Healthy Equine Research Articles

Intraocular lens power calculation for the equine eye

BackgroundPhacoemulsification and intraocular lens (IOL) implantation during cataract surgery in horses occur with increasing frequency. To reduce the postoperative refractive error it is necessary to determine the proper IOL power. In the present study retinoscopy, keratometry and ultrasonographic biometry were performed on 98 healthy equine eyes from 49 horses. The refractive state, corneal curvature (keratometry) and the axial location of all optical interfaces (biometry) were measured. The influences of breed, height at the withers, gender and age on values obtained and the comparison between the left and right eye were evaluated statistically. Corresponding IOL power were calculated by use of Binkhorst and Retzlaff theoretical formulas.ResultsMean ± SD refractive state of the horses was + 0.32 ± 0.66 D. Averaged corneal curvature for Haflinger, Friesian, Pony, Shetland pony and Warmblood were 21.30 ± 0.56 D, 20.02 ± 0.60 D, 22.61 ± 1.76 D, 23.77 ± 0.94 D and 20.76 ± 0.88 D, respectively. The estimated postoperative anterior chamber depth (C) was calculated by the formula C = anterior chamber depth (ACD)/0.73. This formula was determined by a different research group. C and axial length of the globe averaged for Haflinger 9.30 ± 0.54 mm and 39.43 ± 1.26 mm, for Friesian 10.12 ± 0.33 mm and 42.23 ± 1.00 mm, for Pony 8.68 ± 0.78 mm and 38.85 ± 3.13 mm, for Shetland pony 8.71 ± 0.81 mm and 37.21 ± 1.50 mm and for Warmblood 9.39 ± 0.51 mm and 40.65 ± 1.30 mm. IOL power was calculated with the Binkhorst and Retzlaff theoretical formulas. Calculated IOL power for the several breeds ranged from 18.03 D to 19.55 D. The mean value across all horses was 18.73 D determined with Binkhorst formula and 18.54 D determined with Retzlaff formula.ConclusionsMean result of this study is: an 18.5 D IOL seemed to be the most appropriate to achieve emmetropia after IOL implantation in horses. Cataract surgery without IOL implantation results in hyperopic and visual compromised horses. Retinoscopy, keratometry and ultrasonographic biometry should be performed on every affected horse and postoperative visual outcome should be determined.

Allogeneic mesenchymal stem cell transplantation in healthy equine superficial digital flexor tendon: A study of the local inflammatory response

The superficial digital flexor tendon (SDFT) is a structure frequently affected by injuries in high-performance athletic horses, and there are limited therapeutic options. Regenerative medicine has evolved significantly in treating different illnesses. However, understanding the cellular behaviour during mesenchymal stem cell (MSC) transplantation in healthy tissues is not fully known yet. To address the inflammatory response induced by allogeneic MSC transplantation, this study evaluated the local inflammatory response after the application of allogeneic adipose tissue-derived mesenchymal stem cells (AT-MSCs) in the equine tendon compared to an autologous transplant and the control group. Eighteen thoracic limbs (TL) in nine animals were divided into three groups and subjected to the application of AT-MSCs in the healthy tendon. In the allogeneic group (Gallog), the animals received an allogeneic AT-MSC application in the TL. The autologous group (Gauto) received an application of autologous cells in the TL, and in the control group (Gcont), phosphate-buffered saline (PBS) was applied. There were no significant differences among the evaluated groups in the physical, morphological, thermography, and ultrasonography analyses. A higher number of CD3-positive lymphocytes was observed in the Gauto group compared to the control (P < 0.05). Additionally, we did not observe different expressions of CD172 and microvascular density among the groups. The allogeneic transplantation of AT-MSCs did not result in an adverse or inflammatory reaction that compromised the use of these cells in this experiment. Their behaviour was similar to that of autologous transplantation.

Pharmacokinetic evaluation and safety of topical 1% morphine sulfate application on the healthy equine eye.

To determine if corneal epithelial cell integrity is detrimentally affected by short-term administration of 1.0% morphine sulfate. Additionally, we sought to determine if topical 1.0% morphine applied to the equine cornea would result in ocular or systemic absorption. Six healthy horses. Morphine sulfate (1.0%) was applied topically to one eye every four hours for 72 h before horses were euthanized. Serum samples were collected at varying time points during the study and aqueous and vitreous humor were collected immediately after euthanasia. Morphine quantification in serum, aqueous, and vitreous humor was performed by ELISA. Treated and control corneas were submitted for histopathology. Horses were monitored for adverse ocular and systemic effects throughout the study period. All horses developed mild mucoid ocular discharge in the treated eye. One horse developed a fever during treatment. Morphine was detected in the aqueous humor of the treated eye for all horses with mean ± standard deviation of 165.18 ng/mL ± 87.69 ng/mL. Morphine was detected in vitreous humor of the treated eye of 5 of 6 horses with mean ± standard deviation of 4.87 ± 4.46 ng/mL. Morphine was detected in the serum of 5 of 6 horses at varying time points. Maximum systemic concentration reached in a single horse was 6.98 ng/mL. Corneal histopathology revealed no difference in microscopic appearance between morphine-treated and control corneas. Topical administration of 1.0% morphine sulfate did not appear to cause any significant ocular or systemic adverse effects. Topical ophthalmic morphine application resulted in both ocular and systemic absorption.

The regulatory role of the erythrocytic membranes’ lipid components in ensuring oxygen-binding capacity of hemoglobin in cardiovascular diseases

Cross-talk of subchondral bone and articular cartilage could be an important aspect in the etiology of osteoarthritis. Previous research has provided some evidence of transport of small molecules (~370 Da) through the calcified cartilage and subchondral bone plate in murine osteoarthritis models. The current study, for the first time, uses a neutral diffusing computed tomography (CT) contrast agent (iodixanol, ~1550 Da) to study the permeability of the osteochondral interface in equine and human samples. Sequential CT monitoring of diffusion after injecting a finite amount of contrast agent solution onto the cartilage surface using a micro-CT showed penetration of the contrast molecules across the cartilage-bone interface. Moreover, diffusion through the cartilage-bone interface was affected by thickness and porosity of the subchondral bone as well as the cartilage thickness in both human and equine samples. Our results revealed that porosity of the subchondral plate contributed more strongly to the diffusion across osteochondral interface compared to other morphological parameters in healthy equine samples. However, thickness of the subchondral plate contributed more strongly to the diffusion in slightly osteoarthritic human samples.

Glycosaminoglycans (GAGs) determination in healthy and damaged equine articular cartilage

espanolRESUMEN: El proposito de este estudio fue establecer la existencia de alguna diferencia en el contenido de GAG entre las superficies articulares que soportan y no soportan peso, comparando articulaciones metacarpofalangicas equinas macroscopicamente sanas y danadas. Las muestras de cartilago se obtuvieron desde la superficie de apoyo de articulaciones macroscopicamente sanas (N1; n=10) y de aquellas macroscopicamente danadas (P1; n=10). Adicionalmente, en una localizacion dorsoproximal se obtuvieron muestras de la superficie del cartilago sin apoyo, desde articulaciones sanas (N2; n=10) y danadas (P2; n=10). Los GAG fueron extraidos desde 100 mg de cartilago de cada muestra y cuantificados mediante el metodo safranina-O que mide la carga anionica total y por el metodo del carbazol que mide el contenido de acido uronico. Ambos metodos fueron capaces de medir el contenido de GAG no encontrandose diferencias entre zonas intraarticulacion (1 y 2). Al comparar el contenido de GAG entre articulaciones sanas y danadas, ambos metodos evidenciaron una disminucion significativa de GAG en las articulaciones danadas (1 y 2). Estos resultados muestran que en un proceso patologico articular cronico se afecta el cartilago en su conjunto y no solo aquellas zonas que muestran lesiones macroscopicas al soportar mayor impacto mecanico. EnglishABSTRACT: The purpose of this study was to establish if there was any difference in the GAGs content between loaded and unloaded surfaces of the joint. Furthermore, the results were compared between macroscopically healthy and damaged joints. Cartilage samples were obtained from two different zones of the equine metacarpophalangeal joint (metacarpal condyles). Samples were collected from the loaded surface of macroscopically healthy joints (N1; n=10) and from macroscopically damaged cartilage (P1; n=10). Additionally, cartilage samples were collected from unloaded areas at the most dorso-proximal zone of the joint in macroscopically healthy joints (N2; n=10) and from the macroscopically pathological joints but without damaged cartilage on the site of sampling (P2; n=10). The GAGs were extracted from 100 mg of cartilage of each sample and quantified through the safranine - O method that measured the total anionic charges, and through the carbazole method that measured the uronic acid content. Both methods measured the GAGs content, showing no differences between intra-joint zones (1 and 2), but when the GAGs content was compared between healthy and pathological joints, both methods showed a significantly decreased GAGs content in the damaged joints (1 and 2). These results show that the whole articular cartilage could be affected in a chronic pathological process and is not only a local process occurring in the macroscopically damaged cartilage associated with the loaded area.

The effect of ambient temperature on infrared thermographic images of joints in the distal forelimbs of healthy racehorses

The aim of the study was to describe the dependence on ambient temperature of distal joint temperature at the forelimbs of racehorses. The study also investigated the influence of differing ambient temperatures on the temperature difference between joints: this was measured ipsilaterally (i.e. between the carpal and fetlock joints along each forelimb) and contralaterally (i.e. between the same joints of the left and right forelimbs). Sixty-four healthy racehorses were monitored over 10 months. At each session, three thermographic images were taken of the dorsal, lateral and medial aspects of the distal forelimbs. Temperature measurements were made from regions of interest (ROIs) covering the carpal and fetlock joints. There was a strong correlation between ambient temperature and absolute joint temperature at all ROIs. The study also observed a moderate correlation between ambient temperature and the ipsilateral temperature differences between joints when measured from the medial and lateral aspects. No significant correlation was noted when measured dorsally. The mean contralateral temperature differences between joints were all close to 0°C. The data support previous reports that the temperature distribution between the forelimbs of the healthy equine is generally symmetric, although some horses differ markedly from the average findings.

Effects of xylazine and dexmedetomidine on equine articular chondrocytes in vitro

ObjectiveTo assess the effects of xylazine and dexmedetomidine on equine chondrocytes, in vitro. Study designProspective, experimental study. Study materialEquine articular chondrocytes from five male horses. MethodsChondrocytes were isolated from healthy equine articular cartilage of the metacarpo/metatarsophalangeal joints. Cell viability was assessed using the WST-8 assay by exposing chondrocytes to xylazine (0.5, 1, 2, 4, 8, 16.6, 25, 50 mg mL−1) or dexmedetomidine (0.001, 0.005, 0.01, 0.05, 0.175, 0.25 mg mL−1) for 15, 30 and 60 minutes. Based on the results of these tests, cells were treated with xylazine (1, 4, 25 mg mL−1) or dexmedetomidine (0.05, 0.175, 0.25 mg mL−1) for 15 minutes to further evaluate: cell viability by neutral red uptake; cell membrane integrity by lactate dehydrogenase release and by fluorescence microscopy with Hoechst 33342 and propidium iodide (PI), and apoptosis by flow cytometry using double staining with annexin V-fluorescein isothiocyanate/PI and by cell morphology. ResultsBoth drugs reduced cell viability in a dose-dependent manner. Specifically, all xylazine concentrations, except 0.5 mg mL−1 and 1 mg mL−1, significantly reduced cell viability, whereas the effects of dexmedetomidine were evident only at 0.175 mg mL−1 and 0.25 mg mL−1. The highest concentrations of xylazine (25 mg mL−1) and dexmedetomidine (0.25 mg mL−1) caused loss of membrane integrity. Cell morphology and flow cytometry analyses demonstrated signs of late apoptosis in xylazine-treated cells, and signs of late apoptosis and necrosis in dexmedetomidine-treated cells. Conclusions and clinical relevanceThis study offers new insights into the potential chondrotoxicity induced by dexmedetomidine and xylazine. Therefore, the intra-articular administration of α2-agonists should be conducted with care, especially for doses of ≥ 4 mg mL−1 of xylazine and 0.175 mg mL−1 and 0.25 mg mL−1 of dexmedetomidine.

Solute transport at the interface of cartilage and subchondral bone plate: Effect of micro-architecture

Cross-talk of subchondral bone and articular cartilage could be an important aspect in the etiology of osteoarthritis. Previous research has provided some evidence of transport of small molecules (~370Da) through the calcified cartilage and subchondral bone plate in murine osteoarthritis models. The current study, for the first time, uses a neutral diffusing computed tomography (CT) contrast agent (iodixanol, ~1550Da) to study the permeability of the osteochondral interface in equine and human samples. Sequential CT monitoring of diffusion after injecting a finite amount of contrast agent solution onto the cartilage surface using a micro-CT showed penetration of the contrast molecules across the cartilage-bone interface. Moreover, diffusion through the cartilage-bone interface was affected by thickness and porosity of the subchondral bone as well as the cartilage thickness in both human and equine samples. Our results revealed that porosity of the subchondral plate contributed more strongly to the diffusion across osteochondral interface compared to other morphological parameters in healthy equine samples. However, thickness of the subchondral plate contributed more strongly to the diffusion in slightly osteoarthritic human samples.

A Microbiological Map of the Healthy Equine Gastrointestinal Tract.

Horses are exquisitely sensitive to non-specific gastrointestinal disturbances as well as systemic and extraintestinal conditions related to gut health, yet minimal data are available regarding the composition of the microbiota present in the equine stomach, small intestine, and cecum and their relation to fecal microbiota. Moreover, there is minimal information regarding the concordance of the luminal and mucosal microbial communities throughout the equine gut. Illumina-based 16S rRNA gene amplicon sequencing of the luminal and mucosal microbiota present in seven regions of the gastrointestinal tract of nine healthy adult horses revealed a distinct compositional divide between the small and large intestines. This disparity in composition was more pronounced within the luminal contents, but was also detected within mucosal populations. Moreover, the uniformity of the gut microbiota was much higher in the cecum and colon relative to that in the stomach, jejunum and ileum, despite a significantly higher number of unique sequences detected in the colon. Collectively, the current data suggest that while colonic samples (a proxy for feces) may provide a reasonable profile of the luminal contents of the healthy equine large intestine, they are not informative with regard to the contents of the stomach or small intestine. In contrast to the distinct difference between the highly variable upper gastrointestinal tract microbiota and relatively uniform large bowel microbiota present within the lumen, these data also demonstrate a regional continuity present in mucosal microbial communities throughout the length of the equine gut.

Development of a novel ex vivo equine corneal model.

To develop an ex vivo equine corneal organ culture model. Specifically, to assess the equine cornea's extracellular matrix and cellularity after 7 days using two different culture techniques: either (i) immersion system or (ii) air/liquid interface system, to determine the best ex vivo equine corneal model. Fourteen healthy equine corneas of various breeds. Equine corneas with 2 mm of perilimbal sclera were freshly harvested from 7 horses undergoing humane euthanasia. One corneal-scleral ring (CSR) from each horse was randomly placed in the (i) immersion condition organ culture system (IC), with the contralateral CSR being placed in the (ii) air/liquid interface organ culture system (ALC) for 7 days. All corneas were evaluated using serial daily gross photography, histology, qPCR, and TUNEL assay. corneal-scleral rings placed in the IC (i) had complete loss of corneal transparency on gross photography by 7 days, showed a significant level of corneal stromal disorganization, significantly increased α-SMA levels on qPCR, and apoptosis on TUNEL assay compared to controls. The ALC (ii) had weak stromal disorganization on histopathologic examination and was not significantly different from normal equine corneal controls on all other evaluated parameters. The air-liquid interface organ culture system maintains the equine cornea's extracellular matrix and preserves corneal transparency, while the immersion condition results in near complete degradation of normal equine corneal architecture after 7 days in culture. The air-liquid organ culture is a viable option to maintain a healthy equine cornea in an ex vivo setting for wound healing studies.

Inflammatory response to the administration of mesenchymal stem cells in an equine experimental model: effect of autologous, and single and repeat doses of pooled allogeneic cells in healthy joints.

BackgroundMesenchymal stem cells (MSCs) transplantation has become a promising therapeutic choice for musculoskeletal injuries. Joint-related disorders are highly prevalent in horses. Therefore, these animals are considered as suitable models for testing MSC-based therapies for these diseases. The aim of this study was to investigate the clinical and inflammatory responses to intra-articular single and repeat dose administration of autologous or of pooled allogeneic MSCs in healthy equine healthy joints. Six horses were intra-articularly injected with a single autologous dose of bone marrow derived MSCs (BM-MSCs) and two separate doses of allogeneic BM-MSCs pooled from several donors. All contralateral joints were injected with Lactated Ringer’s Solution (LRS) as the control vehicle. Signs of synovitis and lameness were evaluated at days 0, 1, 2, 3, 5 and 10 after injection. Total protein (TP), white blood cell count (WBC) and neutrophil count (NC) in synovial fluid were also measured at the same time-points.ResultsA mild synovial effusion without associated lameness was observed after all BM-MSCs injections. The second allogeneic injection caused the lowest signs of synovitis. Local temperature slightly increased after all BM-MSCs treatments compared to the controls. TP, WBC and NC in synovial fluids also increased during days 1 to 5 after all BM-MSCs injections. Both, clinical and synovial parameters were progressively normalized and by day 10 post-inoculation appeared indistinguishable from controls.ConclusionsIntra-articular administration of an allogeneic pool of BM-MSCs represents a safe therapeutic strategy to enhance MSCs availability. Importantly, the absence of hypersensitivity response to the second allogeneic BM-MSCs injection validates the use of repeat dose treatments to potentiate the therapeutic benefit of these cells. These results notably contribute to the development of stem cell based therapies for equine and human joint diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-016-0692-x) contains supplementary material, which is available to authorized users.

Synovial fluid pretreatment with hyaluronidase facilitates isolation of CD44+ extracellular vesicles

Extracellular vesicles (EVs) in synovial fluid (SF) are gaining increased recognition as important factors in joint homeostasis, joint regeneration, and as biomarkers of joint disease. A limited number of studies have investigated EVs in SF samples of patients with joint disease, but knowledge on the role of EVs in healthy joints is lacking. In addition, no standardized protocol is available for isolation of EVs from SF. Based on the high viscosity of SF caused by high concentrations of hyaluronic acid (HA) – a prominent extracellular matrix component – it was hypothesized that EV recovery could be optimized by pretreatment with hyaluronidase (HYase). Therefore, the efficiency of EV isolation from healthy equine SF samples was tested by performing sequential ultracentrifugation steps (10,000g, 100,000g and 200,000g) in the presence or absence of HYase. Quantitative EV analysis using high-resolution flow cytometry showed an efficient recovery of EVs after 100,000g ultracentrifugation, with an increased yield of CD44+ EVs when SF samples were pretreated with HYase. Morphological analysis of SF-derived EVs with cryo-transmission-electron microscopy did not indicate damage by high-speed ultracentrifugation and revealed that most EVs are spherical with a diameter of 20–200 nm. Further protein characterization by Western blotting revealed that healthy SF-derived EVs contain CD9, Annexin-1, and CD90/Thy1.1. Taken together, these data suggest that EV isolation protocols for body fluids that contain relatively high amounts of HA, such as SF, could benefit from treatment of the fluid with HYase prior to ultracentrifugation. This method facilitates recovery and detection of CD44+ EVs within the HA-rich extracellular matrix. Furthermore, based on the findings presented here, it is recommended to sediment SF-derived EVs with at least 100,000g for optimal EV recovery.

Central corneal thickness measurements and ultrasonographic study of the growing equine eye.

The aim of this study was to determine the central corneal thickness (CCT) of healthy equine eyes with high-frequency ultrasound (HF) and ultrasound biomicroscopy (UBM) and to measure the axial dimensions with B-mode ultrasound. Additionally, the influence of the age on the measured dimensions of the globe was analyzed. Fifty warm blood horses (mean age 23.0 ± 21.12 months) were divided into 3 age groups (group 1: 0-6 months; group 2: ≥7-30 months; and group 3: ≥31-78 months). Corneal measurements were performed with a 22-MHz and a 50-MHz transducer. Crystalline lens thickness (CLT), vitreous chamber depth (VCD), and axial globe length (AGL) were measured at a frequency of 10 MHz. Anterior chamber depth (ACD) was calculated. The eyes were measured under general anesthesia nonrelated to this study. The mean CCT of 50 eyes was 818 ± 41 μm. The mean values ± SD in mm for axial dimensions were AGL 34.7 ± 2.7, ACD 3.4 ± 0.7, CLT 10.0 ± 0.9, and VCD of 20.4 ± 1.5. A positive correlation with increasing age was found for all dimensions. No significant differences were found between the gender and the left or right eye. HF and UBM are valuable high-resolution imaging tools for CCT measurements in horses. The here-obtained values support previous findings that a positive correlation with increasing age exists for CCT, ACD, CLT, VCD, and AGL in horse.

Autologous processed plasma: cytokine profile and effects upon injection into healthy equine joints.

This experimental controlled study was performed to evaluate the composition of autologous processed plasma (APP), and the effects of APP intra-articular injection into healthy equine metacarpophalangeal joints. The effects on joints were analysed with a short-phase protocol and a prolonged-phase protocol using saline-injected joints as controls. For the short protocol, horses received one intra-articular APP injection. Synovial fluid samples were collected prior to the injection and 3, 6, 24, 48, and 16 h after treatment. For the prolonged protocol, the joints received three weekly injections of APP, and samples were collected at 0, 7, 14, 21, and 28 days before APP administration. IL1-ra level was found to be increased in APP compared to plasma. Upon intra-articular administration of APP, transient (up to 24 h) increases in white blood cell (WBC) counts along with elevated protein and prostaglandin E2 (PGE2) concentrations were observed in the treated joints. Over the 28-day observation period, APP did not elicit changes relative to baseline levels, but WBC counts, PGE2 and chondroitin sulphate concentrations were lower than those found in the control. In conclusion, APP intra-articular injection induced a mild and transitory inflammatory response but no inflammation reaction was observed over a longer period of treatment and observation.

Effect of incubation temperature on the diagnostic sensitivity of the glanders complement fixation test.

The complement fixation test (CFT) is the only serological test prescribed by the World Organisation for Animal Health (OIE) for the diagnosis of glanders in international trading of equids. However, false-positive reactions have caused financial losses to the animal owners in the past, and false-negative tests have resulted in the introduction of glanders into healthy equine populations in previously glanders-free areas. Both warm (incubation at 37°C for 1 h) and cold (overnight incubation at 4°C) procedures are recommended by the OIE for serodiagnosis of glanders. In a comparison of the sensitivity and specificity of the two techniques, using the United States Department of Agriculture antigen, warm CFT was found to be significantly less sensitive (56.8%; p < 0.0005) than the cold CFT (83.6%). Cold CFT thus increases the detection rate of glanders but a lower diagnostic specificity has to be accepted. The immunoblot was used as the gold standard.

Evaluation of ultrasound velocity in enucleated equine aqueous humor, lens and vitreous body.

BackgroundSonographic ophthalmic examinations have become increasingly important in veterinary medicine. If the velocity of ultrasound in ocular tissues is known, the A-mode ultrasound method may be used to determine the axial intraocular distances, such as anterior chamber depth, lens thickness, axial length of the vitreous and axial globe length, which are required for intraocular lens (IOL) power calculations. To the authors' knowledge, the velocity of ultrasound in the ocular tissues of the horse was not previously determined. In the present study, 33 lenses, 29 samples of aqueous and 31 of vitreous from 35 healthy equine eyes have been examined. The corresponding ultrasound velocities are reported in dependence of age, temperature, gender and elapsed time after enucleation.ResultsThe velocity of ultrasound at 36°C in equine aqueous, lens and vitreous are 1529 ±10 m/s, 1654± 29 m/s and 1527 ±16 m/s respectively, and the corresponding conversion factors are 0.998± 0.007, 1.008 ±0.018 and 0.997 ±0.010. A linear increase of the speed of ultrasound with increasing temperature has been determined for aqueous and vitreous. No temperature dependence was found for the speed of ultrasound in the lens. The ultrasound velocity did not significantly differ (95%) on the basis of gender, age or time after enucleation during the first 72 hours after death.ConclusionsCompared to human eyes, the ultrasound velocity in equine lental tissue deviates by one percent. Therefore, axial length measurements obtained with ultrasound velocities for the human eye must be corrected using conversion factors. For the aqueous and vitreous, deviations are below one percent and can be neglected in clinical settings.

Histological study of the external, middle and inner ear of horses.

Clinical, anatomical and histological aspects of the equine acoustic organ have been poorly investigated and illustrated in literature so far. It is understood that an intact acoustic organ and hearing function are of vital importance for the well-being of flight animals like horses. The knowledge of the acoustic organ is usually transferred analogously from other mammals to horses. The purpose of this study was to provide a detailed and complete histological description of the healthy equine auditory organ, and to determine its congruity to other mammalians. Anatomical dissections and histological preparations were carried out on ten cadaver heads. Specimens of various parts of the equine acoustic organ were taken and evaluated histologically. The histological composition of external, middle and inner ear structures are predominantly congruent to those of other mammals, especially to human beings. Unique inwardly directed rete pegs within the osseous ear canal and the prominent tensor tympani muscle are described for the first time. Results obtained in this study can be employed as references for further research on the equine acoustic organ and improve the understanding of the clinical development of hearing loss, otitis externa/media/interna or tympanosclerosis.

Therapeutic potential of Pirfenidone for treating equine corneal scarring.

To evaluate the safety and efficacy of Pirfenidone (PFD) in the treatment of equine corneal fibrosis using an in vitro model. Healthy donor equine corneas were collected and used to generate primary equine corneal fibroblasts (ECFs) by growing cultures in minimal essential medium supplemented with 10% fetal bovine serum. Equine corneal myofibroblasts (ECMs), used as a model of equine corneal fibrosis, were produced by growing ECF cultures in serum-free medium containing transforming growth factor β1 (1 ng/mL). Trypan blue viability assays and changes in ECF morphology were utilized to determine the optimal PFD dose for this in vitro model. Trypan blue viability, phase-contrast microscopy, and TUNEL assays were used to evaluate the cytotoxicity of PFD. Scratch and MTT assays were used to evaluate the effect of PFD on cellular migration and proliferation. Real-time PCR, immunoblot analysis, and immunocytochemistry were employed to determine the efficacy of PFD to inhibit ECM formation in vitro. Topical PFD application at 200 μg/mL successfully decreased αSMA expression when compared to the TGFβ1 only treatment group (P < 0.01). PFD application ≤ 200 μg/mL did not affect ECF phenotype or cellular viability and did not result in significant cytotoxicity. Pirfenidone safely and effectively inhibits TGFβ1-induced equine corneal fibrosis in vitro. In vivo studies are warranted.

Manual corneal thickness measurements of healthy equine eyes using a portable spectral‐domain optical coherence tomography device

Corneal thickness measurements of the equine globe using spectral-domain optical coherence tomography (SD-OCT) have not been reported. To determine corneal thickness measurements and the intra- and interoperator reliability of a portable SD-OCT device in equine eyes. Prospective observational study. Horses free of ocular disease were used for this study. Gentle manual restraint, in combination with detomidine hydrochloride and a head stand, were employed to ensure proper animal positioning. Corneal pachymetry measurements were obtained from both eyes of each animal 3 times by 2 operators in succession. A 6 mm corneal pachymetry protocol was performed using a portable SD-OCT device. All measurements were obtained manually by one operator (C.G.P.) using the integrated calliper function. Measurements included epithelial thickness, stromal thickness, Descemet's membrane thickness and total corneal thickness. All recorded measurements were analysed to determine both intra- and interoperator reliability. Thirty horses with a mean age of 10.6 ± 6.4 years were examined. Mean epithelial, stromal, Descemet's membrane and total corneal thickness values obtained were, respectively, 174.7 ± 13.6, 599.2 ± 45.4, 38.4 ± 15.3 and 812.0 ± 44.1 μm for operator A and 175.9 ± 12.9, 599.2 ± 44.9, 38.4 ± 15.0 and 812.9 ± 42.9 μm for operator B. A positive correlation was found between Descemet's membrane thickness and age, whereby Descemet's membrane thickness increased by 2 μm/year (P<0.0001). The coefficients of variation for both operators were <4% for all measurements. Intraclass correlations ranged from 0.92 to 0.98. Manual corneal thickness measurement using a portable SD-OCT device provides epithelial, stromal, Descemet's membrane and total corneal thickness measurements with clinically acceptable intra- and interoperator reliability in healthy equine eyes.

The serological evidence in humans supports a negligible risk of zoonotic infection from porcine circovirus type 2

There are two porcine circovirus (PCV) genotypes, PCV-1 and PCV-2. In pigs, PCV-1 infection is asymptomatic but PCV-2 infection can cause severe respiratory disease and other pathology. Although humans ingest PCV-contaminated foods and are exposed to PCV through other sources, the potential of PCV-2 as a zoonotic agent in humans and other species has not been fully explored. Here, four recombinant proteins derived from the PCV-2 capsid gene were examined as antigens using the Luciferase Immunoprecipitation System (LIPS) assay for serological analysis of PCV-2 infection. PCV-2-CAP-Δ1 was the optimum recombinant protein in the LIPS assay with a sensitivity of 93% and specificity of 100% using porcine samples. Testing of healthy human blood donors, equine and bovine serum samples failed to demonstrate the presence of anti-PCV-2 antibodies. Additionally, analysis of two high-risk human groups, cystic fibrosis patients taking porcine derived oral supplements and type I diabetes patients who had undergone porcine islet cell transplantation, showed no evidence of anti-PCV-2 antibodies. These results extend the extensively demonstrated use of LIPS as a robust approach for identifying humoral responses and provide evidence that PCV-2 is likely not infectious in humans.