or lower extremity bypass. However, the long-term benefit of a bypass procedure is predicated on conduit patency. Several studies suggest decreased patency with EVH compared with open vein harvest. Possible reasons for this discrepancy have been investigated by microscopic, electron microscopic, and functional studies of venous endothelium and contractile function of harvested veins with conflicting results. This review details the results of these studies. In addition, the clinical results of coronary bypass graft and lower extremity bypass with open vein harvest and EVH are described in regard to early wound complications and short- and long-term patency.