Histological evidence of superficial inflammation is associated with lower recurrence of equine sarcoids following surgical removal: A follow-up study of 106 tumours in 64 horses.

Although the equine sarcoid is the most common skin neoplasm in domesticated horses, histopathological characteristics have not previously been evaluated for association with recurrence. The aim of this retrospective cohort study was to investigate clinical and histopathological features of excised equine sarcoids and to evaluate their association with recurrence at the original surgical site and at new sites. Clinical records and excisional biopsies from 106 equine sarcoids from 64 horses referred to Leahurst Equine Hospital, University of Liverpool, between March 2010 and February 2015 were retrieved. Biopsies were re-evaluated histologically. Clinical data were obtained from hospital records, and owner-reported follow-up data were obtained by telephone questionnaire. Associations between clinical and histopathological features of sarcoids and their recurrence at the surgical site were determined using uni- and multivariable mixed effects logistic regression. Recurrence of sarcoids at the surgical site occurred in 30 horses (46.9%). Sarcoids developed at a distant site in 21 horses (32.8%). In the final mixed effects logistic regression model, only superficial inflammation was associated with reduced odds of recurrence at the surgical site (adjusted odds ratio, 0.32; 95% confidence intervals, 0.10-0.96; P = 0.04). This suggests that the inflammatory process may play a role in protecting horses against the recurrence of sarcoids.

European Forensic Veterinary Pathology Comes of Age.

During the mid-1700s, development of the veterinary profession was largely focussed on equine medicine and surgery. Subsequently, rather erratic development encompassed other species and eventually led to specialization in different disciplines. Teaching of veterinary pathology was well established in Europe and North America by the late 19th century. Specialization in this discipline was boosted in the 1940s by the formation, in the USA, of the Register of Veterinary Pathology and American College of Veterinary Pathologists. National societies followed soon afterwards in Europe. The European Society of Veterinary Pathology evolved during this period and the European College of Veterinary Pathologists (ECVP) was created in 1995 to promote high standards in the discipline. As an accrediting body, its emphasis is on training and harmonization across Europe. There is an increasing demand for high-grade forensic veterinary pathology reports which address the requirements of the legal system, but so far only a few countries have defined protocols for these reports. In recognition of the need for a specific qualification that benchmarks the competences and experience expected of forensic veterinary pathologists, the ECVP recently launched the Certificate in Forensic Veterinary Pathology.

Histological examination of the interspinous ligament in horses with overriding spinous processes.

The equine interspinous ligament (ISL) consists of an oblique crossing arrangement of collagenous bundles which are thought to counteract the tensile and rotational forces of distraction between the spinous processes (SPs) in the caudal thoracic and cranial lumbar spine. The aim of this controlled histological study was to assess the structural anatomy and innervation of the ISL in horses with clinically significant overriding (dorsal) SPs (ORSPs) and to compare the findings with the ISL of normal horses. Samples of the ISL were obtained from 10 horses that underwent subtotal ostectomy for treatment of ORSPs. Control samples were obtained from horses without spinal pathology. Histological staining of ISL sections with haematoxylin and eosin was performed to assess the morphology of the ligaments and with Alcian blue-periodic acid-Schiff to determine the proteoglycan and glycosaminoglycan content. Immunohistochemistry for S100 was performed for quantitative evaluation of nerves within the ISL. The ISL in horses with ORSPs had an altered collagen fibre alignment and arrangement of the ligamentous layers when compared to healthy controls. A significant increase in fibrocartilaginous tissue with evidence of fibrocartilaginous metaplasia was detected (P=0.001). The number of nerves in the ISL samples was significantly higher in horses with ORSPs than in controls (P=0.017). Structural alterations of the ISL, including loss of fibre alignment and fibrocartilaginous metaplasia, are associated with ORSPs in the equine thoracolumbar spine. In addition, an increase in innervation of the ISL in horses with ORSPs, compared to normal, may explain the thoracolumbar pain experienced by some horses with ORSPs.

Serum and blister-fluid elevation and decreased epidermal content of high-mobility group box 1 protein in drug-induced Stevens-Johnson syndrome/toxic epidermal necrolysis.

BACKGROUND: High-mobility group box 1 (HMGB1) is a damage-associated molecular-pattern protein. Stevens-Johnson syndrome (SJS)/toxic epidermal necrolysis (TEN) are serious, immune-mediated skin-blistering conditions. OBJECTIVES: To determine serum and/or blister-fluid total HMGB1 levels in SJS/TEN cohorts, and HMGB1 expression in formalin-fixed, paraffin-embedded (FFPE) SJS/TEN skin vs. healthy and maculopapular exanthema (MPE) skin. Methods Serum HMGB1 was quantified in Malawian nevirapine-induced hypersensitivity, Taiwanese SJS/TEN and Spanish SJS/TEN cohorts. FFPE skin (healthy skin, MPE, SJS/TEN) was stained and assessed for HMGB1 expression. RESULTS: Serum total HMGB1 was not significantly elevated in patients with nevirapine-induced SJS/TEN (3·98 ± 2·17 ng mL-1 ), MPE (3·92 ± 2·75 ng mL-1 ) or drug reaction with eosinophilia and systemic symptoms (4·73 ± 3·00 ng mL-1 ) vs. tolerant controls (2·97 ± 3·00 ng mL-1 ). HMGB1 was significantly elevated in Taiwanese patients with SJS/TEN, highest during the acute phase (32·6 ± 26·6 ng mL-1 ) vs. the maximal (19·7 ± 23·2 ng mL-1 ; P = 0·007) and recovery (24·6 ± 25·3 ng mL-1 ; P = 0·027) phases. In blister fluid from Spanish patients with SJS/TEN, HMGB1 (486·8 ± 687·9 ng mL-1 ) was significantly higher than in serum (8·8 ± 7·6 ng mL-1 ; P <0·001). Preblistered SJS/TEN skin showed decreased epidermal nuclear HMGB1 expression in upper epidermis vs. healthy or MPE skin but retained basal/suprabasal expression. CONCLUSIONS: Epidermal HMGB1 expression was decreased in SJS/TEN skin. Retained basal/suprabasal epidermal HMGB1 expression may exacerbate localized injury in SJS/TEN.

Histocytic-like Atypical Mast Cell Tumours in Horses.

This report describes a series of four equine mast cell tumours (MCTs) with atypical morphological features. The tumours were 1-2 cm in diameter and mostly localized to the eyes (one eyelid, two conjunctiva). Histologically, they were composed of very large (up to 35 µm) round pleomorphic cells with a large central to paracentral nucleus and abundant granular cytoplasm. A large number of viable mature eosinophils were detected intermingled with the large round cells. Histochemical staining (toluidine blue and Perls' Prussian blue) and immunohistochemistry (KIT, mast cell tryptase, lysozyme and proliferating cell nuclear antigen) confirmed the mast cell origin of the atypical cells and identified an aberrant KIT protein expression in three cases. Based on morphological and immunohistochemical features, we propose to call the lesions equine histiocytic-like atypical MCTs.

Neuroaxonal Dystrophy in a Flock of Pied Imperial Pigeons (Ducula bicolor).

Five juvenile pied imperial pigeons (Ducula bicolor) presented with neurological signs including torticollis, ataxia and poor flying ability. All were humanely destroyed and submitted for post-mortem examination. Microscopically, the most significant findings were in the brain and spinal cord. Spheroid formation was evident within the medulla, pons, diencephalon, cortical grey and subcortical white matter, spinal cord white and grey matter and the granular and molecular cell layers of the cerebellum. There was no evidence of associated inflammation. Immunohistochemistry revealed positive labelling within the spheroids for S100 axons and phosphorylated neurofilaments including SMI31, neurofilament cocktail and microtubule-associated protein 2. Transmission electron microscopy confirmed the light microscopical findings of frequent axonal spheroids. These results are consistent with neuroaxonal dystrophy, which has not been described previously in pigeons. This highlights the importance of considering neuroaxonal dystrophy in juvenile birds with neurological signs. A genetic basis is suspected in this group.

Structure and Innervation of the Equine Supraspinous and Interspinous Ligaments.

Pain related to the osseous thoracolumbar spine is common in the equine athlete, with minimal information available regarding soft tissue pathology. The aims of this study were to describe the anatomy of the equine SSL and ISL (supraspinous and interspinous ligaments) in detail and to assess the innervation of the ligaments and their myofascial attachments including the thoracolumbar fascia. Ten equine thoracolumbar spines (T15-L1) were dissected to define structure and anatomy of the SSL, ISL and adjacent myofascial attachments. Morphological evaluation included histology, electron microscopy and immunohistochemistry (S100 and Substance P) of the SSL, ISL, adjacent fascial attachments, connective tissue and musculature. The anatomical study demonstrated that the SSL and ISL tissues merge with the adjacent myofascia. The ISL has a crossing fibre arrangement consisting of four ligamentous layers with adipose tissue axially. A high proportion of single nerve fibres were detected in the SSL (mean = 2.08 fibres/mm2 ) and ISL (mean = 0.75 fibres/mm2 ), with the larger nerves located between the ligamentous and muscular tissue. The oblique crossing arrangement of the fibres of the ISL likely functions to resist distractive and rotational forces, therefore stabilizing the equine thoracolumbar spine. The dense sensory innervation within the SSL and ISL could explain the severe pain experienced by some horses with impinging dorsal spinous processes. Documentation of the nervous supply of the soft tissues associated with the dorsal spinous processes is a key step towards improving our understanding of equine back pain.

Clinical outcome, PDGFRß and KIT expression in feline histiocytic disorders: a multicentre study.

Information about histiocytic disease in cats is limited. The aim of this study was to document clinical findings and outcome in feline histiocytic disorders, and characterize the expression of PDGFRß and KIT in order to identify potential treatment targets. Morphologically diagnosed feline histiocytic tumours were reviewed and characterized by immunohistochemistry (IHC). Five cases of feline progressive histiocytosis (FPH), eight histiocytic sarcomas (HS) and two haemophagocytic histiocytic sarcomas (HaeHS) were confirmed. PDGFRß was variably positive in most histiocytic cases, while KIT was negative in all. Clinical presentation, treatment and outcome were also evaluated. Partial responses were recorded in measurable disease with tyrosine kinase inhibitors and lomustine, and radiotherapy achieved long-term control in some cases. Survival times were shortest in HaeHS and disseminated disease. PDGFRß, but not KIT, may represent a therapeutic target in feline histiocytic disorders but more studies are needed to investigate other potential treatment targets.

Immunohistochemical expression of MDR1-Pgp 170 in canine cutaneous and oral melanomas: pattern of expression and association with tumour location and phenotype.

Canine melanoma (CMM) more commonly affects the oral mucosa and the cutis. CMM shares several features with human melanomas (HMM), included resistance to a broad variety of antineoplastic chemotherapy agents. P-glycoprotein 1 (Pgp) expression is a well-recognized feature of multi-drug resistance and the purpose of this study was to investigate its expression in treatment naïve CMM. We also investigated Pgp association with tumour location and histological features. Histology records of CMM were retrieved, including patients from 2012-2014. Twenty-five cases of CMM were included in this study. Results revealed that Pgp is expressed in CMM and oral tumours were more likely to have a membranous Pgp expression (100%) than cutaneous tumours (66.6%) (P = 0.010). Cytoplasmic and nuclear Pgp expression could also be identified. Results of this study bring useful data that help in understanding one of the possible mechanisms responsible of intrinsic chemotherapy resistance in canine CMM.

Blunt Force Trauma in Veterinary Forensic Pathology.

Veterinary pathologists commonly encounter lesions of blunt trauma. The development of lesions is affected by the object's mass, velocity, size, shape, and angle of impact and by the plasticity and mobility of the impacted organ. Scrape, impact, and pattern abrasions cause localized epidermal loss and sometimes broken hairs and implanted foreign material. Contusions are best identified after reflecting the skin, and must be differentiated from coagulopathies and livor mortis. Lacerations-traumatic tissue tears-may have irregular margins, bridging by more resilient tissue, deviation of the wound tail, crushed hairs, and unilateral abrasion. Hanging or choking can cause circumferential cervical abrasions, contusions and rupture of hairs, hyoid bone fractures, and congestion of the head. Other special forms of blunt trauma include fractured nails, pressure sores, and dog bites. Ocular blunt trauma causes extraocular and intraocular hemorrhages, proptosis, or retinal detachment. The thoracic viscera are relatively protected from blunt trauma but may develop hemorrhages in intercostal muscles, rib fractures, pulmonary or cardiac contusions or lacerations with subsequent hemothorax, pneumothorax, or cardiac arrhythmia. The abdominal wall is resilient and moveable, yet the liver and spleen are susceptible to traumatic laceration or rupture. Whereas extravasation of blood can occur after death, evidence of vital injury includes leukocyte infiltration, erythrophagocytosis, hemosiderin, reparative lesions of fibroblast proliferation, myocyte regeneration in muscle, and callus formation in bone. Understanding these processes aids in the diagnosis of blunt force trauma including estimation of the age of resulting injuries.

Cutaneous Heterotopic Brain Tissue (Neuroglial Choristoma) with Dysplastic Features in a Kitten.

A 3-month-old, male European shorthair kitten exhibited an ill-defined, soft mass on the skin of the frontal head, which was present since birth. The surgically resected tissue was representative of a discrete dermal and subcutaneous mass comprising islands of neurons, glial and meningothelial elements, sometimes atypical or dysplastic, separated by dense collagenous connective tissue. There was no evident connection between this tissue and the brain. Immunohistochemical examination confirmed the presence of neurons and a pleocellular glial population, supporting a diagnosis of cutaneous neuroglial choristoma believed to be secondary to sequestered (resolved) meningoencephalocoele. Ectopic brain tissue is very rare in small animals. Some atypical features displayed by this tissue may be misdiagnosed as neoplasia. Communication between surgeon and pathologist to clarify the relationship of the lesion to surrounding structures is helpful to avoid misdiagnosis.

Presentation, clinical pathological and post-mortem findings in three related Scottish terriers with ligneous membranitis.

Ligneous conjunctivitis and gingivitis were diagnosed in three related Scottish terrier dogs presented for investigation of severe conjunctivitis and respiratory signs. Hypoplasminogenaemia was confirmed in one of the three affected dogs. Supportive treatment was not effective, and the dogs died or were euthanased because of the disease. Post-mortem analysis of two of the dogs revealed multiple abnormalities including severe proliferative fibrinous lesions affecting the conjunctiva, gingiva, trachea, larynx and epicardium and multiple fibrous adhesions throughout the thoracic and abdominal cavities. One dog had internal hydrocephalus and lacked a cerebellar vermis. Ligneous membranitis was confirmed on histopathology. This is a rare condition in dogs but an important differential diagnosis for severe conjunctivitis and gingivitis.

Canine pancreatic islet cell tumours secreting insulin-like growth factor type 2: a rare entity.

Insulin-like growth factor type II (IGF-II) is the main cause of non-islet cell tumour hypoglycaemia (NICTH) and insulin is thought to be the only factor causing hypoglycaemia in insulinomas. However, two case reports of pancreatic neuroendocrine tumours (PNETs) producing IGF-II have been previously published: a human and a canine patient. In this study, we investigated clinical, histopathological, immunohistochemical and ultrastructural features, and biological behaviour of canine pancreatic IGF-II-omas, a subgroup of PNETs that has not been previously characterized. Case records of 58 dogs with confirmed PNETs and hypoglycaemia were reviewed: six patients were affected by IGF-II-omas. Surgery was performed in all cases and two dogs had metastases. Four patients remained alive and in remission at 370, 440, 560 and 890 days post-diagnosis; two died of non-tumour-related causes. IGF-II-omas can be differentiated from insulinomas through hypoinsulinaemia, IGF-II positive and insulin negative immunostaining. The prevalence of this neoplasia is low, accounting for just 6% of PNETs.

Equine Cutaneous Mast Cell Tumours Exhibit Variable Differentiation, Proliferation Activity and KIT Expression.

Equine cutaneous mast cell tumours (CMCTs) are generally considered to be benign skin lesions, although recurrent and multicentric tumours have been described. For canine CMCTs, grading and prognostic approaches are well established and aberrant KIT expression as well as high proliferation indices are associated with poor outcome. However, in the case of equine CMCTs, morphological features, proliferative activity and KIT expression pattern have not been assessed or related to biological behaviour, and there is discussion as to whether CMCTs are true neoplastic processes. The present study describes 45 equine CMCTs in terms of their morphology and KIT and PCNA expression by immunohistochemistry. KIT expression was classified as membranous (I), cytoplasmic and focally stippled (II) or diffuse cytoplasmic (III). A large proportion of the tumours were multinodular or diffuse dermal infiltrates of mast cells with mild anisokaryosis, a low proliferative rate and a dominance of KIT pattern I, representing well-differentiated CMCTs. In approximately one third of the cases, the mast cells exhibited more infiltrative growth, moderate to marked anisokaryosis and a higher degree of proliferation. These were classified as poorly differentiated CMCTs and exhibited only KIT patterns II and III. These findings indicate that there is a subgroup of poorly differentiated equine CMCTs, in which there is an association between aberrant KIT expression, high proliferative rate and potential aggressive behaviour, all features that confirm at least the poorly differentiated CMCT as a true neoplastic processes.

Ehlers-Danlos syndrome associated with fatal spontaneous vascular rupture in a dog.

A 7-month-old male cross breed dog was presented with hyperextensible skin and atrophic scarring. A diagnosis of Ehlers-Danlos syndrome was made based on clinical signs, histopathology and electron microscopy. Two weeks after presentation, the dog died suddenly. Post-mortem examination revealed haemothorax and rupture of the left subclavian artery. Histological findings, including Goldner's modified Masson's trichrome staining and transmission electron microscopy of the subclavian artery, revealed abnormalities in the structure and arrangement of collagen fibrils, suggesting that the defective collagen formation extended to the vasculature. To the authors' knowledge, this is the first report of Ehlers-Danlos syndrome with vascular involvement in animals.

Metastatic cancer of unknown primary in 21 dogs.

The aim of this retrospective study was to describe clinical features, treatment and outcome of 21 dogs with metastatic cancer of unknown primary (MCUP), a biopsy-proven malignancy being diagnosed at a metastatic stage, in which the anatomical origin of the primary tumour cannot be detected. All dogs underwent total-body computed tomography. Signalment, type and duration of clinical signs, metastasis site, pathology results, treatment and outcome were recorded. Carcinoma was the most common diagnosis (57.1%), followed by sarcoma, melanoma and mast cell tumour. The median number of disease sites per dog was 2, with bones, lymph nodes, lungs and spleen being the most frequent metastatic locations. The median survival for all dogs was 30 days. Overall, a primary site was not identified in 20 (95.2%) dogs. MCUP encompasses a variety of different pathologic entities and harbours a poor prognosis.

Aberrant subcellular immunolocalization of NOTCH-1 activated intracellular domain in feline mammary tumours.

NOTCH-1 is a transmembrane receptor protein. Ligand proteins expressed on the surface of neighbouring cells bind to the NOTCH-1 extracellular domain by juxtacrine signalling and release the NOTCH intracellular domain (NICD) to alter gene expression. Forty feline mammary lesions (34 malignant and six hyperplastic) were submitted for immunohistochemical analysis of NICD expression using an anti-feline NICD monoclonal antibody. Associations between NICD expression in carcinomas and morphological parameters, as well as overall survival (OS), were investigated. NICD nuclear expression was observed in hyperplastic lesions (100%) while cytoplasmic localization was evident in carcinomas (0% nuclear positive; 87.5% cytoplasmic positive; 12.5% negative). Cytoplasmic NICD localization was statistically associated with carcinomas, while nuclear labelling was associated with hyperplasia. No significant correlation between positive or negative NICD expression and OS or morphological parameters was detected. NOTCH-1 activation, immunohistochemically identified by the NICD active form, appears to play a role in feline mammary carcinoma biology as the majority of tumours express this protein. Nuclear localization is consistent with the established NICD metabolic intranuclear pathway while cytoplasmic accumulation suggests aberrant NOTCH-1 signalling typical of malignant tumour progression.

Characterization and potential applications of progenitor-like cells isolated from horse amniotic membrane.

The aim of this work was to isolate, for the first time, progenitor-like cells from the epithelial (AECs) and mesenchymal (AMCs) portions of the horse amniotic membrane, and to define the biological properties of these cells. AECs displayed polygonal epithelial morphology, while AMCs were fibroblast-like. Usually, six to eight passages were reached before proliferation decreased, with 13.08 and 26.5 cell population doublings attained after 31 days for AECs and AMCs, respectively. Immunocytochemical studies performed at passage 3 (P3) showed that both cell populations were positive for the expression of specific embryonic markers (TRA-1-60, SSEA-3, SSEA-4 and Oct-4). Meanwhile, RT-PCR performed at P1 and P5 showed expression of mesenchymal stem/stromal cell markers (CD29, CD105, CD44 and CD166) with negativity for CD34 at P1, although this marker began to be expressed by P5. The cells also expressed MHC-I at both P1 and P5, but lacked MHC-II expression at P1. Both AECs and AMCs demonstrated high plasticity, differentiating in vitro toward the osteogenic, adipogenic, chondrogenic and neurogenic lineages. Equine amnion-\derived cells could also be frozen and recovered without loss of their functional integrity in terms of morphology, presence of specific stemness markers and differentiation ability, although the renewal capacity was lower than that observed for freshly isolated cells. To investigate potential therapeutic effects and cell tolerance in vivo, horse amnion-derived cells were allogeneically injected into three horses with tendon injuries, resulting in a quick reduction in tendon size and ultrasonographic cross-sectional area measurements. These results suggest that horse amnion-derived cells may be useful for cell therapy applications.

Canine invasive lobular carcinoma of the mammary gland: morphological and immunohistochemical characterizations of three cases.

Invasive lobular carcinoma (ILC) represents 15% of invasive human breast tumours. This report describes the morphological and immunohistochemical features of three canine mammary tumours comparable with human ILC. These tumours were composed of a non-delimited proliferation of discrete cells infiltrating fibrous connective tissue. Multifocal in-situ carcinoma associated with invasive lesions was present. Invasive tumour cells and in-situ lesions expressed cytokeratin and CK34betaE12, but not E-cadherin. Based on these morphological and immunohistochemical characteristics, the tumours were classified as canine ILC.