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Organoid cultures could constitute a valuable in vitro model to explore new treatments for canine (c) medullary thyroid carcinoma (MTC). The study's objectives were to establish and characterize 3D organoid cultures of cMTC using histology and immunohistochemistry (IHC) and to evaluate the effect of antitumor drugs on organoids' viability. Five cMTC tissue samples were used to develop organoid cultures of which one organoid line, named cMTC N°2, could be passaged for an extended period. This cMTC N°2 organoid line was further compared to the primary tumor regarding morphology and IHC expression of thyroid transcription factor-1 (TTF-1), thyroglobulin, calcitonin, synaptophysin, vimentin, Ki-67, cyclooxygenase-2 (COX-2), P-glycoprotein and vascular endothelial growth factor (VEGF). Quality control of the cMTC N°2 organoid line was achieved by a single nucleotide polymorphism (SNP) array of the organoids, primary tumor and healthy blood cells of the same dog. The effect of carboplatin, meloxicam and toceranib phosphate (TOC) on cMTC N°2 organoids' viability was evaluated. The cMTC N°2 organoid line was cultured for 94 days and showed similar histological features with the primary tumor. Immunolabeling for TTF-1, thyroglobulin, calcitonin and VEGF was similar between the primary tumor and cMTC N°2 organoids. Compared to the primary tumor, organoids showed higher immunolabeling for vimentin and Ki-67, and lower immunolabeling for synaptophysin, COX-2 and P-glycoprotein. The SNP genotype was similar for each chromosome between healthy blood cells, primary tumor and cMTC N°2 organoids. Carboplatin, meloxicam and TOC had no effect on cMTC N°2 organoid cell viability within achievable in vivo concentration range. In conclusion, the cMTC N°2 organoid line is a promising first milestone towards an established in vitro organoid model to explore pathophysiology and new treatment modalities in cMTC. This article is protected by copyright. All rights reserved.

Supplemental Material, sj-pdf-1-vet-10.1177_03009858211067478 for Computer-assisted mitotic count using a deep learning���based algorithm improves interobserver reproducibility and accuracy by Christof A. Bertram, Marc Aubreville, Taryn A. Donovan, Alexander Bartel, Frauke Wilm, Christian Marzahl, Charles-Antoine Assenmacher, Kathrin Becker, Mark Bennett, Sarah Corner, Brieuc Cossic, Daniela Denk, Martina Dettwiler, Beatriz Garcia Gonzalez, Corinne Gurtner, Ann-Kathrin Haverkamp, Annabelle Heier, Annika Lehmbecker, Sophie Merz, Erica L. Noland, Stephanie Plog, Anja Schmidt, Franziska Sebastian, Dodd G. Sledge, Rebecca C. Smedley, Marco Tecilla, Tuddow Thaiwong, Andrea Fuchs-Baumgartinger, Donald J. Meuten, Katharina Breininger, Matti Kiupel, Andreas Maier and Robert Klopfleisch in Veterinary Pathology

Supplemental material, sj-pdf-1-vet-10.1177_03009858221140818 for Prognostic clinical and histopathological features of canine cutaneous epitheliotropic T-cell lymphoma by Martina Dettwiler, Elizabeth A. Mauldin, Sara Jastrebski, Deborah Gillette, Darko Stefanovski and Amy C. Durham in Veterinary Pathology

Supplemental Material, sj-pdf-1-vet-10.1177_03009858211018813 for Validation of Immunohistochemistry for Canine Proteins Involved in Thyroid Iodine Uptake and Their Expression in Canine Follicular Cell Thyroid Carcinomas (FTCs) and FTC-Derived Organoids by Jana Jankovic, Martina Dettwiler, Martin González Fernández, Eve Tièche, Kerstin Hahn, Simon April-Monn, Matthias S. Dettmer, Martin Kessler, Sven Rottenberg and Miguel Campos in Veterinary Pathology

Canine cutaneous epitheliotropic T-cell lymphoma (CETL) and immune-mediated T-cell predominant dermatoses (IMD) share several clinical and histopathological features, but differ substantially in prognosis. The discrimination of ambiguous cases may be challenging, as diagnostic tests are limited and may prove equivocal. This study aimed to investigate transcriptional differences between CETL and IMD, as a basis for further research on discriminating diagnostic biomarkers. We performed 100bp single-end sequencing on RNA extracted from formalin-fixed and paraffin-embedded skin biopsies from dogs with CETL and IMD, respectively. DESeq2 was used for principal component analysis (PCA) and differential gene expression analysis. Genes with significantly different expression were analyzed for enriched pathways using two different tools. The expression of selected genes and their proteins was validated by RT-qPCR and immunohistochemistry. PCA demonstrated the distinct gene expression profiles of CETL and IMD. In total, 503 genes were upregulated, while 4986 were downregulated in CETL compared to IMD. RT-qPCR confirmed the sequencing results for 5/6 selected genes tested, while the protein expression detected by immunohistochemistry was not entirely consistent. Our study revealed transcriptional differences between canine CETL and IMD, with similarities to human cutaneous lymphoma. Differentially expressed genes are potential discriminatory markers, but require further validation on larger sample collections.

Insect bite hypersensitivity (IBH) is a Th-2, IgE-mediated dermatitis of horses caused by bites of insects of the genus Culicoides that has common features with human atopic dermatitis. Together with Th-2 cells, the epithelial barrier plays an important role in development of type I hypersensitivities. In order to elucidate the role of the epithelial barrier and of the skin immune response in IBH we studied the transcriptome of lesional whole skin of IBH-horses (IBH-LE; n = 9) in comparison to non-lesional skin (IBH-NL; n = 8) as well as to skin of healthy control horses (H; n = 9). To study the "baseline state" of the epithelial barrier, we investigated the transcriptome of non-lesional epidermis in IBH-horses (EPI-IBH-NL; n = 10) in comparison with healthy epidermis from controls (EPI-H; n = 9). IBH-LE skin displayed substantial transcriptomic difference compared to H. IBH-LE was characterized by a downregulation of genes involved in tight junction formation, alterations in keratins and substantial immune signature of both Th-1 and Th-2 types with particular upregulation of IL13, as well as involvement of the hypoxic pathway. IBH-NL shared a number of differentially expressed genes (DEGs) with IBH-LE, but was overall more similar to H skin. In the epidermis, genes involved in metabolism of epidermal lipids, pruritus development, as well as IL25, were significantly differentially expressed between EPI-IBH-NL and EPI-H. Taken together, our data suggests an impairment of the epithelial barrier in IBH-affected horses that may act as a predisposing factor for IBH development. Moreover, these new mechanisms could potentially be used as future therapeutic targets. Importantly, many transcriptional features of equine IBH skin are shared with human atopic dermatitis, confirming equine IBH as a natural model of skin allergy.

Supplemental Material, Combined_supplemental_materials-Bertram_et_al_2 for Computerized Calculation of Mitotic Count Distribution in Canine Cutaneous Mast Cell Tumor Sections: Mitotic Count Is Area Dependent by Christof A. Bertram, Marc Aubreville, Corinne Gurtner, Alexander Bartel, Sarah M. Corner, Martina Dettwiler, Olivia Kershaw, Erica L. Noland, Anja Schmidt, Dodd G. Sledge, Rebecca C. Smedley, Tuddow Thaiwong, Matti Kiupel, Andreas Maier and Robert Klopfleisch in Veterinary Pathology

Dieser Fallbericht beschreibt eine neue Erbkrankheit beim Braunvieh in der Schweiz. Diese auch beim Deutschen Fleckvieh als FH2 vorkommende Krankheit entspricht einer beim Menschen als Fanconi-Bickel-Syndrom bekannten Glykogenspeicherkrankheit. Sie wird durch Mutationen im SLC2A2 Gen verursacht, das für den Glukosetransporter GLUT2 kodiert. Ein Original Braunviehkalb mit homozygotem Genotyp für die FH2-assoziierte SLC2A2 Leserasterverschiebung wird vorgestellt. Die klinische Untersuchung des Kalbes zeigte ein verzögertes Wachstum, Polyurie und Polydipsie sowie schlechte Fell- und Klauenhornqualität. Die pathologische Untersuchung der Nieren zeigte makroskopisch eine unilaterale Hypoplasie sowie eine Aufhellung der Rinde. Histologisch war eine Tubulonephrose der proximalen Tubuli sowie Protein- und Glukosehaltiger Inhalt sichtbar. Eine übermässige Glykogenspeicherung war in keinem Organ feststellbar. Diese histopathologischen Befunde unterscheiden sich von den zuvor beim Fleckvieh beschriebenen Veränderungen. Beim beschriebenen Fall scheint primär eine Nierenfunktionsstörung die Wachstumsretardierung zu erklären. Den Züchtern steht ein direkter Gentest zur Ausmerzung des Defektallels zur Verfügung., INTRODUCTION This case report describes a new genetic disease of the Braunvieh breed in Switzerland. The bovine disorder also occurs in German Fleckvieh, and corresponds to human Fanconi-Bickel syndrome which is an inherited glycogen storage disease caused by mutations of the SLC2A2 gene encoding the glucose transporter GLUT2. This case report describes a single affected Original Braunvieh calf genotyped as homozygous for the FH2-associated SLC2A2 frame shift mutation. The clinical examination showed stunted growth, polyuria and polydipsia, as well as poor claw horn and coat quality. Necropsy revealed a pale cortex of the kidneys and a unilateral renal hypoplasia. Histology showed tubulonephrosis of the proximal tubules with protein- and glucose-rich contents. Glycogen accumulation was not evident in any organ. This finding is different from the reported lesions in two previously described GLUT2-deficient Fleckvieh heifers. In the presented case, growth retardation mainly seems to be associated with renal dysfunction. A direct gene test is available to eliminate the mutant allele from the population.

Nemaline myopathy (NM) is a congenital neuromuscular disorder with heterogeneous clinical signs such as muscle weakness, affecting humans, seldom dogs and cats. For human NM, several causative gene mutations have been identified. The hallmark of this disease is the presence of sarcoplasmic inclusions (nemaline rods) in skeletal muscle fibres, which can be visualised in sections of muscle specimens. Ultrastructurally, an enlargement and streaming of the Z lines can be detected. Here, the case of two 10-week-old female Border collie littermates with progressive gait dysfunction, muscle stiffness and exercise intolerance, which were euthanased because of poor prognosis, is described. Histology of formalin-fixed paraffin-embedded muscle samples revealed sarcoplasmic rods visible in tissue sections stained with haematoxylin and eosin, the Gomori trichrome and the phosphotungstic acid haematoxylin stain. In electron microscopy, the rods were confirmed as streaming of the Z lines. Therefore, a congenital NM was diagnosed.

Supplemental Material, DS1_VET_10.1177_0300985818755254 for Validation of Digital Microscopy Compared With Light Microscopy for the Diagnosis of Canine Cutaneous Tumors by Christof A. Bertram, Corinne Gurtner, Martina Dettwiler, Olivia Kershaw, Kristina Dietert, Laura Pieper, Hannah Pischon, Achim D. Gruber, and Robert Klopfleisch in Veterinary Pathology