Benign Bilateral Adenomyoepithelioma of the Mammary Gland in a Ring-tailed Lemur (Lemur catta).

Naturally occurring mammary tumours are uncommon in prosimians. A 20-year-old female ring-tailed lemur (Lemur catta) developed bilateral enlargement of the mammary glands. Surgical removal revealed that both masses were comprised of multiple nodules and cystic areas that entirely replaced the normal glands. Histologically, a benign neoplastic biphasic cellular proliferation, composed of luminal-epithelial and basal-myoepithelial components, was identified. Immunohistochemical analysis for expression of cytokeratin (CK) AE1/AE3, CK7, CK5 + 8, CK14, vimentin, p63 and 14-3-3σ highlighted the biphasic nature of the neoplasm. A low mitotic count, low Ki67 labelling index, expression of oestrogen receptor-α, lack of expression of human epidermal growth factor receptor and a 3-year disease-free period without recurrence supported the benign nature of the tumour. Macroscopically, histologically and immunohistochemically this neoplasm resembled benign adenomyoepithelioma of the breast in women. This is the first complete report of a naturally occurring mammary tumour in a ring-tailed lemur.

Muscle Pathology in Free-Ranging Stranded Cetaceans.

Despite the profound impact that skeletal muscle disorders may pose for the daily activities of wild terrestrial and marine mammals, such conditions have been rarely described in cetaceans. In this study, the authors aimed to determine the nature and prevalence of skeletal muscle lesions in small and large odontocetes and mysticetes ( n = 153) from 19 different species. A macroscopic evaluation of the epaxial muscle mass and a histologic examination of the longissimus dorsi muscle were performed in all cases. The only macroscopically evident change was variable degrees of atrophy of the epaxial muscles ( longissimus dorsi, multifidus, spinalis) in emaciated specimens. The histopathological study revealed single or combined morphological changes in 91.5% of the cases. These changes included the following: degenerative lesions (75.2%), muscle atrophy (37.9%), chronic myopathic changes (25.5%), parasitic infestation (9.2%), and myositis (1.9%). The skeletal muscle is easily sampled during a necropsy and provides essential microscopic information that reflects both local and systemic conditions. Thus, skeletal muscle should be systematically sampled, processed, and examined in all stranded cetaceans.

Comparative histology of muscle in free ranging cetaceans: shallow versus deep diving species.

Different marine mammal species exhibit a wide range of diving behaviour based on their breath-hold diving capabilities. They are classically categorized as long duration, deep-diving and short duration, shallow-diving species. These abilities are likely to be related to the muscle characteristics of each species. Despite the increasing number of publications on muscle profile in different cetacean species, very little information is currently available concerning the characteristics of other muscle components in these species. In this study, we examined skeletal muscle fiber type, fiber size (cross sectional area and lesser diameter), intramuscular substrates, and perimysium-related structures, by retrospective study in 146 stranded cetaceans involving 15 different species. Additionally, we investigated diving profile-specific histological features. Our results suggest that deep diving species have higher amount of intramyocyte lipid droplets, and evidence higher percentage of intramuscular adipose tissue, and larger fibre sizes in this group of animals.

Heart regeneration after myocardial infarction using synthetic biomaterials.

Myocardial infarction causes almost 7.3 million deaths each year worldwide. However, current treatments are more palliative than curative. Presently, cell and protein therapies are considered the most promising alternative treatments. Clinical trials performed until now have demonstrated that these therapies are limited by protein short half-life and by low transplanted cell survival rate, prompting the development of novel cell and protein delivery systems able to overcome such limitations. In this review we discuss the advances made in the last 10years in the emerging field of cardiac repair using biomaterial-based delivery systems with focus on the progress made on preclinical in vivo studies. Then, we focus in cardiac tissue engineering approaches, and how the incorporation of both cells and proteins together into biomaterials has opened new horizons in the myocardial infarction treatment. Finally, the ongoing challenges and the perspectives for future work in cardiac tissue engineering will also be discussed.

Morphologic and immunohistochemical features of Merkel cells in the dog.

Merkel cells (MCs) are specialized skin receptors characterized by their particular location and close association with nerve terminals. They also are cells with a presumptive neuroendocrine function and are considered as part of the diffuse neuroendocrine system. By using commercially available monoclonal and polyclonal antibodies in samples of dog skin, MCs were properly distinguished from other clear cell types in the epidermis. They expressed cytokeratins 7, 8, 20, EpCAM, NSE, CGA, SYN, S100 protein, and NF, presented diverse cytological features and arrangements depending on the location considered, and showed pronounced heterogeneity with markedly different expression and distribution patterns for antibodies used. Anti-CK20 presented as the most reliable and specific antibody for their identification. The present study increases our knowledge of MCs and establishes a basis for future studies of the role(s) of the MCs in diseased tissues of the dog skin, including the cutaneous neuroendocrine (Merkel cell) tumour.

Primary central nervous system T-cell lymphoma in a common dolphin (Delphinus delphis).

This report describes the pathological findings in an adult female short-beaked common dolphin (Delphinus delphis) stranded alive in the Canary Islands. Necropsy examination revealed the presence of a nodular neoplastic growth in the central nervous system (CNS) at the level of the thalamus. Microscopical examination revealed the mass to be a lymphoma and immunohistochemical labelling demonstrated a T-cell origin. No significant lesions were observed in other organs, including lymphoid organs. This is the first report of a primary T-cell lymphoma in the CNS in cetaceans.

Biliary cirrhosis caused by Campula spp. in a dolphin and four porpoises.

Biliary cirrhosis produced by Campula spp. is described in 1 striped dolphin Stenella coeruleoalba and 4 harbour porpoises Phocoena phocoena. The hepatic lesions consisted of severe proliferation of fibrous connective tissue with loss of the lobular pattern, nodular regeneration of the hepatic tissue, bile duct hyperplasia and severe inflammatory infiltrate composed of eosinophils, macrophages, lymphocytes and plasma cells. These lesions were associated with severe infestation by Campula spp. Although inflammatory and degenerative hepatic lesions are frequently found in stranded dolphins, biliary cirrhosis has not been previously reported in cetaceans. Massive infestation by these parasites should be included as a cause of hepatic failure resulting in stranding of marine mammals.

Adipose-derived stem cells combined with neuregulin-1 delivery systems for heart tissue engineering.

Myocardial infarction (MI) is the leading cause of death worldwide, and extensive research has therefore been performed to find a cure. Neuregulin-1 (NRG) is a growth factor involved in cardiac repair after MI. We previously described how biocompatible and biodegradable microparticles, which are able to release NRG in a sustained manner, represent a valuable approach to avoid problems related to the short half-life after systemic administration of proteins. The effectiveness of this strategy could be improved by combining NRG with several cytokines involved in cardiac regeneration. The present study investigates the potential feasibility of using NRG-releasing particle scaffold combined with adipose-derived stem cells (ADSC) as a multiple growth factor delivery-based tissue engineering strategy for implantation in the infarcted myocardium. NRG-releasing particle scaffolds with a suitable size for intramyocardial implantation were prepared by TROMS. Next, ADSC were adhered to particle scaffolds and their potential for heart administration was assessed in a MI rat model. NRG was successfully encapsulated reaching encapsulation efficiencies of 92.58 ± 3.84%. NRG maintained its biological activity after the microencapsulation process. ADSCs adhered efficiently to particle scaffolds within a few hours. The ADSC-cytokine delivery system developed proved to be compatible with intramyocardial administration in terms of injectability through a 23-gauge needle and tissue response. Interestingly, ADSC-scaffolds were present in the peri-infarted tissue 2 weeks after implantation. This proof of concept study provides important evidence required for future effectiveness studies and for the translation of this approach.

Biodegradation and heart retention of polymeric microparticles in a rat model of myocardial ischemia.

Poly-lactide-co-glycolide (PLGA) microparticles emerged as one of the most promising strategies to achieve site-specific drug delivery. Although these microparticles have been demonstrated to be effective in several wound healing models, their potential in cardiac regeneration has not yet been fully assessed. The present work sought to explore PLGA microparticles as cardiac drug delivery systems. PLGA microparticles were prepared by Total Recirculation One-Machine System (TROMS) after the formation of a multiple emulsion. Microparticles of different size were prepared and characterized to select the most suitable size for intramyocardial administration. Next, the potential of PLGA microparticles for administration in the heart was assessed in a MI rat model. Particle biodegradation over time and myocardial tissue reaction were studied by routine staining and confocal microscopy. Results showed that microparticles with a diameter of 5 µm were the most compatible with intramyocardial administration in terms of injectability through a 29-gauge needle and tissue response. Particles were present in the heart tissue for up to 3 months post-implantation and no particle migration toward other solid organs was observed, demonstrating good myocardial retention. CD68 immunolabeling revealed 31%, 47% and below 4% microparticle uptake by macrophages 1 week, 1 month, and 3 months after injection, respectively (P<0.001). Taken together, these findings support the feasibility of the developed PLGA microparticles as vehicles for delivering growth factors in the infarcted myocardium.

Use of CD10 as a marker of canine mammary myoepithelial cells.

CD10 is an important cell marker in the diagnosis of acute lymphoblastic leukaemia and of breast myoepithelial (ME) cells in humans. The objective of this study was to assess the value of CD10 as a marker of canine ME cells using immunohistochemistry on routinely processed normal, dysplastic and neoplastic mammary tissue. Five different CD10 positive cell types were identified on the basis of cell morphology, pattern of immunoreactivity, and on the co-expression of additional cell lineage-specific markers. Type 1 cells were typical fusiform cells with a ME cell phenotype (calponin- and cytokeratin [CK] 14-positive, CK8/18-negative). Type 2 cells were typical or atypical polyhedral cells with a luminal epithelial (LE) cell phenotype (calponin- and CK14-negative, CK8/18-positive). Type 3 cells had a type 1 phenotype with variable morphology, and type 4 were atypical neoplastic cells with a mixed ME/LE phenotype. Type 5 cells were typical fusiform cells with a stromal phenotype. Type 1 cells were considered normal ME cells and were found in all sample types; type 2 cells were considered normal or neoplastic LE cells and were also found in all sample types; types 3 and 4 cells were restricted to tumour samples and to malignant tumours, respectively, and type 5 cells were found in all sample types, although predominantly in neoplastic tissue. The findings indicate that the CD10 antigen is a sensitive (although not specific) marker of canine ME cells in normal, dysplastic and neoplastic mammary tissue. Differences in the distribution and staining intensity of CD10-positive cells suggest a number of potential roles for this protein in the pathogenesis of canine mammary neoplasia.

Capture myopathy in live-stranded cetaceans.

A group of 51 cetaceans that had been stranded alive on the coasts of the Canary Islands, experienced human capture/rescue interactions and then died, were necropsied over a 12-year period. Of these cetaceans, 25 had haemodynamic lesions indicative of multiorganic vascular shock, degenerative muscle lesions affecting both skeletal and cardiac muscles and myoglobinuric nephrosis typical of capture myopathy (CM). Because macroscopic lesions in muscles and kidneys were not always obvious, a standard protocol was developed where the longissimus dorsi muscle was examined histologically for segmental hypercontraction, contraction band necrosis and segmental muscular degeneration and cardiomyocytes studied for hypereosinophilic wavy fibres, sarcolemmal and perinuclear vacuolation and contraction band necrosis. Light microscopic skeletal and cardiac muscle lesions in all CM animals were confirmed as ante mortem by immunohistochemical assay for myoglobin loss from and fibrinogen entry into affected myofibres. All animals had tubular nephrosis with casts and tubular myoglobin. The oxidative stress-related marker HSP70 was demonstrated immunohistochemically in tubular epithelium. Although the syndrome related to death of live-stranded cetaceans is multifactorial, this study documents that a clinicopathological syndrome comparable to CM of terrestrial wildlife has a role in stranding outcomes.

Complex polysaccharide inclusions in the skeletal muscle of stranded cetaceans.

Skeletal muscle samples were examined post-mortem in 148 cetaceans over a 12-year period. Histological analysis included haematoxylin and eosin (HE) and periodic acid-Schiff (PAS) staining with and without diastase digestion. In addition, histological muscle sections were immunostained for ubiquitin and fast and slow heavy-chain myosin isoforms. PAS-positive, diastase-resistant inclusions were detected in 26 animals from 11 different species. Older cetaceans were preferentially affected. These intrafibre inclusions varied from large aggregates to multiple coarse granules and were typically associated with type II fibres. All diastase-resistant inclusions were positive for ubiquitin. These features resembled those inclusions described as complex polysaccharide in horses. Based on these histological findings and the ubiquitin staining pattern, a morphological diagnosis of complex polysaccharide storage myopathy is proposed.

Exuberant mucometra associated with atresia of the cervix in a queen.

This short communication reports the clinical, ultrasonographic and histopathological findings in a cat with atresia of the uterine cervix and mucometra. After 6 months of continuous oestrous behaviour, a remarkable abdominal enlargement was observed in a 14-year-old queen. A presumptive diagnosis of mucometra was concluded after the ultrasound evaluation and based on clinical signs and blood analyses. Ovariohysterectomy revealed a notable symmetrical distension (4-5 cm in diameter) of both uterine horns that were filled with fluid (690 ml); microbiological analyses confirmed the aseptic nature of the uterine fluid. Ovarian follicular cysts and cystic subsurface epithelial structures, >1.5 cm in diameter, were present in both ovaries and no corpora lutea were observed. Gross and microscopic evaluation of the uterus confirmed the development of cystic endometrial hyperplasia and the absence of an internal cervical os. The endometrial hyperplasia and mucometra could have developed as a consequence of repeated oestrogenic stimulation.

Weissella ceti sp. nov., isolated from beaked whales (Mesoplodon bidens).

During an investigation into the microbiota of beaked whales (Mesoplodon bidens), nine isolates were obtained from different organs of four animals. The isolates were Gram-positive-staining, catalase-negative, short rod-shaped or coccoid organisms. A phylogenetic analysis based on 16S rRNA gene sequences of these isolates allocated them to the genus Weissella, showing 96.3 % and 96.0 % 16S rRNA gene sequence similarity with Weissella viridescens NRIC 1536(T)and Weissella minor NRIC 1625(T), respectively. On the basis of phenotypic, physiological and phylogenetic evidence, it is proposed that the new isolates from whales represent a novel species of the genus Weissella, Weissella ceti sp. nov. The type strain of Weissella ceti is 1119-1A-09(T) ( = CECT 7719(T) = CCUG 59653(T)).

Recovery of Mycoplasma agalactiae from the ears of goats experimentally infected by the intramammary route.

The role of inapparent carriers of Mycoplasma agalactiae and the strategies used to colonise the external ear canal in goats remain unclear. This study examined the ability of M. agalactiae to colonise the ears of goats infected experimentally by the intramammary route. The right mammary glands of 15 lactating goats were inoculated with 10(10) colony forming units (cfu) of M. agalactiae. The goats were randomly assigned to three groups of five animals each and sampled at slaughter at 5, 15 or 45 days post-infection (dpi). A further four goats served as uninfected controls. Right and left ear swabs were collected for detection of M. agalactiae by culture before and after sacrifice. M. agalactiae was detected in 19/20 (95%) ear swabs from goats sampled at 15 and 45dpi, whereas all ear swabs collected before inoculation, ear swabs collected from the group sampled at 5dpi and ear swabs from control goats at the time of sacrifice were negative for M. agalactiae. Blood samples collected at 6, 12, 24, 48 and 72h post-infection for detection of M. agalactiae by culture were also negative. There were differences in the antigenic profiles of isolates recovered from the ears compared to the 7MAG strain used to inoculate the animals and most isolates from the mammary gland, milk and supramammary lymph nodes.

Congenital biventricular cardiac diverticula in a dog.

Multiple diverticula of the right and left cardiac ventricular walls were incidental findings at necropsy in a juvenile, male, mixed-breed dog with no clinical history of heart disease. Each saccular diverticulum had a narrow communication into the corresponding ventricular chamber. Histologically, the diverticular walls consisted mainly of collagen without elastin fibers and atrophic cardiomyocytes. No inflammation or other lesions were associated with the diverticula or the rest of the myocardium. Based on the history and pathologic findings, a diagnosis of congenital biventricular fibrous diverticula was made.

Cross-reactivity of anti-human, anti-porcine and anti-bovine cytokine antibodies with cetacean tissues.

The cross-reactivity of monoclonal antibodies specific for human, porcine and bovine cytokines was evaluated for three cetacean species: Atlantic spotted dolphins (Stenella frontalis), striped dolphins (Stenella coeruleoalba) and fin whales (Balaenoptera physalus). Formalin-fixed and snap-frozen tissue sections of lung, spleen, liver and mesenteric lymph node were evaluated. T and B lymphocytes and monocytes/macrophages were detected by use of anti-human CD3, IgG and lysozyme polyclonal antibodies (pAbs), respectively. These reagents were successfully applied to both fixed and frozen tissues. Anti-human interleukin (IL)-1 alpha, IL-1 beta, IL-8, tumour necrosis factor (TNF)-alpha and CD25, anti-porcine IL-2, IL-6, IL-10, and anti-bovine IL-4 and interferon (IFN)-gamma antibodies produced immunolabelling in cetacean snap-frozen lymph node sections similar to that obtained with tissue from the species of origin, but they did not react with formalin-fixed tissue sections. Anti-porcine IL-12 pAb did not react with snap-frozen cetacean tissue samples. Macrophages and lymphocytes were the most common cells immunolabelled with the anti-cytokine antibodies. This panel of anti-cytokine antibodies may be used to evaluate cytokine expression in snap-frozen tissue samples from the cetacean species tested.

Morphological and immunohistochemical characterization of spontaneous mammary gland tumors in the guinea pig (Cavia porcellus).

Ten spontaneous mammary gland tumors affecting guinea pigs (GP) were analyzed histologically and immunohistochemically. Histologically, 3 were benign (2 simple adenomas and 1 benign mixed tumor) and 7 were malignant (1 simple solid carcinoma and 6 simple tubulopapillary carcinomas). Immunohistochemical data revealed the glandular immunoprofile of all the tumors and suggested their ductal origin on the basis of cytokeratin 20 expression. Interestingly, cytokeratin 7 was detected in basal/myoepithelial cells. Further, all tumors were positive for type alpha estrogen and progesterone receptors, suggesting a role for steroid hormones in the development of these neoplasias in GP. This article describes the morphological and immunohistochemical features of the normal mammary gland and spontaneous mammary gland tumors in GP.

Correlating the immune response with the clinical-pathological course of persistent mastitis experimentally induced by Mycoplasma agalactiae in dairy goats.

To correlate the clinical course of mycoplasma mastitis with its immune response, right mammary glands of 15 lactating goats were inoculating with 10(10) colony-forming units (cfu) of Mycoplasma agalactiae (Ma). Before sacrificing the animals at 5, 15 or 45 days post-inoculation (dpi), blood Ma antibody titres and milk mycoplasma colony and somatic cell counts were monitored. Ma colonised the mammary gland and milk counts increased to over 10(12)cfu/ml within 5 dpi. During this period, an innate immune response involving neutrophils and macrophages was observed, and Ma antigen appeared in the degenerated acinar epithelium. From 7 dpi, a specific antibody response coincided with reduced viable mycoplasmas in milk. The humoral immune response was limited; by 37 dpi, all animals scored negative for anti-Ma antibodies, and around 10(8)cfu/ml were shed. Results indicate an early immune response to Ma inoculation unable to control mycoplasmal invasion. An ensuing humoral response, despite reducing the mycoplasma burden, leads to chronic, persistent infection.