A technique for shotgun proteomic analysis of the precorneal tear film in dogs with naturally occurring primary glaucoma.

OBJECTIVE: To introduce a protocol for the characterization of protein patterns in tears of dogs with primary angle closure glaucoma (PACG) and primary open-angle glaucoma (POAG). ANIMALS: Nineteen dogs (25 eyes). METHODS: Tear samples were collected using a Schirmer tear strip, from dogs with PACG (PACG-affected eyes, n = 8; unaffected eyes predisposed to PACG, n = 7), POAG (n = 4), and healthy controls (n = 6). Protein precipitation and trypsin digestion were performed for analyses via liquid chromatography-tandem mass spectrometry. Proteins were identified using the SwissProt protein sequence database. Relative protein expression in 17 eyes (15 dogs) was evaluated using Proteome Discoverer 2.0. Pathway analyses were performed to investigate molecular mechanisms associated with primary glaucoma. RESULTS: Unique peptides were identified in 505 proteins, with Major allergen Can f 1 and albumin identified with high confidence. Proteins unique to tears from diseased eyes (PACG: n = 7; POAG: n = 14) were identified. Nucleoside diphosphate was unique to tears in PACG eyes naïve to therapy, while retinal binding protein and NSFL1 cofactor p47 were unique to medicated PACG eyes. Relative expression of 34 proteins differed between disease states. Pathway analyses identified that the 'inflammatory response' was among the top disease/disorders in dogs with primary glaucoma (PACG and POAG) but not in healthy controls. CONCLUSION: Tear samples suitable for mass spectrometry were readily obtained from pet dogs without needing specialized equipment. Further studies to validate the findings and explore potential candidate biomarkers for early disease detection and potential therapeutic targets are indicated.

Optical coherence tomography of the retina, nerve fiber layer, and optic nerve head in dogs with glaucoma.

OBJECTIVE: To evaluate the retina and optic nerve head (ONH) in canine eyes predisposed to glaucoma using optical coherence tomography (OCT). ANIMALS: Twenty-five eyes (24 dogs). METHODS: Measures of peripapillary retinal, retinal nerve fiber layer (RNFL), and ganglion cell complex (GCC) thickness and ONH parameters were obtained in vivo by OCT of the unaffected eye in dogs diagnosed with unilateral primary glaucoma (predisposed; n = 12) and compared with measures of healthy control eyes (normal; n = 13). Repeatability and intrarater reliability were explored using intraclass correlation coefficients (ICC). RESULTS: Compared to normal eyes, predisposed eyes had a thinner retina in the temporal (P = 0.005), inferior quadrants (P = 0.003), and decreased inner retinal thickness (superior: P = 0.003, temporal: P = 0.001, inferior: P < 0.001, nasal: P = 0.001). Predisposed eyes had a thinner RNFL compared to normal eyes (P = 0.005), and when analyzed in quadrants, it was thinner in the superior (P < 0.001), temporal (P = 0.034), and nasal quadrants (P = 0.001). Repeatability (ICC 0.763-0.835) and intrarater reliability (ICC 0.824-0.942) were good to excellent for measures of retinal thickness and adequate for RNFL measurements (ICC 0.701-0.798). Reliable measurements of optic disk area were obtained and were similar between groups (P = 0.597). Measurements of parameters relying on automated software detection (GCC, optic cup, optic rim) had inadequate repeatability and reliability. CONCLUSION: Statistically significant differences in retinal and RNFL thicknesses were identified in normal and predisposed eyes. Reliable and consistent measurements of variables with manual adjustment of software detected parameters were obtained. Validation of OCT as a diagnostic tool for clinical assessment in canine glaucoma is warranted.

Development of a vision impairment score for the assessment of functional vision in dogs: Initial evidence of validity, reliability, and responsiveness.

AIM: To describe the development and initial validation of a questionnaire measuring functional vision in dogs. METHODS: A 17-item survey was designed to quantify functional vision in dogs. The Vision Impairment Score (VIS) was determined by summing responses to each question. Questions were assigned to one of five subcategories: overall vision, daily activities, peripheral vision, near vision, and distance vision. Content validity was established during development phases, and construct validity via comparing results of known groups (blind vs sighted; normal vs impaired vision; surgery to improve vision vs nonrestorative surgery), and through factor analysis. Concurrent criterion validity was determined with use of a validated health-related quality-of-life (HRQL) assessment tool. Reliability and responsiveness assessments were investigated using intraclass correlation coefficient (ICC) and effect size (ES), respectively. RESULTS: Responses (221) from 201 dog owners were included. Compared to sighted dogs (n = 153), blind dogs (n = 48) had a higher VIS and greater impairment in all subcategories. Among sighted dogs, a higher VIS was obtained in dogs with low vision compared to those with normal vision (P < 0.001). A higher VIS was associated with poorer HRQL (P < 0.001). Perfect reliability was obtained for 6/17 questions, and excellent reliability for 11/17 questions (intraclass correlation 1.0 and >0.9, respectively), and the VIS was highly responsive to therapeutic intervention (effect size 1.46). CONCLUSION: Results suggest the VIS may be clinically useful in assessing and obtaining a quantifiable measure of functional vision in dogs. Ongoing validation of the tool for clinical use is needed.

Solitary (primary) uveal T-cell lymphoma in a horse.

A 22-year-old Australian stockhorse gelding was presented with anterior uveitis in the right eye which was nonresponsive to anti-inflammatory therapy. Clinical examination revealed corneal edema and vascularization, marked hypopyon, and thickening of the dorsal iris, which was confirmed by ultrasonography. Hematologic and biochemical analyses, abdominal and thoracic ultrasonography, and abdominocentesis with cytologic and biochemical analysis revealed no significant abnormalities. Cytological examination of an aqueous humor sample revealed a population of predominantly large lymphoblasts with high nuclear-to-cytoplasmic ratio, round or irregular nuclei, clumped nuclear chromatin, multiple large prominent nucleoli, and a small volume of basophilic cytoplasm. The cytologic diagnosis was intraocular lymphoma. Biopsy of the right submandibular lymph node revealed no evidence of neoplastic invasion. Euthanasia and a complete necropsy were performed and revealed no evidence of neoplasia in any tissue other than the right eye, which had an extensive, well-defined infiltrate of neoplastic lymphocytes expanding the ciliary body and iris, infiltrating the ciliary epithelium, and extending into the pars plana and peripheral choroid. Immunohistochemistry confirmed that neoplastic cells expressed the T-cell marker CD3. To the authors' knowledge, this is the first description of primary, solitary uveal T-cell lymphoma in a horse. Although apparently rare, lymphoma should be considered in horses with uveitis, even when inflammation is unilateral and in the absence of extraocular signs of neoplasia. Aqueocentesis and cytological examination provided an antemortem diagnosis in this case and should be considered as a diagnostic tool for investigation of uveal thickening and hypopyon.