In Vivo Tumorigenicity of the 20q11.21 Amplicon in an Engraftment Model of hPSCs and Differentiated Liver Cells.

Human pluripotent stem cells (hPSCs) are a promising source of somatic cells for clinical applications and disease modelling. However, during culture they accumulate genetic aberrations such as amplification of 20q11.21 which occurs in approximately 20% of extensively cultured hPSC lines and confers a BCL2L1-mediated survival advantage. During the production of the large number of cells required for transplantation and therapy these aberrations may become unavoidable which has important safety implications for therapies and may also impact upon disease modelling. Presently, these risks are poorly understood; whilst it is apparent that large-scale genetic aberrations can pose an oncogenic risk, the risks associated with smaller, more insidious changes have not been fully explored. In this report, the effects of engraftment of human embryonic stem cells (hESCs) and hESC-derived hepatocyte-like cells (HLCs) with and without amplification of the 20q11.21 minimal amplicon and isochromosome 20q (i20q) in SCID-beige mice are presented. The cells were tracked in vivo using a luminescent reporter over a period of approximately four months. Intrasplenic injection of hESCs showed greater engraftment potential and the formation of more severely disruptive lesions in the liver and spleen of animals injected with cells containing 20q11.21 compared with i20q and wild type. HLCs with 20q11.21 engrafted more successfully and formed more severely disruptive lesions than wild type cells or cells with i20q. These results reinforce the notion that karyotyping of therapeutic hPSC is required for transplant, and suggest that screening for known common aberrations is necessary. Further work to identify commonly arising genetic aberrations should be performed and routine screening for hPSCs intended for therapeutic use should be used.

MRI ischemic and hemorrhagic lesions in arterial and venous territories characterize central nervous system intravascular lymphoma in dogs.

Intravascular lymphoma (IVL) is characterized by the proliferation of large malignant lymphocytes within the lumen of blood vessels. This retrospective, multi-center, case series study aimed to describe the MRI features of confirmed central nervous system IVL in dogs and compare them with histopathological findings. Medical record databases from seven veterinary centers were searched for cases of histologically confirmed IVL. Dogs were included if an MRI was performed. The MRI studies and histopathology samples were reviewed to compare the MRI changes with the histopathological findings. Twelve dogs met the inclusion criteria (12 brains and three spinal cords). Imaging of the brains revealed multifocal T2-weighted/FLAIR hyperintense and T1-weighted iso-hypointense lesions, with variable contrast enhancement; areas of abnormal diffusion both in arterial and venous territories in diffusion-weighted imaging; and meningeal enhancement. On gradient echo images (GRE), the changes comprised tubular susceptibility artifacts, consistent with the "susceptibility vessel sign", and additional variably sized/shaped intraparenchymal susceptibility artifacts. Spinal cord lesions presented as fusiform T2-weighted hyperintensities with scattered susceptibility artifacts on GRE and variable parenchymal and meningeal contrast enhancement. On histopathology, subarachnoid hemorrhages and neuroparenchymal areas of edema and necrosis, with or without hemorrhage, indicating ischemic and hemorrhagic infarctions, were found. These lesions were concurrent with severely dilated meningeal and parenchymal arteries and veins plugged by neoplastic lymphocytes and fibrin. Due to the unique angiocentric distribution of IVL, ischemic and hemorrhagic infarcts of variable chronicity affecting both the arterial and venous territories associated with thrombi formation can be detected on MRI.

Quantitative histologic evaluation reveals different degree of liver atrophy in cachectic and starved dogs.

Cases of neglect in dogs are among the forensic cases submitted most commonly for postmortem examination. Starvation is a form of primary protein-energy malnutrition in which the availability of food is severely restricted or absent; cachexia is a form of protein-energy malnutrition secondary to progressive metabolic derangement during chronic diseases. Despite both conditions leading to an emaciated appearance of the cadaver, discrimination between the two is crucial in forensic cases. We hypothesized that among emaciated dogs, the degree of liver atrophy in starved animals is higher than in cachectic ones, and that this can be investigated microscopically, regardless of the degree of cadaver decomposition. We studied 46 animals: 23 starved, 11 cachectic, and 12 control dogs. Portal tracts were identified by the presence of a bile duct and associated vascular structures recognizable by a thin rim of collagen still visible regardless of the degree of cadaver decomposition. The number of portal tracts per lpf (10×) was used as an indirect measure of atrophy. The number of portal tracts in starved dogs was significantly higher (p < 0.01) compared to both cachectic and control dogs, indicating a higher degree of liver atrophy in starvation. Measuring the density of portal tracts offers a reliable additional tool for discrimination between starvation and cachexia.

Pulmonary bleeding in racehorses: A gross, histologic, and ultrastructural comparison of exercise-induced pulmonary hemorrhage and exercise-associated fatal pulmonary hemorrhage.

Exercise-induced pulmonary hemorrhage (EIPH) is a common condition of Thoroughbred racehorses that is usually responsible for reduced performance, while exercise-associated fatal pulmonary hemorrhage (EAFPH) is characterized by severe pulmonary bleeding of unknown pathogenesis resulting in sudden death during strenuous exercise. The aim of the study was to characterize and compare anamnestic data together with pulmonary gross, histologic, and ultrastructural findings in racehorses with EIPH (n = 10), EAFPH (n = 10), and control horses (n = 5). No differences in anamnesis were identified between the 3 groups. Grossly cranial lobe reddening and edema scores were significantly more prevalent and severe in the EAFPH group compared with the EIPH and control groups. Histologically, hemorrhage scores were higher in the EAFPH group, while hemosiderophages, iron encrustations of collagen and elastin fibers, and vascular remodeling scores were significantly higher in EIPH group compared with the EAFPH and control groups. In all groups, caudal lung locations exhibited a significantly higher score for vascular remodeling, hemosiderophage accumulation, iron encrustation, and type II pneumocyte hyperplasia when compared with cranial, dorsal, and ventral locations. Ultrastructural analysis of perivascular collagen showed fibrils with significantly larger diameters in the EAFPH group compared with the EIPH group but not compared with the control group. This study demonstrates that lungs of horses that experienced EAFPH show significantly less vascular remodeling and other long-term pulmonary abnormalities that characterize horses with EIPH.

Elephant Endotheliotropic Herpesvirus 4 and Clostridium perfringens Type C Fatal Co-Infection in an Adult Asian Elephant (Elephas maximus).

Elephant endotheliotropic herpesvirus hemorrhagic disease (EEHV-HD) is an acute, often fatal, multisystemic hemorrhagic disease and one of the most significant causes of mortality of Asian elephants in captivity. Most fatal cases of EEHV-HD are associated with EEHV1A and EEHV1B in juveniles. This case report describes the clinical and pathological features of a fatal co-infection of Clostridium perfringens type C and EEHV-HD, caused by EEHV4, in an adult female Asian elephant. Although fatal clostridial enterotoxemia has been occasionally reported in elephants, this report highlights the importance of having both EEHV-HD and clostridial enterotoxemia as potential differential diagnoses in cases of widespread tissue necrosis and internal hemorrhage in elephants, regardless of the animal age group, due to their macroscopic similarities, frequent co-occurrence and cumulative morbid potential.

Leukocyte numbers and intestinal mucosal morphometrics in horses with no clinical intestinal disease.

Healthy horses and other animals have large numbers of resident leukocytes in the intestinal wall, but there is scant information regarding which and how many leukocytes are normally present in the equine intestinal wall. Our aim was to provide a reference range of leukocytes in the intestinal mucosal and submucosal propria of normal horses. We included in our study intestinal tissues from 22 Thoroughbred racehorses with no clinical intestinal disease, which had been euthanized because of catastrophic musculoskeletal injuries. Neutrophils, lymphocytes, eosinophils, macrophages, and plasma cells were counted in 5 random 17,600-µm2 areas of villus lamina propria of the duodenum, jejunum, and ileum, and deep lamina propria of the duodenum, jejunum, ileum, right ventral colon, left ventral colon, left dorsal colon, right dorsal colon, and small colon. Other features investigated in the same intestinal segments included villus height and width (small intestine), presence of ciliated protozoa, Paneth cells number, subcryptal leukocyte layers (number of leukocyte layers between the bottom of the crypts and the muscularis mucosae), and submucosal leukocytes. Lymphocytes were the most numerous cells in all segments analyzed, followed by plasma cells, eosinophils, macrophages, and neutrophils. Eosinophil numbers were significantly higher in both lamina propria and submucosa of the large intestine than in the small intestine. The duodenum had shorter and thinner villi than either jejunum or ileum. The data provided from our study will be useful for diagnosticians examining inflammatory processes in the intestinal tract of horses.

Disseminated Nocardiosis Caused by Nocardia farcinica in Two Puppy Siblings.

Systemic nocardiosis due to Nocardia farcinica has not been reported in canine outbreaks. Two 14-week-old female Dogue de Bordeaux siblings presented with fever and severe, acute onset limb lameness; traumatic lesions with evidence of infection were identified over the lame limbs of both dogs. The patients were euthanised owing to lack of therapeutic response and rapid escalation to systemic infection with central nervous system manifestations. The post-mortem changes consisted of multiple disseminated abscesses, mainly affecting the skin and subcutis at the limb traumatic injuries, local and hilar lymph nodes, lung, kidney and brain. Bacterial culture and identification via MALDI-TOF and 16S rRNA sequencing revealed Nocardia farcinica from several of these sites in both dogs. Clinical significance of the isolate was supported by cytology of the post-mortem organs' impression smears showing numerous branching filamentous bacteria associated with inflammation. The organism displayed marked multidrug-resistance. No history of immunosuppression was available, and immunohistochemistry ruled out viral pathogens as canine distemper and parvovirus. N. farcinica should be considered as a potential differential cause of sudden lameness and systemic infection in dogs with traumatic skin lesions over the limbs. This is the first reported small-scale outbreak of systemic nocardiosis in dogs due to N. farcinica.

Intratendinous Fibroma of the Superficial Digital Flexor Tendon within the Carpal Sheath of a Horse.

A 20-year-old British Warmblood gelding was presented for a progressively worsening right forelimb lameness which developed following an intense dressage training session. Initial ultrasound examination revealed a triangular, intrathecal, hypoechoic region within the superficial digital flexor tendon (SDFT) in the proximal, palmar carpal region distal to the accessory carpal bone (ACB), extending 7cm distally into the proximal metacarpal region. No significant improvement in clinical presentation was observed following an eight-week rehabilitation programme. Repeat ultrasound examination revealed an enlarged cross-sectional area of affected tendon. Due to a poor clinical response to conservative treatment, combined with increasingly marked severity of the lameness, the horse was humanely euthanised and the affected SDFT was submitted for macroscopic and histopathological examination at the University of Liverpool. This revealed an approximately 70mm-elongated, focally extensive mass located within the SDFT. On cross-section, the mass was poorly delineated, irregularly triangular, focally haemorrhagic, firm, 15 × 12mm wide and eccentrically placed towards the palmar aspect of the tendon. Histopathological examination identified a moderately to highly cellular, infiltrative, poorly demarcated mesenchymal neoplasm comprised of streams of moderately atypical spindloid cells including bizarre mitoses. Intratendinous fibroma is an uncommonly recorded human neoplasm and this case represents the third reported case of this entity in the horse, the first in an adult horse and the first to be identified in the SDFT.

The use of Corticotropin-releasing factor (CRF) as a marker of agonic stress in abused dogs: Preliminary results.

In Veterinary Forensic Medicine, determination of the degree of animal suffering is an essential element for the prosecution of perpetrators of animal abuse. The purpose of this study is to find a suitable immunohistochemical marker for the assessment of suffering to be routinely used in Veterinary Forensic Pathology, by analyzing the expression of Corticotropin-releasing factor (CRF) in formalin-fixed brains of dogs as a measurement of the agonic stress. CRF, a key peptide element in exogenous and endogenous stressors adaptation, can regulate endocrine-behavioral responses to stress stimulating pituitary ACTH release and consequent adrenal secretion of glucocorticoids. Since CRF acts in days or weeks, this study investigates its role as a potential distinctive marker between sudden death and death associated with a longer agonic period. The study used immunohistochemistry (IHC) to evaluate the CRF expression in the brain of dogs that suffered sudden death, as compared to dogs that died after long-term agonic stress. IHC labelling analysis was performed with machine-learning-based software and the results were statistically evaluated. Our results demonstrate for the first time that CRF is a promising marker of stress in abused patients also in Veterinary Medicine.

A Case Series of Five Horses with Superficial Digital Flexor Tendon Lesions in the Carpal Canal.

This case series discusses the clinical presentation, ultrasonographic findings, treatment, and outcome of 5 horses with superficial digital flexor tendon (SDFT) lesions within the carpal canal. The horses' ages ranged from 11-28 years, and presented with an acute, unilateral forelimb lameness which worsened following proximal limb flexion. The presence of a lesion within the SDFT of the carpal canal was accompanied by a mild swelling of the palmar carpal region in most cases. Diagnostic anesthesia of the ulnar nerve, when performed, abolished the lameness. Ultrasound evaluation revealed an increased cross-sectional area (CSA) of the superficial digital flexor tendon compared to the contralateral, sound limb and a loss of tendon fiber architecture within the palmar aspect of the affected tendon, most readily identifiable upon longitudinal ultrasound images. Three out of the five horses returned to their previous level of performance, and two were euthanized. A back at the knee carpal region conformation was associated with a poorer prognosis in these cases. An intratendinous fibroma was identified on histopathological analysis of one-horse following euthanazia, which has been recorded in a complementary short communication.

Clinical features, diagnosis, and survival analysis of dogs with glioma.

BACKGROUND: Gliomas in dogs remain poorly understood. OBJECTIVES: To characterize the clinicopathologic findings, diagnostic imaging features and survival of a large sample of dogs with glioma using the Comparative Brain Tumor Consortium diagnostic classification. ANIMALS: Ninety-one dogs with histopathological diagnosis of glioma. METHODS: Multicentric retrospective case series. Signalment, clinicopathologic findings, diagnostic imaging characteristics, treatment, and outcome were used. Tumors were reclassified according to the new canine glioma diagnostic scheme. RESULTS: No associations were found between clinicopathologic findings or survival and tumor type or grade. However, definitive treatments provided significantly (P = .03) improved median survival time (84 days; 95% confidence interval [CI], 45-190) compared to palliative treatment (26 days; 95% CI, 11-54). On magnetic resonance imaging (MRI), oligodendrogliomas were associated with smooth margins and T1-weighted hypointensity compared to astrocytomas (odds ratio [OR], 42.5; 95% CI, 2.42-744.97; P = .04; OR, 45.5; 95% CI, 5.78-333.33; P < .001, respectively) and undefined gliomas (OR, 84; 95% CI, 3.43-999.99; P = .02; OR, 32.3; 95% CI, 2.51-500.00; P = .008, respectively) and were more commonly in contact with the ventricles than astrocytomas (OR, 7.47; 95% CI, 1.03-53.95; P = .049). Tumor spread to neighboring brain structures was associated with high-grade glioma (OR, 6.02; 95% CI, 1.06-34.48; P = .04). CONCLUSIONS AND CLINICAL IMPORTANCE: Dogs with gliomas have poor outcomes, but risk factors identified in survival analysis inform prognosis and the newly identified MRI characteristics could refine diagnosis of tumor type and grade.

Pharmacological Activation of Nrf2 Enhances Functional Liver Regeneration.

BACKGROUND AND AIMS: The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) regulates an array of cytoprotective genes, yet studies in transgenic mice have led to conflicting reports on its role in liver regeneration. We aimed to test the hypothesis that pharmacological activation of Nrf2 would enhance liver regeneration. APPROACH AND RESULTS: Wild-type and Nrf2 null mice were administered bardoxolone methyl (CDDO-Me), a potent activator of Nrf2 that has entered clinical development, and then subjected to two-thirds partial hepatectomy. Using translational noninvasive imaging techniques, CDDO-Me was shown to enhance the rate of restoration of liver volume (MRI) and improve liver function (multispectral optoacoustic imaging of indocyanine green clearance) in wild-type, but not Nrf2 null, mice following partial hepatectomy. Using immunofluorescence imaging and whole transcriptome analysis, these effects were found to be associated with an increase in hepatocyte hypertrophy and proliferation, the suppression of immune and inflammatory signals, and metabolic adaptation in the remnant liver tissue. Similar processes were modulated following exposure of primary human hepatocytes to CDDO-Me, highlighting the potential relevance of our findings to patients. CONCLUSIONS: Our results indicate that pharmacological activation of Nrf2 is a promising strategy for enhancing functional liver regeneration. Such an approach could therefore aid the recovery of patients undergoing liver surgery and support the treatment of acute and chronic liver disease.

Two-Year Follow-Up of 4-mm-Long Implants Used as Distal Support of Full-Arch FDPs Compared to 10-mm Implants Installed after Sinus Floor Elevation. A Randomized Clinical Trial.

BACKGROUND: In edentulous patients, bone resorption cannot allow the installation of standard implants and it is demanded to use short implants in the residual alveolar bone or longer implants in grafted bone. AIM: To compare the survival and bone level changes of standard plus short 4-mm implants used as distal support of a maxillary full-arch fixed dental prostheses (FDPs) with standard (10-mm) implants placed in association with a bilateral sinus floor augmentation procedure. MATERIAL AND METHODS: Full-arch FDPs supported by six implants were randomly placed in both groups. In the control group, all implants were 10 mm long and 4.1 mm in diameter. The distal implant in both sides of the maxilla was installed after 4 months from bilaterally sinus floor elevation. In the test group (short group), the distal implant in both sides of the maxilla was 4 mm long and 4.1 mm in diameter. No sinus floor elevations were performed in the test group. Clinical assessments and X-rays were taken at prosthesis delivering and after 6, 12, 18, and 24 months. Patient-reported outcome measures (PROMs) were also evaluated before surgery and after 6, 12, and 24 months. RESULTS: The changes over time of the bone level for the short implants were -0.01 ± 0.11 mm, -0.04 ± 0.13 mm, -0.17 ± 0.29 mm, and -0.28 ± 0.37 mm after 6, 12, 18, and 24 months from prosthesis delivering, respectively. For the standard implants, bone changes were -0.21 ± 0.33 mm (p = 0.103), -0.30 ± 0.32 mm (p = 0.023), -0.40 ± 0.37 mm (p = 0.144), and -0.54 ± 0.49 mm (p = 0.128), respectively. A statistically relevant difference was found only at 12 months after loading between the two groups. CONCLUSIONS: Similar results on implant survival rate and marginal bone loss were observed for the short and standard implants, placed in association with a bilateral sinus floor augmentation procedure, used as distal support of a maxillary full-arch FDP. A statistically relevant difference was found only at 12 months after loading between the two groups (p = 0.023).

Cranial solitary osseous plasmacytoma and subjacent amyloid deposition in a dog.

Solitary osseous plasmacytomas affecting the vertebrae, the zygomatic arch, and ribs occur in dogs. In this report, we describe clinical and imaging features of a solitary osseous plasmacytoma affecting the skull with deposition of amyloid forming a mass-like lesion. To the authors' knowledge, no similar cases have been reported before.

Gastric Intravascular Lymphoma in a Dog: Case Report and Literature Review.

Intravascular lymphoma (IVL) is a rare, high-grade, extranodal lymphoma characterized by selective proliferation of neoplastic lymphocytes within the lumen of small vessels. A 10 yr old female intact mixed-breed dog was presented with a 7 mo history of vomiting and anorexia. Physical examination revealed abdominal discomfort. Ultrasonography and endoscopy identified a submucosal gastric mass. Excision was performed by partial gastrectomy and histopathology and immunohistochemistry confirmed a T-cell IVL. The owner declined chemotherapy, and the dog was instead treated palliatively with prednisolone. Two months after surgery, vomiting recurred and abdominal ultrasonography revealed a large gastric ulcer with focal peritonitis. The dog was euthanized 4 mo after initial presentation and postmortem examination confirmed IVL recurrence in the stomach and an isolated nodule of neoplastic cells in the omentum. No involvement of other organs was found following histopathological examination. This is the first description of primary gastric intravascular lymphoma causing chronic vomiting in a dog.

Long-term follow-up of single crowns supported by short, moderately rough implants-A prospective 10-year cohort study.

OBJECTIVES: To evaluate prospectively the clinical and radiographic outcomes after ten years of short (6 mm) implants with a moderately rough surface supporting single crowns in the posterior region. MATERIAL AND METHODS: Forty 6 mm modified sandblasted large-grit acid-etched (mod-SLA), soft tissue level implants were installed in the distal segments of 35 consecutive patients. After 6 weeks of healing, abutments were tightened, and single porcelain-fused-to-metal crowns were cemented. Implant survival, marginal bone loss, and clinical crown/implant ratio were evaluated at various time intervals up to 10 years after loading. RESULTS: Two out of the 40 implants were lost before loading, one implant was lost after 7 years because of peri-implantitis. One patient with two implants died and was excluded from analysis. Two patients did not come at the 10-year follow-up and were considered as drop out (2 implants). The survival rate was 91.7% (n = 36). Thirty-three implants were available for marginal bone loss evaluation. A mean marginal bone loss after 10 years of function was 0.8 ± 0.7 mm. Between 5 and 10 years, the loss was 0.2 ± 0.4 mm. No technical complications were registered during the 10-year period. The clinical crown/implant ratio increased with time from 1.6 at the delivery of the prosthesis to 2.0 after 10 years of loading with no increase between 5 and 10 years. CONCLUSION: Short (6 mm) implants with a moderately rough surface supporting single crowns in the posterior region and loaded after 6-7 weeks maintained full function for at least 10 years with low marginal bone resorption.

Introducing forensic entomology in cases of suspected animal neglect.

Cases of arthropod-infested, abandoned or abused animals are sometimes brought to the attention of veterinarians by animal welfare authorities, with the requirement for a full postmortem examination towards criminal or civil proceedings. In these situations, entomology is an important support tool for the pathologists' investigation since the presence of arthropod life cycle stages serve as reliable forensic markers, especially for blowflies which form the first waves of activity following death. In the present study, 70 cadavers from a total of 544 referred to the Institute of Veterinary Science, University of Liverpool, between 2009 and 2014 displayed evidence of infestation. Here, the authors introduce principles of applied entomology and simplified approaches for estimating the minimum time since death, relevant in the context of routine submissions and the broad remit of individual cases. Despite often limited availability of scene of the crime and local thermal data, the interpretation of the minimum postmortem interval has nonetheless proved valuable as an adjunct to the expert pathology report. However, future developments and enhanced accuracy in this area of animal welfare require resource and training in expertise, and agreed standardisation of both laboratory and field procedures.

Molecular isoforms of high-mobility group box 1 are mechanistic biomarkers for epilepsy.

Approximately 30% of epilepsy patients do not respond to antiepileptic drugs, representing an unmet medical need. There is evidence that neuroinflammation plays a pathogenic role in drug-resistant epilepsy. The high-mobility group box 1 (HMGB1)/TLR4 axis is a key initiator of neuroinflammation following epileptogenic injuries, and its activation contributes to seizure generation in animal models. However, further work is required to understand the role of HMGB1 and its isoforms in epileptogenesis and drug resistance. Using a combination of animal models and sera from clinically well-characterized patients, we have demonstrated that there are dynamic changes in HMGB1 isoforms in the brain and blood of animals undergoing epileptogenesis. The pathologic disulfide HMGB1 isoform progressively increased in blood before epilepsy onset and prospectively identified animals that developed the disease. Consistent with animal data, we observed early expression of disulfide HMGB1 in patients with newly diagnosed epilepsy, and its persistence was associated with subsequent seizures. In contrast with patients with well-controlled epilepsy, patients with chronic, drug-refractory epilepsy persistently expressed the acetylated, disulfide HMGB1 isoforms. Moreover, treatment of animals with antiinflammatory drugs during epileptogenesis prevented both disease progression and blood increase in HMGB1 isoforms. Our data suggest that HMGB1 isoforms are mechanistic biomarkers for epileptogenesis and drug-resistant epilepsy in humans, necessitating evaluation in larger-scale prospective studies.