Cardiorespiratory Effects and Desflurane Requirement in Dogs Undergoing Ovariectomy after Administration Maropitant or Methadone.

General anesthesia for ovariectomy in dogs is based on a balanced anesthesia protocol such as using analgesics along with an inhalant agent. While opioids such as fentanyl and methadone are commonly used for their analgesic potency, other drugs can also have analgesic effects. Maropitant, an antiemetic for dogs and cats, has also been shown to exert analgesic effects, especially on visceral pain. The aim of this study was to compare the cardiorespiratory effects and analgesic properties of maropitant and methadone combined with desflurane in dogs undergoing ovariectomy. Two groups of 20 healthy mixed-breeds bitches undergoing elective ovariectomy received intravenous either maropitant at antiemetic dose of 1 mg kg-1 or methadone at the dose of 0.3 mg kg-1. Cardiorespiratory variables were collected before premedication, 10 min after sedation and during surgery. Recovery quality and postoperative pain were evaluated 15, 30, 60, 120, 240 and 360 min postoperatively. Results showed that maropitant produced analgesia and reduced the requirement of desflurane in amounts similar to those determined by methadone (5.39 ± 0.20% and 4.91 ± 0.26%, respectively) without significant difference, while maintaining heart rate, arterial blood pressure, respiratory rate and carbon dioxide end-tidal partial pressure even at a more satisfactory level. Therefore, maropitant may be recommended as an analgesic drug for abdominal surgery not only in healthy dogs but also in those with reduced cardiorespiratory compensatory capacities or at risk of hypotension, especially when combined with a sedative such as dexmedetomidine.

Characterization of the Sarcidano Horse Coat Color Genes.

The goal of this study was to contribute to the general knowledge of the Sarcidano Horse, both by the identification of the genetic basis of the coat color and by updating the exact locations of the genotyping sites, based on the current EquCab3.0 genome assembly version. One-hundred Sarcidano Horses, living in semi-feral condition, have been captured to perform health and biometric checks. From that total number, 70 individual samples of whole blood were used for DNA extraction, aimed to characterize the genetic basis of the coat color. By genotyping and sequencing analyses of the MC1R Exon 1 and ASIP Exon 3, a real image of the coat color distribution in the studied population has been obtained. Chestnut and Black resulted in the most representative coat colors both from a phenotypic and genotypic point of view, that is suggestive of no human domestication or crossbreeding with domestic breed. Due to its ancient origin and genetic isolation, an active regional plan for the conservation of this breed would be desirable, focused on maintenance of resident genotypes and genetic resources. Collection and management of DNA, sperm, embryos, with the involvement of research centers and Universities, could be a valid enhancing strategy.

Strain Elastography of Injured Equine Superficial Digital Flexor Tendons: A Reliability Study of Manual Measurements.

Early diagnosis of tendon injuries and accurate long-term monitoring of the healing process are key for equine veterinarians that use conventional ultrasonography. The development of strain elastography could improve the management of clinical cases. The aim of the study was to assess the intraobserver repeatability and interobserver reproducibility of manual measurements of the colored areas of the tendons within elastograms and to standardize this manual modality by comparing the analysis of the images with ImageJ. Twenty elastograms of the injured superficial digital flexor tendons (SDFTs) of horses were analyzed by two different operators after an acute injury was diagnosed with ultrasonography. Statistical analysis demonstrated excellent intraobserver repeatability (intraclass correlation coefficient, ICC = 0.949) and good interobserver reproducibility (ICC = 0.855) for manual measurements performed with tools available on the ultrasound unit. A good agreement between manual measurements and measurements performed with ImageJ (ICC = 0.849) was then demonstrated. Despite its subjectivity, the manual modality proved to be a valid method for analyzing images obtained with strain elastography.

Histological and Immunohistochemical Features of Trichoblastoma in a Sarda Breed Sheep.

Skin tumors with adnexal differentiation are commonly reported in dogs and cats, while only anecdotal evidence is available in sheep. Here we illustrate the macroscopic, histologic, and immunohistochemical features of a cutaneous lesion with adnexal differentiation in a 6-year-old female Sarda breed sheep, surgically treated for a horn-like mass located in the left pinna. Additionally, we investigate a possible contribution of Ovine Papillomaviruses (OaPVs). Histologically, the dermis was expanded by an expansive and unencapsulated multilobulated nodule composed of cuboidal to spindle basaloid cells arranged in variably-sized cytokeratins (CK) AE1-AE3, CK 5/6 and CK 34 beta E12, p63-positive winding cords with a characteristic palisade arrangement of neoplastic cells in the periphery of the tumor. Based on these results, the cutaneous neoplasm was diagnosed as a trabecular trichoblastoma with spindle cells and rare structures resembling papillary mesenchymal bodies. Additionally, multiple enlarged sebaceous lobules clustered around dilated ducts suggestive of sebaceous gland hyperplasia were detected near the trichoblastoma. No PV DNA was found in the examined tissues, suggesting that ovine PVs are not involved in the pathogenesis of the present skin tumors with adnexal differentiation. Further investigations and efforts are required to elucidate the prevalence of skin tumors with adnexal differentiation in this species.

Sheep embryonic stem-like cells engrafted into sheep femoral condyle osteochondral defects: 4-year follow-up.

BACKGROUND: Articular cartilage lacks a regenerative response. Embryonic stem cells (ESCs) are a source of pluripotent cells for cartilage regeneration. Their use, however, is associated with a risk of teratoma development, which depends on multiple factors including the number of engrafted cells and their degree of histocompatibility with recipients, the immunosuppression of the host and the site of transplantation. Colonies of sheep embryonic stem-like (ES-like) cells from in vitro-produced embryos, positive for stage-specific embryonic antigens (SSEAs), alkaline phosphatase (ALP), Oct 4, Nanog, Sox 2 and Stat 3 gene expression, and forming embryoid bodies, were pooled in groups of two-three, embedded in fibrin glue and engrafted into osteochondral defects in the left medial femoral condyles of 3 allogeneic ewes (ES). Empty defects (ED) and defects filled with cell-free glue (G) in the condyles of the controlateral stifle joint served as controls. After euthanasia at 4 years post-engraftment, the regenerated tissue was evaluated by macroscopic, histological and immunohistochemical (collagen type II) examinations and fluorescent in situ hybridization (FISH) assay to prove the ES-like cells origin of the regenerated tissue. RESULTS: No teratoma occurred in any of the ES samples. No statistically significant macroscopic or histological differences were observed among the 3 treatment groups. FISH was positive in all the 3 ES samples. CONCLUSIONS: This in vivo preclinical study allowed a long-term evaluation of the occurrence of teratoma in non-immunosuppressed allogeneic adult sheep engrafted with allogeneic ES-like cells, supporting the safe and reliable application of ES cells in the clinic.

Development of a Device-Assisted Nerve-Regeneration Procedure in Disruptive Lesions of the Brachial Plexus.

BACKGROUND: We describe the development of a new surgical procedure to be used in the treatment of disruptive brachial plexus (BP) lesions. It is centered on an artificial device designed to assist nerve regeneration by providing a confined and protected environment. Nerve fibers can repair inside the device, while the adverse massive scar-tissue formation is limited to the outside of the device. METHODS: Steps in the development of the procedure were (1) definition of the rationale, (2) design of the device, (3) choice of an in vivo translational model, (4)refinement of the surgical procedure, and (5) performance of an in vivo pilot study as a proof of concept. An interdisciplinary team from several laboratories was involved in this work over a period of 6 years. RESULTS: Results showed the absence of significant scar tissue in the regenerate and the presence of myelinated fibers aligned proximodistally between the stumps. This surgical approach can be seen not only as a definitive treatment but also as an early examination and stabilization before some different surgery will be later performed. It may also be used as additional protection for traditional surgery like end-to-end coaptation. CONCLUSIONS: We conclude that the availability of a suitable device-assisted early treatment, even if not to be considered definitive, could help in addressing the BP lesions at an earlier stage and this may improve the final outcome. Our evidence justifies further experimentation on this approach.

Second Generation Needling Techniques for the Treatment of Chondral Defects in Animal Model.

Purpose To compare the macroscopic, histological, and immunohistochemical characteristics of the repair tissue of chondral defects treated with microfracture and nanofracture in an ovine model. Methods Full-thickness chondral lesions were created in the medial femoral condyle of both knees in four adult sheep and were treated with microfracture on one side and with nanofracture on the contralateral side. Chondral repair was assessed after 12 months by macroscopic, histological, and immunohistochemical analyses. Results Histological cartilage repair significantly improved in the samples treated with nanofracture for cellular morphological characteristics and cartilage architecture. The immunohistochemical analysis showed a significantly higher immunoreactivity to type II collagen in the defects treated with nanofracture. Conclusion Nanofracture provided better repair tissue than microfracture, with a more satisfactory cartilage architecture renovation and tissue having greater type II collagen content. Clinical Relevance Mesenchymal stem cell stimulation is the most frequently used primary cartilage repair procedure. Nanofracture represents a novel technique to stimulate bone marrow that results into a successful repair of chondral defects.

Host cell tropism, genome characterization, and evolutionary features of OaPV4, a novel Deltapapillomavirus identified in sheep fibropapilloma.

Investigating papillomavirus (PV) diversity is crucial to fully comprehend pathogenicity, genetic features, and evolution of taxa hosted by domestic and wild animal species. This study reports the identification of OaPV4, a novel ovine PV type within Deltapapillomaviruses 3. The study of OaPV4 genomic features combined to in situ hybridization and immunohistochemistry investigations allowed extrapolating several general biological features of ovine PVs, such as their cellular tropism, pathogenicity, and evolutionary history. Based on results, ovine PVs can be grouped into a polyphyletic ancient group of viruses, which splits in two main subgroups having peculiar cellular tropism and pathogenicity. Results add up to animal PV diversity and are crucial to future studies aimed to investigate the correlation between animal PV and cutaneous benign and malign proliferations.

The use of embryonic cells in the treatment of osteochondral defects of the knee: an ovine in vivo study.

PURPOSE: the aim of this study was to determine whether local delivery of embryonic stem-like (ESL) cells into osteochondral defects in the femoral condyles of sheep would enhance regeneration of hyaline articular cartilage. METHODS: male ESL cells embedded in fibrin glue were engrafted into osteochondral defects in the medial condyles (ESL-M) of the left femur in 22 ewes. An identical defect was created in the medial condyle of the contralateral stifle joint and left untreated as a control (empty defect, ED). The ewes were divided into 5 groups. Four sheep each were euthanized at 1, 2, 6, and 12 months from surgery, and 6 ewes were euthanized 24 months post-implantation. To study the effect of varying loads on the long-term regeneration process, an identical defect was also created and ESL cell engraftment performed in the lateral condyle (ESL-L) of the left stifle joint of the animals in the 12- and 24-month groups. The evaluation of regenerated tissue was performed by biomechanical, macroscopic, histological, immunohistochemical (collagen type II) and fluorescent in situ hybridization (FISH) assays. RESULTS: no significant differences were found between treated and control sites in the biomechanical assays at any time point. ESL cell grafts showed significantly greater macroscopic evidence of regeneration as compared to controls at 24 months after surgery; significantly better histological evidence of repair in ESL-M samples versus controls was found throughout the considered period. At 24 months from surgery there was significantly improved integration of graft edges with the host tissue in the ESL-M as compared to the ESL-L samples, demonstrating that load bearing positively affects the long-term regeneration process. CONCLUSIONS: ESL cells enhanced the regeneration of hyaline cartilage. FISH confirmed that the regenerative tissue originated from ESL cells. CLINICAL RELEVANCE: ESL cells are able to self-renew for prolonged periods without differentiation and, most importantly, to differentiate into a large variety of tissues.

Subchondral bone remodeling: comparing nanofracture with microfracture. An ovine in vivo study.

PURPOSE: microfracture, providing direct stimulation of chondrogenic mesenchymal stem cells (MSCs) in the subchondral bone, remains the most frequently used primary cartilage repair technique. However, the newly formed type I collagen-rich fibrocartilaginous tissue has poor biomechanical properties and a tendency to degenerate. To overcome these limitations the nanofracture technique was introduced. Our purpose was to compare subchondral bone remodeling 6 months after microfracture versus nanofracture (subchondral needling) treatment in an ovine model. METHODS: full-thickness chondral lesions were created in the load-bearing area of the medial femoral condyles in four adult sheep. Each animal was then treated on one side with microfracture and on the contralateral side with nanofracture. Subchondral bone remodeling was assessed by micro-CT using a Bruker(®) SKYSCAN and CTVOX 2.7 software (Bruker Corp., Billerica, MA, USA) for image reconstruction; trabecular bone density measurements were performed through a color-representation structure thickness analysis. RESULTS: at the six-month endpoint, the microfracture-treated samples showed limited perforation depth and cone-shaped channels with large diameters at the joint surface. The channel walls displayed a high degree of regularity with significant trabecular bone compaction leading to a sealing effect with limited communication with the surrounding trabecular canals. Condyles treated with nanofracture showed channels characterized by greater depth and smaller diameters and natural irregularities of the channel walls, absence of trabecular compaction around the perforation, remarkable communication with trabecular canals, and neo-trabecular remodeling inside the channels. CONCLUSIONS: nanofracture is an effective and innovative repair technique allowing deeper perforation into subchondral bone with less trabecular fragmentation and compaction when compared to microfracture; it results in better restoration of the normal subchondral bone architecture at six months. CLINICAL RELEVANCE: our data support the use of smaller-diameter and deeper subchondral bone perforation for MSC stimulation; this technique may prove to be an attractive alternative to standard microfracture procedures.

Magnetic resonance imaging features of progressive ethmoid hematoma in 2 horses.

Progressive ethmoid hematoma is frequently reported among the pathologies of the upper airways in horses. While several hypotheses have been proposed such as repeated trauma, inflammatory processes and/or chronic infections, congenital phenomena, complication of hemangiomas, and neoplastic processes, the aetiology of this condition is yet unknown. Clinical symptoms are non-specific and includes haemorrhagic nasal discharge, respiratory noise not related to physical exercise and, in most severe cases, phenomena of coughing. The purpose of this case report is to contribute to a correct diagnostic approach to this particular pathology of upper respiratory tract in horses. Magnetic Resonance allows for a very detailed imaging of the development of the disease and suggests appropriate therapeutic choices.

Treatment with embryonic stem-like cells into osteochondral defects in sheep femoral condyles.

BACKGROUND: Articular cartilage has poor intrinsic capacity for regeneration because of its avascularity and very slow cellular turnover. Defects deriving from trauma or joint disease tend to be repaired with fibrocartilage rather than hyaline cartilage. Consequent degenerative processes are related to the width and depth of the defect. Since mesenchymal stem cells (MSCs) deriving from patients affected by osteoarthritis have a lower proliferative and chondrogenic activity, the systemic or local delivery of heterologous cells may enhance regeneration or inhibit the progressive loss of joint tissue. Embryonic stem cells (ESCs) are very promising, since they can self-renew for prolonged periods without differentiation and can differentiate into tissues from all the 3 germ layers. To date only a few experiments have used ESCs for the study of the cartilage regeneration in animal models and most of them used laboratory animals. Sheep, due to their anatomical, physiological and immunological similarity to humans, represent a valid model for translational studies. This experiment aimed to evaluate if the local delivery of male sheep embryonic stem-like (ES-like) cells into osteochondral defects in the femoral condyles of adult sheep can enhance the regeneration of articular cartilage. Twenty-two ewes were divided into 5 groups (1, 2, 6, 12 and 24 months after surgery). Newly formed tissue was evaluated by macroscopic, histological, immunohistochemical (collagen type II) and fluorescent in situ hybridization (FISH) assays. RESULTS: Regenerated tissue was ultimately evaluated on 17 sheep. Samples engrafted with ES-like cells had significantly better histologic evidence of regeneration with respect to empty defects, used as controls, at all time periods. CONCLUSIONS: Histological assessments demonstrated that the local delivery of ES-like cells into osteochondral defects in sheep femoral condyles enhances the regeneration of the articular hyaline cartilage, without signs of immune rejection or teratoma for 24 months after engraftment.

Repeated platelet concentrate injections enhance reparative response of microfractures in the treatment of chondral defects of the knee: an experimental study in an animal model.

PURPOSE: To assess the histology and biomechanics of repair cartilage after microfractures with and without repeated local injections of platelet concentrate for the treatment of full-thickness focal chondral defects of the knee. METHODS: A full-thickness chondral lesion on the medial femoral condyle was created in 30 sheep and treated with microfractures. Animals were divided into 2 groups, according to postoperative treatment: in group 1 we performed 5 weekly injections of autologous conditioned plasma, whereas group 2 did not undergo further treatments. Animals were killed at 3, 6, and 12 months after treatment. Macroscopic, histologic, and biomechanical evaluations were performed. Differences between groups at each time interval and differences over time within groups were analyzed for each outcome. Significance was set at P < .05. RESULTS: Group 1 showed significantly better macroscopic, histologic, and biomechanical results than group 2 at each time interval. Analysis of time effect within groups showed that in group 1, quality of repair tissue significantly improved from 3 to 6 months after treatment and remained stable over time for all the outcomes; in group 2 a significant histologic and mechanical deterioration was observed between 6 and 12 months' follow-up. CONCLUSIONS: Five repeated local injections of autologous conditioned plasma after microfractures in the treatment of full-thickness cartilage injuries promoted a better and more durable reparative response than isolated microfractures, although they did not produce hyaline cartilage. CLINICAL RELEVANCE: Periodical intra-articular injections of platelet concentrate after microfractures may improve cartilage repair and prevent further degenerative changes.

Sheep embryonic stem-like cells transplanted in full-thickness cartilage defects.

Articular cartilage regeneration is limited. Embryonic stem (ES) cell lines provide a source of totipotent cells for regenerating cartilage. Anatomical, biomechanical, physiological and immunological similarities between humans and sheep make this animal an optimal experimental model. This study examines the repair process of articular cartilage in sheep after transplantation of ES-like cells isolated from inner cell masses (ICMs) derived from in vitro-produced (IVP) vitrified embryos. Thirty-five ES-like colonies from 40 IVP embryos, positive for stage-specific embryonic antigens (SSEAs), were pooled in groups of two or three, embedded in fibrin glue and transplanted into osteochondral defects in the medial femoral condyles of 14 ewes. Empty defect (ED) and cell-free glue (G) in the controlateral stifle joint served as controls. The Y gene sequence was used to detect ES-like cells in the repair tissue by in situ hybridization (ISH). Two ewes were euthanized at 1 month post-operatively, three each at 2 and 6 months and four at 12 months. Repairing tissue was examined by biomechanical, macroscopic, histological, immunohistochemical (collagen type II) and ISH assays. Scores of all treatments showed no statistical significant differences among treatment groups at a given time period, although ES-like grafts showed a tendency toward a better healing process. ISH was positive in all ES-like specimens. This study demonstrates that ES-like cells transplanted into cartilage defects stimulate the repair process to promote better organization and tissue bulk. However, the small number of cells applied and the short interval between surgery and euthanasia might have negatively affected the results.