Veterinary Ophthalmology (2015) 18, 5, 371–380
DOI:10.1111/vop.12209
Clinical and histological characteristics of canine ocular gliovascular syndrome Amy Treadwell,* Carolina Naranjo,† Tiffany Blocker,‡ Mitzi Zarfoss§ and Richard R. Dubielzig¶ *Carolina Veterinary Specialists, Charlotte, NC, USA; †Departamento de Medicina y Cirugıa Animales, Universidad Complutense de Madrid, Madrid, Spain; ‡Irvine, CA, USA; §PETS Referral Center, Berkeley, CA, USA; and ¶Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, WI, USA
Address communications to: C. Naranjo Tel.: +34-913943861 Fax: +34-913943808 e-mail: [email protected]
Abstract Objective To characterize the clinical, diagnostic, and histopathologic findings in dogs with canine ocular gliovascular syndrome (COGS). Procedures The archives at the Comparative Ocular Pathology Laboratory of Wisconsin (COPLOW) were used to identify eyes with COGS. Histopathological inclusion criteria included: a neovascular membrane extending from the optic nerve head or retina, clusters of spindle cells lacking vascularization within the vitreous, and histological signs of glaucoma. Special and immunohistochemical (IHC) staining techniques were performed. Clinical data, treatments, and outcomes were obtained from case records and information provided by submitting veterinarians. Results Thirty-seven eyes of 36 dogs were identified with COGS. The average age at diagnosis was 8.8 years (2.2). The relative risk for a Labrador retriever affected by COGS was significantly greater (9.3 times) (P < 0.0001) when compared to all other dog breeds within the COPLOW database. Most dogs presented with hyphema and secondary glaucoma; average intraocular pressure was 39 mmHg (19). Average time to enucleation or evisceration was 27 days. Vitreal cells stained positive with IHC for glial fibrillary acidic protein in 14 of 17 globes, and vascular endothelial growth factor was expressed in the vitreal cells in five of five globes. Conclusions We have defined a syndrome associated with vitreal glial cell aggregates and neovascular proliferation from the optic nerve or retina, which leads to neovascular glaucoma. The inflammation and secondary glaucoma resulting from this syndrome appear poorly responsive to conventional medical therapies. The exact etiology of COGS remains undetermined, but a systemic etiology is unlikely. Key Words: canine ocular gliovascular syndrome, dog, glial cells, intraocular hemorrhage, intravitreal membranes, neovascular glaucoma
INTRODUCTION
Canine ocular gliovascular syndrome (COGS) is a syndrome associated with intravitreal spindle cell aggregates, optic nerve or retinal neovascular membranes, intraocular hemorrhage, and neovascular glaucoma. This condition was previously described in a histological review of five canine globes containing intravitreal cellular membranes, intravitreal hemorrhage, pre-iridal fibrovascular membranes, and glaucoma.1 The name canine ocular gliovascular syndrome was introduced by Naranjo et al.2 at the 2006 American College of Veterinary Ophthalmologists (ACVO) conference in an abstract which described the histological findings of 21 canine globes. © 2014 American College of Veterinary Ophthalmologists
Neovascular glaucoma (NVG) is a well-described cause of visual impairment and blindness in humans. A form of secondary glaucoma, NVG, is associated with a number of underlying conditions recognized in humans: diabetic retinopathy, central retinal vein occlusion, carotid occlusion, ocular ischemic syndrome, trauma, chronic uveitis, ocular radiation, and neoplasia.3,4 Retinal ischemia, common to the aforementioned diseases, triggers the release of angiogenic factors such as vascular endothelial growth factor (VEGF)5 and interleukin-6.6 VEGF has been implicated as an important factor in the pathogenesis of ocular neovascularization and NVG.5,7–9 Pre-iridal fibrovascular membrane (PIFM) formation on the anterior iris surface and extending into the iridocorneal angle results in
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peripheral anterior synechiae, angle closure, obstruction of aqueous outflow, and increase in intraocular pressure (IOP).3,10 Neovascularization in the anterior segment can also lead to hyphema and subsequent worsening of secondary glaucoma. The best treatment for NVG is management of the causative disease. Early recognition of NVG and initiation of therapies are critical. Although familiar to the pathologist,10 NVG is rarely recognized clinically in veterinary medicine. The aims of this retrospective study were to obtain and assess clinical and diagnostic findings, evaluate the course of therapy, and further explore histologic and immunohistochemical features of dogs diagnosed with COGS. These results were analyzed in an effort to determine whether a common etiology exists among affected dogs. MATERIALS AND METHODS
Samples from 37 eyes from 36 dogs (35 globes and two evisceration specimens) affected by COGS were identified using the archives of the Comparative Ocular Pathology Laboratory of Wisconsin (COPLOW) within the time period: January 1992 to July 2009. Twenty globes from the 2006 Naranjo abstract2 were included in this retrospective analysis. Light microscopy of hematoxylin and eosin (H&E)-stained specimens was performed to confirm the inclusion criteria for COGS. Three criteria were used: clusters of cells within the vitreous that lack vascular supply; the presence of neovascular membrane proliferation extending from the optic nerve head and/or peripheral retina; and two or more histological signs of glaucoma (i.e., ganglion cell loss, optic nerve cupping, peripheral anterior synechiae). Submitting veterinarians were contacted via a combination of email, standard mail, and fax. Questionnaires were provided inquiring about the signalment, history, ophthalmic examination findings, diagnostic test results, treatment, and outcome of each dog affected with COGS. Patient records were evaluated when provided. In cases for which a questionnaire was not returned or clinical information was not available, clinical data were limited to information included on the original submission forms that accompanied the specimens to COPLOW. The following histochemical stains and techniques were used to further characterize histologic changes: Periodic acid–Schiff (PAS) method (n = 22), Alcian blue at pH 2.5 (n = 21), Prussian blue (n = 20), and Masson’s trichrome (n = 18). Immunohistochemical staining methods were performed using antibodies for glial fibrillary acid protein (GFAP) (n = 17) and VEGF (n = 5). Immunohistochemistry (IHC) for GFAP was performed as follows: slides were rehydrated, and endogenous peroxidase activity was quenched using 3% hydrogen peroxide (Cumberland Swan Hydrogen Peroxide, Fisher Scientific, Chicago, IL, USA) for 5 min. The remaining procedures were completed in an automated stainer (Lab Vision
Autostainer 720 2D, Thermo Fisher Scientific, Fitchburg, WI, USA) and included a 10-min nonspecific binding blocker (Ultra V Block, Lab Vision, Thermo Fisher Scientific), primary antibody (rabbit polyclonal anti-human GFAP, Dako, Carpinteria, CA, USA) at a dilution of 1:1500 for 30 min, biotinylated secondary antibody (HRP polymer, Lab Vision, Thermo Fisher Scientific) for 15 min, and 3,3-diaminobenzidine chromogen solution (DAB+, Lab Vision, Thermo Fisher Scientific) for 5 min. Slides were rinsed in a buffer solution (PBS with Tween, Lab Vision, Thermo Fisher Scientific) after every step. Finally, slides were rinsed in tap water, counterstained with Harris hematoxylin for 2 min, dehydrated, and cover-slipped. The external positive control consisted of a section of canine brain, while unaffected retina or optic nerve was used as an internal positive control. Negative controls were processed in the same way using buffer solution in place of the primary antibody. IHC for VEGF was performed in a preliminary study of five randomly selected globes at the University of Illinois Veterinary Diagnostic Laboratory as previously described.11 Statistical analysis was performed using the statistical software package SAS v9 (SAS Institute Inc., Cary, NC, USA). Descriptive statistics including mean, median, and standard deviation were calculated for age, initial intraocular pressure, average blood pressure, and time to enucleation. Simple point estimates of the proportion and 95% confidence interval (CI) estimates were calculated for GFAP staining and breed affected. Relative risk for COGS was calculated for Labrador retrievers vs. all other breeds. RESULTS
Clinical characteristics Questionnaires and case history information were obtained for 28 of the 36 affected dogs. Signalment and clinical history Thirty-seven eyes (36 dogs) were identified with COGS. The average age at the time of diagnosis was 8.8 years (years) (2.2) with a median age of 9.4 years (range: 3.5–13). Eighteen males and 18 females were affected. There were five intact and 13 neutered dogs in each group. The right eye (OD) was affected in 20 dogs; the left eye (OS) in 15 dogs. Two submissions did not state whether the globe was right or left. One dog was bilaterally (OU) affected; OD was submitted 5 months before OS. Sixteen dogs were noted to have brown eyes; however, information regarding iris color was not available for the remaining dogs. Breed Labrador retrievers and retriever mixes accounted for 64% of the breeds affected (23/36). There were 20 pure-bred Labrador retrievers and three retriever mixes affected. Other breeds affected included: two English springer spaniels, two basset hounds, two terrier mixes, and one each of golden retriever, Samoyed, English
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bulldog, pug, German shorthaired pointer, collie, and border collie mix. At the conclusion of data acquisition (2009), there were a total of 19 351 canine cases in the COPLOW archives. Of these, 2293 were pure-bred Labrador retrievers. Within the COGS population, 20 of 36 dogs (56%) were pure-bred Labrador retrievers. These data were used to determine that the relative risk of COGS in the Labrador retriever breed which was 9.3 times that of all other dog breeds in the population examined; this value is significant (P < 0.0001). The 95% confidence interval for this relative risk was 4.8–17.9.
Presenting complaint One or more presenting complaints were recorded for each dog. The most common were a red eye in 12 cases, a cloudy eye in seven cases, intraocular hemorrhage in six cases, and a painful eye in five cases. Less commonly reported presenting complaints were glaucoma (n = 4), mucoid discharge (n = 3), uveitis (n = 3), mydriasis (n = 2), blindness (n = 2), cataract (n = 2), elevated third eyelid (n = 1), pain when yawning (n = 1), change in eye color (1), swollen eye (n = 1), and epiphora (n = 1). Nonophthalmic findings Sixteen cases (44%) were reported to have concurrent systemic abnormalities. Allergic skin disease and otitis were reported in three and four dogs, respectively. Hypothyroidism controlled with thyroid hormone replacement was reported in two dogs. Peripheral lymphadenopathy of a single lymph node was noted in two dogs. One of them had a self-limiting submandibular lymphadenomegaly without cytologic evaluation, and the other, a cytologically confirmed reactive popliteal lymph node. One dog had doxycyclineresponsive cutaneous ecchymotic lesions. This dog had a normal coagulation panel, was Ehrlichia canis (E. canis) and heartworm antigen negative, but had a low positive titer to Ehrlichia equi (E. equi), Borrelia burgdorferi (Lyme), and Rickettsia rickettsii (Rocky Mountain spotted fever – RMSF). Additional findings reported were osteoarthritis in two dogs and one dog each was affected by brachycephalic airway syndrome, subcutaneous lipoma, hypertriglyceridemia, cutaneous hemangioma, seizures, splenic nodular hyperplasia, hepatic mass, hematochezia (while on oral prednisone), and elbow dysplasia. Oculo-skeletal dysplasia was not reported in any of the patients in this study. Twenty of 36 dogs (56%) had no abnormal systemic findings. Ophthalmic examination findings (affected eye) Eight of 37 eyes (22%) had reported buphthalmia. In 5/37, adnexal abnormalities were listed for the affected eye including: two eyelid masses, one with microblepharon, one with ectropion, and one with everted third eyelid cartilage. Corneal abnormalities were reported in 24 eyes (65%), corneal edema being the most common (22/37, 59%).
Additional corneal abnormalities reported were as follows: three eyes with exposure keratitis, two eyes with ulceration, and one each of pannus, Haab’s striae, and deep neovascularization. Hyphema was reported in 26 of 37 eyes (70%). Anterior uveitis was also a common finding, with 22 eyes affected (59%). Seventeen of the 22 eyes with anterior uveitis had flare, graded on a scale of 1–4. Flare was reported as mild to moderate in most cases; the flare score was 1/4 in five eyes, 2/4 in six eyes, 3/4 in four eyes, and 4/4 in two eyes. Fibrin and hypopyon were reported in one eye each. Cataract was noted in 13 eyes, and the stage of the cataract was described in nine of them. Cataracts were described as posterior cortical cataracts in five eyes, hypermature in 2, incipient in 1, and immature in 1. Vitreal hemorrhage was reported in seven of 37 eyes (Fig. 1). White intravitreal lesions were described in five eyes. Two had white, fluffy debris within the vitreous, one had a white arborizing structure anterior to the optic nerve head, one had white anterior and posterior vitreal opacities along with a white ‘vascular-like’ structure anterior to the optic nerve head (Fig. 2), and one had a dense white vitreal opacity located ventrally. One additional eye was reported to have pre-existing retinal dysplasia and subsequent retinal detachment.
Contralateral eye abnormalities One or more abnormalities were reported to be present in the contralateral eye of 14/36 dogs (39%). Eleven dogs of those 14 (79%) had lesions involving the posterior segment. Chorioretinal scarring and/or altered tapetal reflectivity was described in four dogs. Anterior vitreal opacities were noted in three dogs, and white/yellow intravitreal aggregates were reported in two dogs. One of these two dogs with intravitreal aggregates had histologic confirmation of COGS
Figure 1. Vitreal hemorrhage seen as a dark red to gray discoloration that obscures the fundus (center and mid left of the picture) in a COGS-affected dog (Photo courtesy of Dr. Glenwood Gum).
© 2014 American College of Veterinary Ophthalmologists, Veterinary Ophthalmology, 18, 371–380
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Figure 2. White anterior and posterior vitreal opacities (Photo courtesy of Dr. Kerry Ketring).
following enucleation of the contralateral eye. The globe from the other dog was not submitted for histopathology. Retinal folds/dysplasia was noted in three dogs. Cataracts were noted in two dogs. Vitreal hemorrhage, a hyaloid remnant, papilledema, glaucoma, anterior uveitis, and keratoconjunctivitis sicca (KCS) were reported, in one dog each.
Diagnostics Schirmer tear test (STT) values were recorded in 14 of 36 dogs, 13 of 14 were considered normal. An abnormal STT (0 mm wetting/min) was recorded for one dog with previously diagnosed KCS in the affected eye. Intraocular pressure values were recorded in 30 of 37 eyes. The average IOP at initial presentation was 39 mmHg (19). The median IOP was 38 mmHg (range: 6–77). Gonioscopy of the contralateral eye was performed in nine dogs, of which six had a normal appearing iridocorneal angle. Ocular ultrasound findings were reported for 11 dogs. Echogenic vitreal debris extending from the peripheral retina and/or optic nerve head was described in seven dogs (Fig. 3), retinal detachment was reported in six dogs, and a posterior segment mass effect was reported in two dogs. Cytological evaluation of aqueous humor in one eye and subretinal fluid in one eye was nondiagnostic in the two dogs with a mass effect. A normal flash electroretinogram was recorded in one dog, and fluorescein angiography in the same dog revealed a filling defect and partial retinal detachment. Indirect systolic blood pressure (BP) measurements were obtained in eight of 36 dogs. The mean systolic BP at presentation was 140 mmHg (22). A variety of additional diagnostics were performed including: complete blood count (CBC) and serum chemistry (n = 23), coagulation
Figure 3. Echogenic vitreal debris extending from the peripheral retina and/or optic nerve head was described in seven dogs. This image is the right globe from a Labrador retriever within the study population and displays multifocal, curvilinear echogenicities extending from the peripheral retina (white arrow).
profile (n = 5), urinalysis (n = 3), infectious disease titers (tick borne disease titers (n = 10), fungal disease titers (n = 3), heartworm antigen (n = 3)), imaging (thoracic radiographs (n = 7), abdominal radiographs (n = 1), abdominal ultrasound (n = 1)), and lymph node cytology (n = 1). No common diagnostic findings were observed. Fourteen of the 23 CBC/chemistry panels were within normal limits. The remaining nine had mild elevations in alkaline phosphatase (ALP), alanine transaminase (ALT), and triglycerides. Diabetes mellitus was not reported in any dog. The specific etiologic agent, which constituted a ‘tick borne disease titer’ or a ‘fungal titer’, was not always stated. The following infectious titers were listed: E. canis, E. equi, Lyme, RMSF, Bartonella spp., Histoplasma spp., Blastomyces spp., Aspergillus spp., Cocccidiodies spp., Toxoplasma spp., and Cryptococcus spp. Reportedly, one dog had a low positive titer to E. equi, Lyme and RMSF and another had a low positive Lyme titer (thought to be secondary to vaccine).
Treatments Over the course of disease, a variety of drug combinations were reportedly used, including methazolamide, dorzolamide, brinzolamide, timolol, lantanoprost, flurbiprofen, neopolydexamethasone, prednisolone acetate, betamethasone, amoxicillin/clavulanic acid, orbifloxacin, doxycycline, itraconazole, fluconazole, carprofen, and
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acetaminophen. Intraocular pressure and inflammation control were not achieved long term on the above listed medical therapies. One dog received an intravitreal injection of 1.25 mg bevacizumab and an intracameral injection of 37.5 lg tissue plasminogen activator (TPA), 0.75 mg triamcinolone, and 0.05 mg moxifloxacin, which appeared to improve signs of uveitis and glaucoma. This improvement was temporary, as signs of uveitis and elevated IOP returned approximately 3 weeks following injection. Repeat bevacizumab injection was declined by the owner. Thirty-five of 37 eyes underwent enucleation of the affected globe, and two eyes underwent evisceration with intraocular prosthesis. Evisceration samples were diagnosed with COGS based on evidence of the three previously mentioned inclusion criteria. The mean time from initial presentation to enucleation/evisceration surgery was 27 days (34) with a range of 0–135 days.
Histopathological characteristics All 37 eyes had intravitreal neovascular membranes characterized by an eosinophilic, ‘glassy’, collagenous matrix surrounding tortuous delicate blood vessels (Fig. 4). Twenty-two of 36 eyes (61%) had membrane extension from the optic nerve. Nine eyes had extension of membranes from the retina, and five eyes had membrane extension from both retina and optic nerve. Intravitreal cellular clusters, also present in all 37 eyes, were located in the anterior vitreous in 30 of 36 globes (83%) (Fig. 5). The origin of the neovascular membrane and the location of these cells in one evisceration specimen could not be accurately determined due to tissue distortion as a result of fixation; therefore, percentages are calculated out of 36. In four specimens, the cells were
Figure 4. Neovascular membrane embedded in an eosinophilic homogeneous matrix arising from the optic nerve head and extending into the vitreous (center of the picture). Hemorrhage can be seen in the vitreous and optic nerve head. Hematoxylin and eosin (HE), original magnification 29.
Figure 5. Cluster of nonvascularized spindle (glial) cells within the anterior vitreous, immediately posterior to the lens, which can be seen in the top of the picture. HE, original magnification 209.
present within the posterior vitreous, and in two specimens, cells were found in both the anterior and posterior vitreous. The intravitreal cellular clusters were characterized by tufts of loosely arranged, plump spindle cells, with no obvious connection to structures within the globe. PIFM was seen in all 37 globes, and peripheral anterior synechiae were present in 36/36 cases in which evaluation was possible (peripheral anterior synechiae could not be assessed in the evisceration sample) (Fig. 6). Intraocular hemorrhage and retinal detachment were common findings among COGS eyes; intraocular hemorrhage was identified in 31 of 35 specimens (89%), and retinal detachment was histologically confirmed in 24 of 35 globes (69%). Optic nerve cupping or gliosis was observed in 27
Figure 6. Pre-iridal fibrovascular membrane carpeting the anterior surface of the iris, with peripheral anterior synechiae, lining, and collapsing the iridocorneal angle. HE, original magnification 109.
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of 35 specimens (77%). PIFM, peripheral anterior synechiae, and optic nerve head could not be examined in two eyes due to tissue loss at sectioning. Retinal ganglion cell (RGC) numbers were decreased or absent in 35 of 37 specimens (95%). Of the 35, 15 specimens had decreased RGC numbers (
DOI:10.1111/vop.12209
Clinical and histological characteristics of canine ocular gliovascular syndrome Amy Treadwell,* Carolina Naranjo,† Tiffany Blocker,‡ Mitzi Zarfoss§ and Richard R. Dubielzig¶ *Carolina Veterinary Specialists, Charlotte, NC, USA; †Departamento de Medicina y Cirugıa Animales, Universidad Complutense de Madrid, Madrid, Spain; ‡Irvine, CA, USA; §PETS Referral Center, Berkeley, CA, USA; and ¶Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, WI, USA
Address communications to: C. Naranjo Tel.: +34-913943861 Fax: +34-913943808 e-mail: [email protected]
Abstract Objective To characterize the clinical, diagnostic, and histopathologic findings in dogs with canine ocular gliovascular syndrome (COGS). Procedures The archives at the Comparative Ocular Pathology Laboratory of Wisconsin (COPLOW) were used to identify eyes with COGS. Histopathological inclusion criteria included: a neovascular membrane extending from the optic nerve head or retina, clusters of spindle cells lacking vascularization within the vitreous, and histological signs of glaucoma. Special and immunohistochemical (IHC) staining techniques were performed. Clinical data, treatments, and outcomes were obtained from case records and information provided by submitting veterinarians. Results Thirty-seven eyes of 36 dogs were identified with COGS. The average age at diagnosis was 8.8 years (2.2). The relative risk for a Labrador retriever affected by COGS was significantly greater (9.3 times) (P < 0.0001) when compared to all other dog breeds within the COPLOW database. Most dogs presented with hyphema and secondary glaucoma; average intraocular pressure was 39 mmHg (19). Average time to enucleation or evisceration was 27 days. Vitreal cells stained positive with IHC for glial fibrillary acidic protein in 14 of 17 globes, and vascular endothelial growth factor was expressed in the vitreal cells in five of five globes. Conclusions We have defined a syndrome associated with vitreal glial cell aggregates and neovascular proliferation from the optic nerve or retina, which leads to neovascular glaucoma. The inflammation and secondary glaucoma resulting from this syndrome appear poorly responsive to conventional medical therapies. The exact etiology of COGS remains undetermined, but a systemic etiology is unlikely. Key Words: canine ocular gliovascular syndrome, dog, glial cells, intraocular hemorrhage, intravitreal membranes, neovascular glaucoma
INTRODUCTION
Canine ocular gliovascular syndrome (COGS) is a syndrome associated with intravitreal spindle cell aggregates, optic nerve or retinal neovascular membranes, intraocular hemorrhage, and neovascular glaucoma. This condition was previously described in a histological review of five canine globes containing intravitreal cellular membranes, intravitreal hemorrhage, pre-iridal fibrovascular membranes, and glaucoma.1 The name canine ocular gliovascular syndrome was introduced by Naranjo et al.2 at the 2006 American College of Veterinary Ophthalmologists (ACVO) conference in an abstract which described the histological findings of 21 canine globes. © 2014 American College of Veterinary Ophthalmologists
Neovascular glaucoma (NVG) is a well-described cause of visual impairment and blindness in humans. A form of secondary glaucoma, NVG, is associated with a number of underlying conditions recognized in humans: diabetic retinopathy, central retinal vein occlusion, carotid occlusion, ocular ischemic syndrome, trauma, chronic uveitis, ocular radiation, and neoplasia.3,4 Retinal ischemia, common to the aforementioned diseases, triggers the release of angiogenic factors such as vascular endothelial growth factor (VEGF)5 and interleukin-6.6 VEGF has been implicated as an important factor in the pathogenesis of ocular neovascularization and NVG.5,7–9 Pre-iridal fibrovascular membrane (PIFM) formation on the anterior iris surface and extending into the iridocorneal angle results in
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peripheral anterior synechiae, angle closure, obstruction of aqueous outflow, and increase in intraocular pressure (IOP).3,10 Neovascularization in the anterior segment can also lead to hyphema and subsequent worsening of secondary glaucoma. The best treatment for NVG is management of the causative disease. Early recognition of NVG and initiation of therapies are critical. Although familiar to the pathologist,10 NVG is rarely recognized clinically in veterinary medicine. The aims of this retrospective study were to obtain and assess clinical and diagnostic findings, evaluate the course of therapy, and further explore histologic and immunohistochemical features of dogs diagnosed with COGS. These results were analyzed in an effort to determine whether a common etiology exists among affected dogs. MATERIALS AND METHODS
Samples from 37 eyes from 36 dogs (35 globes and two evisceration specimens) affected by COGS were identified using the archives of the Comparative Ocular Pathology Laboratory of Wisconsin (COPLOW) within the time period: January 1992 to July 2009. Twenty globes from the 2006 Naranjo abstract2 were included in this retrospective analysis. Light microscopy of hematoxylin and eosin (H&E)-stained specimens was performed to confirm the inclusion criteria for COGS. Three criteria were used: clusters of cells within the vitreous that lack vascular supply; the presence of neovascular membrane proliferation extending from the optic nerve head and/or peripheral retina; and two or more histological signs of glaucoma (i.e., ganglion cell loss, optic nerve cupping, peripheral anterior synechiae). Submitting veterinarians were contacted via a combination of email, standard mail, and fax. Questionnaires were provided inquiring about the signalment, history, ophthalmic examination findings, diagnostic test results, treatment, and outcome of each dog affected with COGS. Patient records were evaluated when provided. In cases for which a questionnaire was not returned or clinical information was not available, clinical data were limited to information included on the original submission forms that accompanied the specimens to COPLOW. The following histochemical stains and techniques were used to further characterize histologic changes: Periodic acid–Schiff (PAS) method (n = 22), Alcian blue at pH 2.5 (n = 21), Prussian blue (n = 20), and Masson’s trichrome (n = 18). Immunohistochemical staining methods were performed using antibodies for glial fibrillary acid protein (GFAP) (n = 17) and VEGF (n = 5). Immunohistochemistry (IHC) for GFAP was performed as follows: slides were rehydrated, and endogenous peroxidase activity was quenched using 3% hydrogen peroxide (Cumberland Swan Hydrogen Peroxide, Fisher Scientific, Chicago, IL, USA) for 5 min. The remaining procedures were completed in an automated stainer (Lab Vision
Autostainer 720 2D, Thermo Fisher Scientific, Fitchburg, WI, USA) and included a 10-min nonspecific binding blocker (Ultra V Block, Lab Vision, Thermo Fisher Scientific), primary antibody (rabbit polyclonal anti-human GFAP, Dako, Carpinteria, CA, USA) at a dilution of 1:1500 for 30 min, biotinylated secondary antibody (HRP polymer, Lab Vision, Thermo Fisher Scientific) for 15 min, and 3,3-diaminobenzidine chromogen solution (DAB+, Lab Vision, Thermo Fisher Scientific) for 5 min. Slides were rinsed in a buffer solution (PBS with Tween, Lab Vision, Thermo Fisher Scientific) after every step. Finally, slides were rinsed in tap water, counterstained with Harris hematoxylin for 2 min, dehydrated, and cover-slipped. The external positive control consisted of a section of canine brain, while unaffected retina or optic nerve was used as an internal positive control. Negative controls were processed in the same way using buffer solution in place of the primary antibody. IHC for VEGF was performed in a preliminary study of five randomly selected globes at the University of Illinois Veterinary Diagnostic Laboratory as previously described.11 Statistical analysis was performed using the statistical software package SAS v9 (SAS Institute Inc., Cary, NC, USA). Descriptive statistics including mean, median, and standard deviation were calculated for age, initial intraocular pressure, average blood pressure, and time to enucleation. Simple point estimates of the proportion and 95% confidence interval (CI) estimates were calculated for GFAP staining and breed affected. Relative risk for COGS was calculated for Labrador retrievers vs. all other breeds. RESULTS
Clinical characteristics Questionnaires and case history information were obtained for 28 of the 36 affected dogs. Signalment and clinical history Thirty-seven eyes (36 dogs) were identified with COGS. The average age at the time of diagnosis was 8.8 years (years) (2.2) with a median age of 9.4 years (range: 3.5–13). Eighteen males and 18 females were affected. There were five intact and 13 neutered dogs in each group. The right eye (OD) was affected in 20 dogs; the left eye (OS) in 15 dogs. Two submissions did not state whether the globe was right or left. One dog was bilaterally (OU) affected; OD was submitted 5 months before OS. Sixteen dogs were noted to have brown eyes; however, information regarding iris color was not available for the remaining dogs. Breed Labrador retrievers and retriever mixes accounted for 64% of the breeds affected (23/36). There were 20 pure-bred Labrador retrievers and three retriever mixes affected. Other breeds affected included: two English springer spaniels, two basset hounds, two terrier mixes, and one each of golden retriever, Samoyed, English
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bulldog, pug, German shorthaired pointer, collie, and border collie mix. At the conclusion of data acquisition (2009), there were a total of 19 351 canine cases in the COPLOW archives. Of these, 2293 were pure-bred Labrador retrievers. Within the COGS population, 20 of 36 dogs (56%) were pure-bred Labrador retrievers. These data were used to determine that the relative risk of COGS in the Labrador retriever breed which was 9.3 times that of all other dog breeds in the population examined; this value is significant (P < 0.0001). The 95% confidence interval for this relative risk was 4.8–17.9.
Presenting complaint One or more presenting complaints were recorded for each dog. The most common were a red eye in 12 cases, a cloudy eye in seven cases, intraocular hemorrhage in six cases, and a painful eye in five cases. Less commonly reported presenting complaints were glaucoma (n = 4), mucoid discharge (n = 3), uveitis (n = 3), mydriasis (n = 2), blindness (n = 2), cataract (n = 2), elevated third eyelid (n = 1), pain when yawning (n = 1), change in eye color (1), swollen eye (n = 1), and epiphora (n = 1). Nonophthalmic findings Sixteen cases (44%) were reported to have concurrent systemic abnormalities. Allergic skin disease and otitis were reported in three and four dogs, respectively. Hypothyroidism controlled with thyroid hormone replacement was reported in two dogs. Peripheral lymphadenopathy of a single lymph node was noted in two dogs. One of them had a self-limiting submandibular lymphadenomegaly without cytologic evaluation, and the other, a cytologically confirmed reactive popliteal lymph node. One dog had doxycyclineresponsive cutaneous ecchymotic lesions. This dog had a normal coagulation panel, was Ehrlichia canis (E. canis) and heartworm antigen negative, but had a low positive titer to Ehrlichia equi (E. equi), Borrelia burgdorferi (Lyme), and Rickettsia rickettsii (Rocky Mountain spotted fever – RMSF). Additional findings reported were osteoarthritis in two dogs and one dog each was affected by brachycephalic airway syndrome, subcutaneous lipoma, hypertriglyceridemia, cutaneous hemangioma, seizures, splenic nodular hyperplasia, hepatic mass, hematochezia (while on oral prednisone), and elbow dysplasia. Oculo-skeletal dysplasia was not reported in any of the patients in this study. Twenty of 36 dogs (56%) had no abnormal systemic findings. Ophthalmic examination findings (affected eye) Eight of 37 eyes (22%) had reported buphthalmia. In 5/37, adnexal abnormalities were listed for the affected eye including: two eyelid masses, one with microblepharon, one with ectropion, and one with everted third eyelid cartilage. Corneal abnormalities were reported in 24 eyes (65%), corneal edema being the most common (22/37, 59%).
Additional corneal abnormalities reported were as follows: three eyes with exposure keratitis, two eyes with ulceration, and one each of pannus, Haab’s striae, and deep neovascularization. Hyphema was reported in 26 of 37 eyes (70%). Anterior uveitis was also a common finding, with 22 eyes affected (59%). Seventeen of the 22 eyes with anterior uveitis had flare, graded on a scale of 1–4. Flare was reported as mild to moderate in most cases; the flare score was 1/4 in five eyes, 2/4 in six eyes, 3/4 in four eyes, and 4/4 in two eyes. Fibrin and hypopyon were reported in one eye each. Cataract was noted in 13 eyes, and the stage of the cataract was described in nine of them. Cataracts were described as posterior cortical cataracts in five eyes, hypermature in 2, incipient in 1, and immature in 1. Vitreal hemorrhage was reported in seven of 37 eyes (Fig. 1). White intravitreal lesions were described in five eyes. Two had white, fluffy debris within the vitreous, one had a white arborizing structure anterior to the optic nerve head, one had white anterior and posterior vitreal opacities along with a white ‘vascular-like’ structure anterior to the optic nerve head (Fig. 2), and one had a dense white vitreal opacity located ventrally. One additional eye was reported to have pre-existing retinal dysplasia and subsequent retinal detachment.
Contralateral eye abnormalities One or more abnormalities were reported to be present in the contralateral eye of 14/36 dogs (39%). Eleven dogs of those 14 (79%) had lesions involving the posterior segment. Chorioretinal scarring and/or altered tapetal reflectivity was described in four dogs. Anterior vitreal opacities were noted in three dogs, and white/yellow intravitreal aggregates were reported in two dogs. One of these two dogs with intravitreal aggregates had histologic confirmation of COGS
Figure 1. Vitreal hemorrhage seen as a dark red to gray discoloration that obscures the fundus (center and mid left of the picture) in a COGS-affected dog (Photo courtesy of Dr. Glenwood Gum).
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Figure 2. White anterior and posterior vitreal opacities (Photo courtesy of Dr. Kerry Ketring).
following enucleation of the contralateral eye. The globe from the other dog was not submitted for histopathology. Retinal folds/dysplasia was noted in three dogs. Cataracts were noted in two dogs. Vitreal hemorrhage, a hyaloid remnant, papilledema, glaucoma, anterior uveitis, and keratoconjunctivitis sicca (KCS) were reported, in one dog each.
Diagnostics Schirmer tear test (STT) values were recorded in 14 of 36 dogs, 13 of 14 were considered normal. An abnormal STT (0 mm wetting/min) was recorded for one dog with previously diagnosed KCS in the affected eye. Intraocular pressure values were recorded in 30 of 37 eyes. The average IOP at initial presentation was 39 mmHg (19). The median IOP was 38 mmHg (range: 6–77). Gonioscopy of the contralateral eye was performed in nine dogs, of which six had a normal appearing iridocorneal angle. Ocular ultrasound findings were reported for 11 dogs. Echogenic vitreal debris extending from the peripheral retina and/or optic nerve head was described in seven dogs (Fig. 3), retinal detachment was reported in six dogs, and a posterior segment mass effect was reported in two dogs. Cytological evaluation of aqueous humor in one eye and subretinal fluid in one eye was nondiagnostic in the two dogs with a mass effect. A normal flash electroretinogram was recorded in one dog, and fluorescein angiography in the same dog revealed a filling defect and partial retinal detachment. Indirect systolic blood pressure (BP) measurements were obtained in eight of 36 dogs. The mean systolic BP at presentation was 140 mmHg (22). A variety of additional diagnostics were performed including: complete blood count (CBC) and serum chemistry (n = 23), coagulation
Figure 3. Echogenic vitreal debris extending from the peripheral retina and/or optic nerve head was described in seven dogs. This image is the right globe from a Labrador retriever within the study population and displays multifocal, curvilinear echogenicities extending from the peripheral retina (white arrow).
profile (n = 5), urinalysis (n = 3), infectious disease titers (tick borne disease titers (n = 10), fungal disease titers (n = 3), heartworm antigen (n = 3)), imaging (thoracic radiographs (n = 7), abdominal radiographs (n = 1), abdominal ultrasound (n = 1)), and lymph node cytology (n = 1). No common diagnostic findings were observed. Fourteen of the 23 CBC/chemistry panels were within normal limits. The remaining nine had mild elevations in alkaline phosphatase (ALP), alanine transaminase (ALT), and triglycerides. Diabetes mellitus was not reported in any dog. The specific etiologic agent, which constituted a ‘tick borne disease titer’ or a ‘fungal titer’, was not always stated. The following infectious titers were listed: E. canis, E. equi, Lyme, RMSF, Bartonella spp., Histoplasma spp., Blastomyces spp., Aspergillus spp., Cocccidiodies spp., Toxoplasma spp., and Cryptococcus spp. Reportedly, one dog had a low positive titer to E. equi, Lyme and RMSF and another had a low positive Lyme titer (thought to be secondary to vaccine).
Treatments Over the course of disease, a variety of drug combinations were reportedly used, including methazolamide, dorzolamide, brinzolamide, timolol, lantanoprost, flurbiprofen, neopolydexamethasone, prednisolone acetate, betamethasone, amoxicillin/clavulanic acid, orbifloxacin, doxycycline, itraconazole, fluconazole, carprofen, and
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acetaminophen. Intraocular pressure and inflammation control were not achieved long term on the above listed medical therapies. One dog received an intravitreal injection of 1.25 mg bevacizumab and an intracameral injection of 37.5 lg tissue plasminogen activator (TPA), 0.75 mg triamcinolone, and 0.05 mg moxifloxacin, which appeared to improve signs of uveitis and glaucoma. This improvement was temporary, as signs of uveitis and elevated IOP returned approximately 3 weeks following injection. Repeat bevacizumab injection was declined by the owner. Thirty-five of 37 eyes underwent enucleation of the affected globe, and two eyes underwent evisceration with intraocular prosthesis. Evisceration samples were diagnosed with COGS based on evidence of the three previously mentioned inclusion criteria. The mean time from initial presentation to enucleation/evisceration surgery was 27 days (34) with a range of 0–135 days.
Histopathological characteristics All 37 eyes had intravitreal neovascular membranes characterized by an eosinophilic, ‘glassy’, collagenous matrix surrounding tortuous delicate blood vessels (Fig. 4). Twenty-two of 36 eyes (61%) had membrane extension from the optic nerve. Nine eyes had extension of membranes from the retina, and five eyes had membrane extension from both retina and optic nerve. Intravitreal cellular clusters, also present in all 37 eyes, were located in the anterior vitreous in 30 of 36 globes (83%) (Fig. 5). The origin of the neovascular membrane and the location of these cells in one evisceration specimen could not be accurately determined due to tissue distortion as a result of fixation; therefore, percentages are calculated out of 36. In four specimens, the cells were
Figure 4. Neovascular membrane embedded in an eosinophilic homogeneous matrix arising from the optic nerve head and extending into the vitreous (center of the picture). Hemorrhage can be seen in the vitreous and optic nerve head. Hematoxylin and eosin (HE), original magnification 29.
Figure 5. Cluster of nonvascularized spindle (glial) cells within the anterior vitreous, immediately posterior to the lens, which can be seen in the top of the picture. HE, original magnification 209.
present within the posterior vitreous, and in two specimens, cells were found in both the anterior and posterior vitreous. The intravitreal cellular clusters were characterized by tufts of loosely arranged, plump spindle cells, with no obvious connection to structures within the globe. PIFM was seen in all 37 globes, and peripheral anterior synechiae were present in 36/36 cases in which evaluation was possible (peripheral anterior synechiae could not be assessed in the evisceration sample) (Fig. 6). Intraocular hemorrhage and retinal detachment were common findings among COGS eyes; intraocular hemorrhage was identified in 31 of 35 specimens (89%), and retinal detachment was histologically confirmed in 24 of 35 globes (69%). Optic nerve cupping or gliosis was observed in 27
Figure 6. Pre-iridal fibrovascular membrane carpeting the anterior surface of the iris, with peripheral anterior synechiae, lining, and collapsing the iridocorneal angle. HE, original magnification 109.
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of 35 specimens (77%). PIFM, peripheral anterior synechiae, and optic nerve head could not be examined in two eyes due to tissue loss at sectioning. Retinal ganglion cell (RGC) numbers were decreased or absent in 35 of 37 specimens (95%). Of the 35, 15 specimens had decreased RGC numbers (
