Sheep corneal endothelium morphology - evaluation with trypan blue and alizarin red

Background: The endothelium is a layer fundamental to maintaining corneal transparency. In ophthalmology, sheep eyes have been used as a model in research related to corneal transplantation. Different techniques have been used to evaluate the corneal endothelium. Concerning vital dyes, corneal endothelial cell analyses have not yet been studied in ovines. The purpose of the present study was to evaluate the morphology of endothelial cells from different regions of the cornea of sheep after staining with alizarin red and trypan blue using an optical microscope. Materials, Methods & Results: Twenty healthy eyes of 10 male sheep obtained from a licensed commercial slaughterhouse were studied. The study was approved by the Research Committee of the Faculty of Veterinary at UFRGS and followed the ethical standards of the Association for Research in Vision and Ophthalmology (ARVO). Immediately after the slaughter, the eyes were enucleated and underwent eye examination. The corneal endothelium was stained with trypan blue and alizarin red and examined and photographed using an optical microscope. The central, superior, inferior, nasal and temporal areas of the cornea were evaluated for cell morphology. Data were compared by t-tests. Differences were considered statistically significant at P < 0.05. Immediately after staining the corneal endothelium, it was possible to examine with an optical microscope, obtain images and analyse the shape of endothelial cells from all regions of the sheep cornea. Polygonal, uniform and continuous cells were observed in all samples studied. Considering all the corneas analysed, cells with 6 sides (75.11%), 5 sides (12.76%) and 4 sides (12.12%) were found. In the central region of the cornea 75.91% of cells with 6 sides, 12.6% of cells with 5 sides and 11.48% with 7 sides were found. In the superior region of the cornea 76.07% of cells with 6 sides, 13.25% with 5 sides and 10.68% with 7 sides were found. In the lower region were found 74.72% of cells with 6 sides, 13% with 5 sides and 12.27% with 7 sides. In the temporal region, 74.14% were 6-sided cells, 11.42% had 5 sides, and 14.43% had 7 sides. Furthermore, in the nasal region, 74.72% of the cells had 6 sides, 13.54% had 5 sides, and 11.73% had 7 sides. No significant differences were found between cell morphology in all corneal regions evaluated. In addition, no significant difference was found when comparing the right eye with the left eye. Discussion: Different methods are used for the analysis of corneal endothelium. For ex vivo research optical microscopy after endothelial staining is an alternative low-cost technique that allows the analysis of all regions of the cornea. Quantitative analyses must characterise the endothelial parameters of the different species. The analysis of the morphology of corneal endothelium with an optic microscope after staining with alizarin red has been described as an effective, rapid and cost-efficient method, since this dye blends with the borated cells, allowing identification. In the present study, using optical microscopy and coloration with alizarin red it was possible to explore and obtain images of the ovine endothelium of all regions of the cornea. In the current study, the endothelium had a predominance of cells will 6 sides in all regions studied. This study allowed us to obtain images of the endothelium as well as quantitative data on the morphology of the different regions of the sheep cornea. This study demonstrated that morphology did not differ between the central and peripheral regions. The findings of this study represent a further source of reproducible data that should be considered when using sheep cornea as ex vivo model for experimental research.

Acute effects of ropivacaine hydrochloride on corneal endothelial cell ultrastructure of horses: ex vivo study / Efeitos agudos do cloridrato de ropivacaína na ultraestrutura das células endoteliais da córnea de equinos: estudo ex vivo

The objective of this study was to evaluate the acute effects of ropivacaine hydrochloride on the corneal endothelium of horses. Forty-eight eyes were obtained from a commercial slaughterhouse and were randomly divided into three groups. In group A, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 60 seconds. In group B, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 15 minutes. In group C, the corneal endothelium was exposed to a balanced saline solution for 60 seconds. Afterwards, all samples were prepared for evaluation with scanning electron microscopy. Random electromicrographs were obtained from each sample. The images were analysed and, with the aid of software, areas with no endothelial cells were measured. The average endothelial loss, expressed as a percentage in relation to the total area, of the samples in group A was 5.28%. The average endothelial loss of samples from group B, expressed as a percentage in relation to the total area, was 20.39%. The damage to the corneal endothelium was significantly greater in group B compared to groups A and C. It was possible to conclude that 0.75% ropivacaine hydrochloride induced acute damage to corneal endothelium cells.

Objetivou-se avaliar os efeitos agudos do cloridrato de ropivacaína no endotélio da córnea de equinos. Quarenta e oito olhos de equinos foram divididos aleatoriamente em três grupos. No grupo A o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 60 segundos. No grupo B o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 15 minutos. No grupo C o endotélio da córnea foi exposto à solução salina balanceada por 60 segundos. As amostras foram preparadas para avaliação com microscopia eletrônica de varredura. Eletromicrografias eletrônicas de varredura foram obtidas aleatoriamente de cada amostra. As imagens foram analisadas e, com o auxílio de um programa para morfometria foram medidas as áreas sem células endoteliais. A perda endotelial média foi expressa em porcentagem em relação à área total das amostras do grupo A foi de 5,28%. A perda endotelial média de amostras do grupo B foi expressa em porcentagem em relação à área total, foi de 20,39%. O dano ao endotélio da córnea foi significativamente maior no grupo B, comparado aos grupos A e C. O cloridrato de ropivacaína a 0,75% induziu dano agudo nas células do endotélio da córnea de equinos.

Effects of daily curve and age on intraocular pressure in Criollo horses / Efeitos da curva diária e da idade na pressão intraocular em cavalos Crioulos

The objective of this study was to evaluate the intraocular pressure (IOP) of healthy Criollo horses using a rebound tonometer throughout the day. In addition, assessments were made in horses of different ages. Twenty-seven horses, male and female, were divided into three groups by age: Group I (3-5 years old), Group II (6-8 years old), and Group III: (9-16 years old). Ophthalmic examinations were performed using the Schirmer tear test, slit-lamp biomicroscopy, fluorescein test and indirect ophthalmoscopy. Seven measurements of IOP were assessed on the same day (at 6:00 am, 9:00 am, 12:00 am, 3:00 pm, 6:00 pm, 9:00 pm and 00:00 pm). A t-test was used when there were two groups of comparisons and ANOVA was used to detect differences in IOP between measurement times and between age categories. The average IOP was 28.4 ± 3.7 mmHg for all eyes. The mean IOP for Groups I, II and III were 29.2 ± 3.5, 28.4 ± 4.3 and 27.7 ± 3.2 mmHg, respectively. There was no statistically difference between right and left eyes. There was a significant difference between Group I and Group III (P = 0.008). There were no statistically significant differences between measurements recorded at different times of the day (P = 0.560). The IOP was not influenced by the circadian rhythm, but older horses showed reduced IOP.

O objetivo deste estudo foi avaliar a pressão intraocular (PIO) com um tonômetro rebote em cavalos da raça Crioula ao longo do dia. Além disso, foram realizadas avaliações em cavalos de diferentes idades. Vinte e sete cavalos, machos ou fêmeas foram divididos em três grupos de acordo com a idade: Grupo I (3 a 5 anos), Grupo II (6 a 8 anos) e Grupo III (9 a 16 anos). O exame oftálmico incluiu teste da lágrima de Schirmer, biomicroscopia com lâmpada de fenda, prova da fluoresceína e oftalmoscopia indireta. Sete medidas de PIO foram realizadas no mesmo dia (às 6h, 9h, 12h, 15h, 18h, 21h e 24h). O teste t foi usado quando havia dois grupos de comparações e a análise de variância (ANOVA) foi usada para detectar diferenças na PIO entre os momentos de medição e entre as categorias de idade. A PIO média foi de 28,4 ± 3,7 mmHg para todos os olhos. A PIO média dos grupos I, II e III foi de 29,2 ± 3,5, 28,4 ± 4,3 e 27,7 ± 3,2 mmHg, respectivamente. Não houve diferença estatística entre os olhos direito e esquerdo. Houve diferença entre o grupo I e o grupo III (P = 0,008). Não houve diferença entre as medidas registradas em diferentes momentos do dia (P = 0,560). A PIO aferida com tonômetro de rebote em cavalos da raça Crioula saudáveis não sofreu influência da hora do dia em que foi realizada, mas foi menor em equinos mais velhos.

Modified eye evisceration in a tropical screech owl (Megascops choliba)

Background: An adult owl was presented with an injury to the right eye that rendered it blind in that eye. The left eye was normal. Removal of the right eye was recommended and a modified eye evisceration was performed. No complications were observed during or after surgery. The objective of this paper is to describe the modified eye evisceration technique that was successfully used in a tropical screech owl (Megascops choliba). Case: An adult owl was presented with an injury to the right eye that rendered it blind in that eye. Two previous surgical treatments have been carried out but have not been successful. Using a portable slit-lamp (Kowa SL-15®), both eyes were examined. The left eye was normal. Upon ophthalmic examination of the right eye, the owl demonstrated blepharospasm and large central corneal ulcer. Removal of the right eye was recommended. The bird received midazolam hydrochloride [Dormire® - 1 mg/kg, IM] and ketamine hydrochloride [Ketamina® - 5 mg/kg IM] as pre-anesthetic medications. Subsequently, the bird was anesthetized with isoflurane (Isoforine®) by facemask for induction, and then maintained with isoflurane vaporized in 100% oxygen through an endotracheal tube. With the aid of a surgical microscope and microsurgery materials, a modified eye evisceration was performed. Post-operatively, the owl received meloxicam [Maxicam® - 0.5 mg/kg, IM] and tramadol hydrochloride [Cronidor® - 15 mg/kg, orally for 4 days]. The day after surgery, the owl was comfortable and its usual appetite was regained. The patient remained hospitalized for 3 weeks and was evaluated daily. The skin sutures were removed 10 days after the surgical procedure and the surgical wound had healed normally. The patient was reintroduced into the wild after 2 months. During the 6 months post-release, the bird was evaluated once a month, and no complications were observed. Discussion: Severe eye trauma and complicated corneal ulcers are common causes of eyeball removal in birds. In birds, there is a high risk of complications during enucleation. The fragility of the orbital bones makes them susceptible to trauma during the surgery. Evisceration involves the removal of the inner contents of the eye while leaving the cornea and the sclera intact. In the current case, evisceration was chosen because the eye was blind, and maintaining a blind eye would be a source of pain and infection. In the modified evisceration technique, the risk of complications is minimal compared to enucleation, mainly because surgical manipulation is minimal. In our case, the total surgery time was 20 min. Another complication reported after enucleation in birds is the possibility of disfiguring the bird because the removal of the globe disturbs the natural head balance. To avoid these complications, the use of an intraocular prosthesis after evisceration in birds has been performed. However, owls have a tubular-shaped globe with scleral ossicles. These factors could hinder or even prevent the accommodation of a cylindrical silicone prosthesis. In the present case, an intraocular prosthesis implant was never considered due to the unavailability of the prosthesis and to avoid the risk of postoperative complications that have been reported from the literature in dogs. In this case, the owl recovered well from anesthesia without complications, and no postoperative hemorrhage was observed. No signs of pain were observed during the postoperative period and the owl had already shown an appetite and fed on the first postoperative day. The previously published reports using the modified evisceration technique also demonstrated an absence of pain signs during the postoperative period.(AU)

Acute effects of ropivacaine hydrochloride on corneal endothelial cell ultrastructure of horses: ex vivo study / Efeitos agudos do cloridrato de ropivacaína na ultraestrutura das células endoteliais da córnea de equinos: estudo ex vivo

ABSTRACT: The objective of this study was to evaluate the acute effects of ropivacaine hydrochloride on the corneal endothelium of horses. Forty-eight eyes were obtained from a commercial slaughterhouse and were randomly divided into three groups. In group A, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 60 seconds. In group B, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 15 minutes. In group C, the corneal endothelium was exposed to a balanced saline solution for 60 seconds. Afterwards, all samples were prepared for evaluation with scanning electron microscopy. Random electromicrographs were obtained from each sample. The images were analysed and, with the aid of software, areas with no endothelial cells were measured. The average endothelial loss, expressed as a percentage in relation to the total area, of the samples in group A was 5.28%. The average endothelial loss of samples from group B, expressed as a percentage in relation to the total area, was 20.39%. The damage to the corneal endothelium was significantly greater in group B compared to groups A and C. It was possible to conclude that 0.75% ropivacaine hydrochloride induced acute damage to corneal endothelium cells.

RESUMO: Objetivou-se avaliar os efeitos agudos do cloridrato de ropivacaína no endotélio da córnea de equinos. Quarenta e oito olhos de equinos foram divididos aleatoriamente em três grupos. No grupo A o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 60 segundos. No grupo B o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 15 minutos. No grupo C o endotélio da córnea foi exposto à solução salina balanceada por 60 segundos. As amostras foram preparadas para avaliação com microscopia eletrônica de varredura. Eletromicrografias eletrônicas de varredura foram obtidas aleatoriamente de cada amostra. As imagens foram analisadas e, com o auxílio de um programa para morfometria foram medidas as áreas sem células endoteliais. A perda endotelial média foi expressa em porcentagem em relação à área total das amostras do grupo A foi de 5,28%. A perda endotelial média de amostras do grupo B foi expressa em porcentagem em relação à área total, foi de 20,39%. O dano ao endotélio da córnea foi significativamente maior no grupo B, comparado aos grupos A e C. O cloridrato de ropivacaína a 0,75% induziu dano agudo nas células do endotélio da córnea de equinos.

Effects of daily curve and age on intraocular pressure in Criollo horses / Efeitos da curva diária e da idade na pressão intraocular em cavalos Crioulos

The objective of this study was to evaluate the intraocular pressure (IOP) of healthy Criollo horses using a rebound tonometer throughout the day. In addition, assessments were made in horses of different ages. Twenty-seven horses, male and female, were divided into three groups by age: Group I (3-5 years old), Group II (6-8 years old), and Group III: (9-16 years old). Ophthalmic examinations were performed using the Schirmer tear test, slit-lamp biomicroscopy, fluorescein test and indirect ophthalmoscopy. Seven measurements of IOP were assessed on the same day (at 6:00 am, 9:00 am, 12:00 am, 3:00 pm, 6:00 pm, 9:00 pm and 00:00 pm). A t-test was used when there were two groups of comparisons and ANOVA was used to detect differences in IOP between measurement times and between age categories. The average IOP was 28.4 ± 3.7 mmHg for all eyes. The mean IOP for Groups I, II and III were 29.2 ± 3.5, 28.4 ± 4.3 and 27.7 ± 3.2 mmHg, respectively. There was no statistically difference between right and left eyes. There was a significant difference between Group I and Group III (P = 0.008). There were no statistically significant differences between measurements recorded at different times of the day (P = 0.560). The IOP was not influenced by the circadian rhythm, but older horses showed reduced IOP.(AU)

O objetivo deste estudo foi avaliar a pressão intraocular (PIO) com um tonômetro rebote em cavalos da raça Crioula ao longo do dia. Além disso, foram realizadas avaliações em cavalos de diferentes idades. Vinte e sete cavalos, machos ou fêmeas foram divididos em três grupos de acordo com a idade: Grupo I (3 a 5 anos), Grupo II (6 a 8 anos) e Grupo III (9 a 16 anos). O exame oftálmico incluiu teste da lágrima de Schirmer, biomicroscopia com lâmpada de fenda, prova da fluoresceína e oftalmoscopia indireta. Sete medidas de PIO foram realizadas no mesmo dia (às 6h, 9h, 12h, 15h, 18h, 21h e 24h). O teste t foi usado quando havia dois grupos de comparações e a análise de variância (ANOVA) foi usada para detectar diferenças na PIO entre os momentos de medição e entre as categorias de idade. A PIO média foi de 28,4 ± 3,7 mmHg para todos os olhos. A PIO média dos grupos I, II e III foi de 29,2 ± 3,5, 28,4 ± 4,3 e 27,7 ± 3,2 mmHg, respectivamente. Não houve diferença estatística entre os olhos direito e esquerdo. Houve diferença entre o grupo I e o grupo III (P = 0,008). Não houve diferença entre as medidas registradas em diferentes momentos do dia (P = 0,560). A PIO aferida com tonômetro de rebote em cavalos da raça Crioula saudáveis não sofreu influência da hora do dia em que foi realizada, mas foi menor em equinos mais velhos.(AU)

Acute effects of ropivacaine hydrochloride on corneal endothelial cell ultrastructure of horses: ex vivo study / Efeitos agudos do cloridrato de ropivacaína na ultraestrutura das células endoteliais da córnea de equinos: estudo ex vivo

The objective of this study was to evaluate the acute effects of ropivacaine hydrochloride on the corneal endothelium of horses. Forty-eight eyes were obtained from a commercial slaughterhouse and were randomly divided into three groups. In group A, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 60 seconds. In group B, the corneal endothelium was exposed to 0.75% ropivacaine hydrochloride for 15 minutes. In group C, the corneal endothelium was exposed to a balanced saline solution for 60 seconds. Afterwards, all samples were prepared for evaluation with scanning electron microscopy. Random electromicrographs were obtained from each sample. The images were analysed and, with the aid of software, areas with no endothelial cells were measured. The average endothelial loss, expressed as a percentage in relation to the total area, of the samples in group A was 5.28%. The average endothelial loss of samples from group B, expressed as a percentage in relation to the total area, was 20.39%. The damage to the corneal endothelium was significantly greater in group B compared to groups A and C. It was possible to conclude that 0.75% ropivacaine hydrochloride induced acute damage to corneal endothelium cells.(AU)

Objetivou-se avaliar os efeitos agudos do cloridrato de ropivacaína no endotélio da córnea de equinos. Quarenta e oito olhos de equinos foram divididos aleatoriamente em três grupos. No grupo A o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 60 segundos. No grupo B o endotélio da córnea foi exposto a cloridrato de ropivacaína a 0,75% por 15 minutos. No grupo C o endotélio da córnea foi exposto à solução salina balanceada por 60 segundos. As amostras foram preparadas para avaliação com microscopia eletrônica de varredura. Eletromicrografias eletrônicas de varredura foram obtidas aleatoriamente de cada amostra. As imagens foram analisadas e, com o auxílio de um programa para morfometria foram medidas as áreas sem células endoteliais. A perda endotelial média foi expressa em porcentagem em relação à área total das amostras do grupo A foi de 5,28%. A perda endotelial média de amostras do grupo B foi expressa em porcentagem em relação à área total, foi de 20,39%. O dano ao endotélio da córnea foi significativamente maior no grupo B, comparado aos grupos A e C. O cloridrato de ropivacaína a 0,75% induziu dano agudo nas células do endotélio da córnea de equinos.(AU)