Disseminated idiopathic lipid keratopathy in a normolipemic cat.

OBJECTIVE: To report a case of idiopathic lipid keratopathy in a normolipemic cat. ANIMAL STUDIED: A 10-year-old neutered female European domestic cat. RESULTS: A cat was evaluated for bilateral white corneal deposits. Slit-lamp examination revealed multiple, well-defined, round, stromal, cream-colored deposits of different sizes associated with generalized superficial corneal vascularization. Blood lipids were normal, and no history of travel to tropical locations or ocular trauma was present. Topical betamethasone/gentamicin 0.1% suspension q 12 hours did not result in any improvement of clinical appearance after one week. Tomography following the initial therapy revealed dense, hyperreflective deposits with posterior shadowing in the anterior and mid stroma of both corneas. A four-week course of itraconazole 0.01% ophthalmic cream was prescribed q 12 hours with no improvement. Four months after the initial examination, a diagnostic superficial keratectomy and amniotic membrane implantation were performed. Histopathological analysis showed membrane bound vacuoles with infiltration of foamy macrophages suggesting a diagnosis of primary lipidosis. The post-surgical period was unremarkable, and ten days later, the patient was re-examined. Opacification from a corneal leukoma was observed in the excision site with mild fibrotic tissue. Two months post-keratectomy, no further changes were detected in the cornea, and the patient was managed only with topical lubricant. CONCLUSIONS: To our knowledge, this is the first report of idiopathic corneal lipidosis in a cat and may be considered as a differential diagnosis of corneal disease in felines.

Preformulation Studies of a Liposomal Formulation Containing Sirolimus for the Treatment of Dry Eye Disease.

PURPOSE: The aim of this study was to develop and characterize a liposomal product containing sirolimus to be administered subconjunctivally for the treatment of nonresponsive keratoconjunctivitis sicca (KCS) or dry eye. METHODS: Formulations were prepared using an ethanol injection method and an adaptation of the heating method in pursuance of the most suitable methodology for future industrial production. Liposomes were loaded with either a high dose of 1 mg/mL of sirolimus or a less toxic dose of 0.4 mg/mL. The effects of critical process and formulation parameters were investigated. Liposomes were characterized in terms of size, zeta potential, polydispersity, differential scanning calorimetry, morphology, entrapment efficiency, phospholipid content, thermal stability, and sterility. The formulation was evaluated clinically in dogs with spontaneous KCS. RESULTS: Sterile liposomal dispersions with sizes ranging from 140 to 211 nm, were successfully obtained. High entrapment efficiency of 93%-98% was achieved. The heating method allowed an easier production of liposomes with high entrapment efficiency, to significantly shorten production time and the elimination of the use of alcohol. The poor stability of the obtained liposomes in aqueous dispersion made the inclusion of a lyophilization step necessary to the manufacturing process. In vivo testing of the liposomal sirolimus formulations in the spontaneous KCS dog model have produced promising results, particularly with a sirolimus dose of 1 mg/mL, indicating the need for further development and study of proposed formulations in the treatment of canine KCS. Clinical improvement in tear production in dogs with spontaneous KCS treated with the 1 mg/mL dose product was observed. CONCLUSIONS: The heating method allowed easier production of high entrapment efficiency liposomes to significantly shorten production time and the elimination of the use of alcohol. Tear production was increased in dogs administered with the formulation.

Proteomic analysis of equine amniotic membrane: characterization of proteins.

INTRODUCTION: Human amniotic membrane (AM) has been used as a biomaterial for surgical wound skin and ocular surface reconstruction for several years. Currently, equine AM has been used for corneal reconstruction in several animal species, and appears to have the same properties as human AM. Despite the observed positive healing abilities of this tissue in horses with ulcerative keratitis the proteins of equine AM have not been described. OBJECTIVE: To identify proteins known to be associated with corneal healing from frozen equine AM. PROCEDURES: Placentas were acquired from healthy live foal births from a local Thoroughbred breeding farm. The amnion was removed from the chorion by blunt dissection, washed with phosphate-buffered saline (PBS), and treated with 0.05% trypsin and 0.02% ethylene diaminetetraacetic acid in PBS. Amnion was attached to nitrocellulose paper (epithelial side up), and cut into 4 × 4 cm pieces. The sheets were frozen at -80 °C. The protein samples were solubilized, and analyzed by 2D gel electrophoresis and shotgun proteomics. RESULTS: A reference identification map of the equine AM proteins was produced and 149 different proteins were identified. From gel-based proteomics, 49 spots were excised and 43 proteins identified by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Shotgun proteomics identified 116 proteins with an overlap of 10 proteins in both analyses. CONCLUSIONS: We have described a reference map for equine AM proteins that may provide a background to explain the positive results found in horses with ulcerative keratopathies using this biomaterial.

Optimal management of equine keratomycosis.

Keratomycosis in the horse exists in several unique clinical forms. This paper discusses the diagnosis and clinical management of keratomycosis in the horse associated with tear film instability, epithelial keratopathy, subepithelial infiltrates, superficial and deep ulcers, plaques, melting ulcers, descemetoceles, iris prolapse, and stromal abscesses. Prompt diagnosis and aggressive treatment of equine keratomycosis can make a major difference in the maintenance of a cosmetic and visual eye.

Metastatic intraocular chondrosarcoma in a dog.

Chondrosarcoma accounts for about 10% of all canine bone tumors and is the second most common primary bone tumor in dogs. In veterinary medicine, chondrosarcomas are classified as skeletal and extraskeletal. Extraskeletal chondrosarcomas are mesenchymal neoplasms of soft tissues and visceral organs that produce neoplastic chondrocytes in a fibrillary matrix. There is no involvement of bone or periosteal tissues in extraskeletal chondrosarcomas. The aim of this report is to describe the first case of a metastatic intraocular extraskeletal chondrosarcoma in a dog.

Effects of topical 0.2 percent Cyclosporine A on corneal neovascularization induced by xenologous amniotic membrane implantation into a corneal stroma micropocket of rats / Efeitos do uso tópico da Ciclosporina A (CsA) 0,2 por cento na neovascularização corneal induzida pelo implante de membrana amniótica xenógena em microbolsa no estroma da córnea de ratos

The objective of the study was to evaluate the topical effects of 0.2 percent Cyclosporine A (CsA) on corneal neovascularization of rats following surgical implantation of equine amniotic membrane into a corneal stroma micropocket. The implantation of xenologous amniotic membrane was performed bilaterally in 90 rats. In the same day of the surgery each right eye started receiving topical CsA twice a day. The left eye received no medication and served as a control. The evaluation of corneal neovascularization was performed by computerized image analysis and histopathological evaluation at 1, 3, 7, 15, 30 and 60 days postoperatively. For the image analysis 10 animals were used per time period, and for the histopathological examination, five animals were used per time period. Image analysis found that corneal neovascularization began on the 3rd postoperative day, reached its peak on the 7th day, and then progressively and rapidly decreased. Statistic analysis indicated that neovascularization of the CsA treated eye on the 7th day was significantly higher than that observed in untreated eyes. On the 30th day, however, this pattern was reversed with the neovascularization observed in the CsA treated eyes declining to the low levels observed on the 3rd day. The degree of neovascularization in the untreated eyes on the 30th day declined to the baseline levels found on day 3 at the 60th day. Histopathological analysis indicated that deposition of collagen in the implanted tissue was completed by the 15th day. Therefore, we concluded that (1) equine amniotic membrane in rat corneal stroma produced an intense neovascularization until the 15th day postoperatively and then regressed, (2) deposition of collagen of the implanted tissue was completed on the 15th day postoperatively, and (3) use of CsA was associated with increase in the corneal neovascularization initially, followed by a quick and intense regression.

Este estudo teve como objetivo a avaliação dos efeitos tópicos da Ciclosporina a 0,2 por cento (CsA) sobre a neovascularização corneana de ratos após implante cirúrgico de membrana amniótica eqüina em microbolsa do estroma corneana. O implante da membrana foi feito bilateralmente em 90 ratos. O tratamento com CsA iniciou-se no mesmo dia da cirurgia, nos olhos direitos dos animais, duas vezes ao dia. Os olhos esquerdos não receberam nenhum tratamento e serviram de controle. A avaliação da neovascularização corneana foi feita por análise de imagem computadorizada e por exame histopatológico aos dias 1, 3, 7, 15, 30 e 60 de pós-cirúrgico. Para a análise de imagem foram utilizados 10 animais por período, e para o exame histopatológico, 5 por período. A análise de imagem demonstrou que a neovascularização iniciou-se no 3º dia pós-cirúrgico, alcançou seu pico no 7º dia e então regrediu rápida e progressivamente até o 60º dia. A análise estatística indicou que a neovascularização no 7º dia nos olhos tratados com CsA foi significantemente mais acentuada do que aquela observada nos olhos não tratados. Entretanto, no 30º dia este fato se reverteu, e a neovascularização observada nos olhos tratados com CsA diminuíra a níveis baixos comparáveis àquela do 3º dia. Já nos olhos não tratados, o grau de neovascularização somente pôde ser comparado àquele nível básico encontrado no 3º dia aos 60 dias de pós-operatório. A análise histopatológica demonstrou que a deposição de colágeno no tecido implantado se completou no 15º dia. Desta maneira, foi possível concluir que (1) a membrana amniótica em estroma corneano de ratos produz intensa neovascularização até o 15º dia de pós-operatório com posterior regressão, (2) a deposição de colágeno do tecido implantado foi completa ao 15º dia de pós-operatório, e que (3) o uso de CsA esteve associado com aumento inicial da neovascularização corneana, seguido de rápida e intensa regressão.

Measurement of retinal nerve fiber layer thickness in normal and glaucomatous Cocker Spaniels by scanning laser polarimetry.

OBJECTIVE: To measure changes in the thickness of the retinal nerve fiber layer in normal and early glaucomatous dogs with scanning laser polarimetry. ANIMALS STUDIED: A total of 45 eyes, 32 normal and 13 glaucomatous eyes, of American Cocker Spaniels with primary glaucoma were used. All eyes were evaluated through a complete neuro-ophthalmic examination, tonometry, gonioscopy, slit-lamp biomicroscopy, and indirect ophthalmoscopy prior to enucleation. METHODS: The retinal nerve fiber layer thickness was measured in anesthetized animals with scanning laser polarimetry (Nerve fiber analyzer, GDx; Laser Diagnostic Technologies, LTD, San Diego, CA, USA). Glaucomatous eyes retained some vision at the time of this study. RESULTS: The mean +/- SD of the retinal nerve fiber layer thickness was 141.69 +/- 18 microm for normal dogs and 105.08 +/- 23.86 microm for visual glaucomatous dogs. The average retinal nerve fiber layer thickness in the superior and inferior retinal quadrants was 148.03 +/- 8.5 and 141.06 +/- 8.73 microm, respectively, for normal dogs, and 106.61 +/- 25.77 and 107.08 +/- 24.99 microm in the superior and inferior retinal quadrants, respectively, for glaucomatous dogs. The superior to nasal retinal nerve fiber layer thickness ratio was 1.45 for normal dogs and 1.26 for visual glaucomatous dogs. CONCLUSIONS: Using scanning laser polarimetry it was possible to detect changes in retinal nerve fiber layer thickness in glaucomatous dogs at early stages of the disease. Therefore, this instrument has the potential to improve the clinical management of canine glaucoma by detecting progressive changes to the retinal nerve fiber layer.

Ascorbic acid levels of aqueous humor of dogs after experimental phacoemulsification.

Phacoemulsification has been successfully employed in humans and animals for lens extraction. This ultrasonic extracapsular surgical technique induces hydroxyl radical formation in the anterior chamber, which accumulates despite irrigation and aspiration. In this paper we determined the total antioxidant status of aqueous humor after phacoemulsification by measuring aqueous humor ascorbic acid levels. Mixed-breed dogs (n = 11; weighing about 10 kg) with normal eyes as determined by slit-lamp biomicroscopy, applanation tonometry, and indirect ophthalmoscopy had phacoemulsification performed in one eye with the other eye used as a control. Samples of aqueous humor were obtained by anterior chamber paracentesis before surgery and at days 1, 2, 3, 7, and 15 after surgery. Total aqueous humor antioxidant status was inferred from the capacity of aqueous humor to inhibit free radical generation by 2,2-azobis (2-amidopropane) chlorine. Ascorbic acid concentrations were measured by high-pressure liquid chromatography with UV detection. Protein content was determined with the biuret reagent. Statistical analysis was performed by anova followed by the paired t-test. Total antioxidant capacity was reduced from 48 to 27 min during the first 24 h with a gradual increase thereafter, remaining statistically lower than the control eye until 7 days postoperatively. Reduced levels of ascorbic acid followed this reduction in antioxidant capacity (from 211 to 99 microm after 24 h), remaining lower than the control eye until 15 days postoperatively. Protein concentration in aqueous humor increased from 0.62 mg/mL to 30.8 mg/mL 24 h after surgery, remaining statistically lower than the control eye until 15 days postoperatively. Paracentesis alone did not significantly alter the parameters measured. These results indicate that after phacoemulsification, the aqueous humor ascorbic acid levels and antioxidant defenses in aqueous humor are reduced, indirectly corroborating free radical production in the anterior chamber as a result of phacoemulsification. The inflammatory process consequent to the surgical procedure demonstrated by increased protein content in aqueous humor can also contribute to free radical production and ascorbic acid consumption.

Amniotic membrane transplantation for the reconstruction of the ocular surface in three cases.

This paper describes the use of amniotic membrane in the reconstructive surgical repair of generalized keratomalacia, ankyloblepharon, and after fibrous histiocytoma removal in two dogs and a cat. Case 1 was an 11-year-old female Yorkshire terrier with severe bullous keratomalacia in the left eye (OS). A frozen canine amniotic membrane graft and a third eyelid flap were applied. At day 80 postoperatively, only a mild scar and corneal vascularization were present. Case 2 was a 4-year-old female Siamese cat with symblepharon of both eyes after rhinotracheitis. Resection of the conjunctiva and frozen canine amniotic membrane transplant were performed. One month later, there was a little corneal scarring and corneal vascularization. Case 3 was a 6-year-old female terrier with a scleral and corneal mass at the 11-12 o'clock position (OS). Resection of the mass and amniotic membrane transplantation were performed. The mass was a fibrous histiocytoma localized to the sclera and cornea. This eye healed with mild conjunctivalization and no pigmentation. Amniotic membrane transplantation can be used as a method of reconstruction of the ocular surface with good repair of the cornea and minimal scarring in small animals.

Morphometric analysis of the corneal endothelium of Yacare caiman (Caiman yacare) using scanning electron microscopy.

The objective of this study was to examine the endothelial surface morphology and to perform morphometric analysis of the corneal endothelial cells of Yacare caiman (Caiman yacare) using scanning electron microscopy. Morphometric analysis with regard to polygonality, mean cell area, cell density and coefficient of variation of mean cell area was performed. Cell areas were measured using image analysis software. The normal corneal endothelium of Yacare caiman consisted of polygonal cells of uniform size and shape with interdigitations of the cell borders. Microvilli appeared as protrusions on the cellular surface. The average cell area was 270 +/- 24 microm(2) and the endothelial cell density was 3704 +/- 324 cells/mm(2). The coefficient of variation of cell area was 0.22. This study demonstrates that the Yacare caiman corneal endothelium is similar to those described in other vertebrates.

Fungal flora of normal eyes of healthy horses from the State of Rio de Janeiro, Brazil.

The conjunctival fungal flora of 32 adult horses with normal eyes (n = 64) from the State of Rio de Janeiro in Brazil was identified in the fall of 2000 using horses of different breeds, both genders and aged 5-19 years old. The culture samples were taken from the conjunctival sac of both eyes with a sterile cotton swab wetted with saline solution, seeded in Sabouraud's dextrose agar with chloramphenicol, and incubated for 5 days at an average temperature of 25 degrees C. The number of fungal colonies per eye varied between 0 and 250 colony forming units (CFUs). There were often differences in colony types between eyes of the same animal. Filamentous fungi of genera were isolated and identified in the following proportion of the total genera of fungal colonies isolated: Aspergillus (32.2%), Penicillium (25.8%), Scopulariopsis (15.9%), Trichoderma (11.2%), Cladosporium (5.6%), Mucor (2.1%), Syncephalastrum (2.1%), Eurotium (1.7%), Geotrichum (0.9%), Rhizopus (0.9%), Gliomastix (0.4%), Fusarium (0.4%), Staphylotrichum (0.4%) and Verticillium (0.4%). Yeast genera represented 9% of the total isolates. Over half the horses had at least one normal eye with either Aspergillus, Penicillium, Trichoderma or Scopulariopsis isolated, which is a departure from other studies of the normal horse eye.