Nasolacrimal sac foreign body extraction using vitreoretinal forceps in 28 dogs.

PURPOSE: To describe a novel technique of nasolacrimal foreign body extraction in dogs by using a 20G vitreoretinal forceps introduced through the superior lacrimal punctum. METHODS: A retrospective review of the medical records of dogs with dacryocystitis due to nasolacrimal foreign bodies between the years 2001 and 2022 was performed. We recorded the breed, age, affected eye, type and number of foreign bodies, concomitant diseases, and the use of imaging techniques. All animals underwent the same procedure of a 20G vitreoretinal forceps insertion through the upper canaliculus reaching the lacrimal sac and retrograde extraction of the foreign bodies. RESULTS: A total of 28 dogs were included, 16 males and 12 females, with a mean (±SD) age of 4.7 (±3.2) years. The most common breeds were Wire-Haired Dachshund (4/28; 14.29%) and Labrador Retriever (3/28; 10.71%). Additional imaging techniques were used, such as orbital ultrasound in 13 cases (13/28; 46.43%) and computed tomography in one case (1/28; 3.57%). The most common type of foreign body retrieved was grass awns, although seeds and plant debris were also found. Dacryocystitis resolved after removal of the foreign body and appropriate medical therapy was ensured in all cases in the 1-month postprocedure follow-up. CONCLUSION: Extraction of nasolacrimal foreign bodies with vitreoretinal forceps is a novel, noninvasive, and easily applicable technique that, although not successful in all cases, can be attempted before performing more aggressive surgery.

Natural disease history of a canine model of oligogenic RPGRIP1-cone-rod dystrophy establishes variable effects of previously and newly mapped modifier loci.

Canine RPGRIP1-cone-rod dystrophy (CRD), a model for human inherited retinal diseases (IRDs), was originally identified as autosomal recessive early-onset blindness. However, later studies revealed extensive phenotypic variability among RPGRIP1 mutants. This led to the identification of a homozygous MAP9 variant as a modifier associated with early-onset disease. Based on further phenotypic variation affecting cone photoreceptor function, we report mapping of L3 as an additional modifier locus, within a 4.1-Mb locus on canine chromosome 30. We establish the natural disease history of RPGRIP1-CRD based on up to 9-year long-term functional and structural retinal data from 58 dogs including 44 RPGRIP1 mutants grouped according to the modifier status. RPGRIP1 mutants affected by both MAP9 and L3 modifiers exhibited the most severe phenotypes with rapid disease progression. MAP9 alone was found to act as an overall accelerator of rod and cone diseases, while L3 had a cone-specific effect. Ultrastructural analysis of photoreceptors revealed varying degrees of rod and cone damage, while the connecting cilia appeared structurally preserved in all groups. We conclude that RPGRIP1-CRD is an oligogenic disease with at least three loci contributing to the pathogenesis. While the RPGRIP1 variant is required for developing the disease, MAP9 and L3 modifiers exacerbate the phenotype, individually and cumulatively. Oligogenic canine RPGRIP1-CRD illustrates the impact of multiple genetic modifiers on disease phenotype and thus has the potential to reveal new targets for broad-spectrum therapies for oligogenic or polygenic forms of human IRDs.

Retinal Vascular Plexuses Are Unequally Affected in Canine Inherited Retinal Degenerations.

Purpose: To characterize the progression of vascular changes that occur in each retinal plexus, in three canine models of inherited retinal degeneration. Methods: In this retrospective cohort study, we examined the retinal imaging records of 44 dogs from a research colony that had undergone optical coherence tomography angiography (OCTA) imaging. Animals enrolled included crd2/NPHP5 and xlpra2/RPGR mutant dogs imaged at different stages of photoreceptor loss, as well as RHOT4R/+ dogs after acute light-induced rod degeneration. Also included were normal controls imaged at similar ages. OCT angiograms of the superficial vascular plexus combined with the intermediate capillary plexus (SVP + ICP), and the deep capillary plexus (DCP) were analyzed using the AngioTool software to calculate vessel density and other vascular parameters. Results: A reduction in vessel density was seen over time in both the SVP + ICP and DCP in all mutant dogs but was more pronounced in the DCP. Scans were subclassified based on outer nuclear layer (ONL) thinning compared to age-matched normal controls. When ONL loss was 0% to 50%, vessel density in the DCP was significantly lower than in age-matched controls. In all cases, when ONL loss exceeded 87.5%, vessel density in the SVP + ICP was significantly reduced as well. In the acute light-induced rod degeneration model, the vascular regression changes were observed mainly in the DCP. Conclusions: Vessel density reduction in dogs undergoing retinal degeneration is first detected by OCTA in the DCP, and only at later stages in the SVP + ICP.

Systemic immunosuppression promotes survival and integration of subretinally implanted human ESC-derived photoreceptor precursors in dogs.

Regenerative therapies aimed at replacing photoreceptors are a promising approach for the treatment of otherwise incurable causes of blindness. However, such therapies still face significant hurdles, including the need to improve subretinal delivery and long-term survival rate of transplanted cells, and promote sufficient integration into the host retina. Here, we successfully delivered in vitro-derived human photoreceptor precursor cells (PRPCs; also known as immature photoreceptors) to the subretinal space of seven normal and three rcd1/PDE6B mutant dogs with advanced inherited retinal degeneration. Notably, while these xenografts were rejected in dogs that were not immunosuppressed, transplants in most dogs receiving systemic immunosuppression survived up to 3-5 months postinjection. Moreover, differentiation of donor PRPCs into photoreceptors with synaptic pedicle-like structures that established contact with second-order neurons was enhanced in rcd1/PDE6B mutant dogs. Together, our findings set the stage for evaluating functional vision restoration following photoreceptor replacement in canine models of inherited retinal degeneration.

Targeting ON-bipolar cells by AAV gene therapy stably reverses LRIT3-congenital stationary night blindness.

SignificanceCanine models of inherited retinal diseases have helped advance adeno-associated virus (AAV)-based gene therapies targeting specific cells in the outer retina for treating blinding diseases in patients. However, therapeutic targeting of diseases such as congenital stationary night blindness (CSNB) that exhibit defects in ON-bipolar cells (ON-BCs) of the midretina remains underdeveloped. Using a leucine-rich repeat, immunoglobulin-like and transmembrane domain 3 (LRIT3) mutant canine model of CSNB exhibiting ON-BC dysfunction, we tested the ability of cell-specific AAV capsids and promotors to specifically target ON-BCs for gene delivery. Subretinal injection of one vector demonstrated safety and efficacy with robust and stable rescue of electroretinography signals and night vision up to 1 y, paving the way for clinical trials in patients.

Retinal structural and microvascular abnormalities in retinal dysplasia imaged by OCT and OCT angiography.

OBJECTIVE: To describe the in vivo structural characteristics of multifocal and geographic retinal dysplasia visualized with advanced retinal imaging including confocal scanning laser ophthalmoscopy (cSLO), optical coherence tomography (OCT), en face OCT, and the novel vascular imaging technique OCT angiography (OCTA). DOGS STUDIED AND PROCEDURES: Two dogs were diagnosed with unilateral multifocal or geographic retinal dysplasia and underwent advanced retinal imaging under general anesthesia at the Retinal Disease Studies Facility of the University of Pennsylvania. RESULTS: In both cases, the morphological pattern of the lesions was similar including outer retinal folds that invaginated and formed tubular retinal rosettes, surrounding a central inner retinal thickening (multifocal) or plaque (geographic). The two dogs had multiple vascular anomalies in the lesions such as increased tortuosity, abnormal change of vessel diameter including aneurysms and capillary network disruption. We also identified increased autofluorescence by AF cSLO with short wavelength light source (488 nm and barrier filter at 500 nm), and several areas of photoreceptor loss associated with the lesions. CONCLUSION: The use of OCTA allowed the identification of microvascular abnormalities associated with multifocal and geographic retinal dysplasia in two dogs. To our knowledge, this is the first report where the dye-free OCTA technique is used to study vascular lesions in canine retinas.

Characterization of the Canine Retinal Vasculature With Optical Coherence Tomography Angiography: Comparisons With Histology and Fluorescein Angiography.

Purpose: To present a methodology for quantification of the canine retinal vasculature imaged by optical coherence tomography angiography (OCTA) and validate this approach by comparison with fluorescein angiography (FA) and confocal imaging of retinal wholemounts labelled by immunohistochemistry (IHC). Methods: Six normal adult dogs underwent retinal OCTA imaging in both eyes. The images extracted from the different microvascular plexuses at eight retinal locations spanning the central and mid-peripheral fundus were analyzed using the AngioTool software. FA was performed in one eye and was compared to the OCTA images. Six eyes from three dogs were processed by IHC to examine the retinal vasculature. Results: A total of four retinal plexuses were identified by OCTA in the canine retina, and their density and topographical pattern varied with eccentricity. OCTA offered improved resolution over FA with the advantage of allowing imaging of the individual plexuses. Detection by OCTA of small vessels within the deep capillary plexus was possible and approached the level of resolution achieved with ex vivo imaging of the retinal vasculature by confocal microscopy/IHC. The plexuses herein described are analogous to human retinal vasculature. Conclusion: OCTA can be used to image and quantify non-invasively the vascular retinal networks of the canine retina. We provide normative data in eight different retinal locations that can be imaged non-invasively with this technology. This could support analysis of retinal vascular changes associated with disease and following therapeutic intervention.

Comparison of the use of a standard cytology brush versus a mini cytology brush to obtain conjunctival samples for cytologic examination in healthy dogs.

OBJECTIVE: To compare the quality of conjunctival samples for cytologic examination obtained with 2 conjunctival exfoliative brushes, a mini cytology brush (MCB) and a standard cytology brush (SCB), in healthy dogs. ANIMALS: 20 client-owned dogs that were free of ocular disease. PROCEDURES: A prospective single-center randomized trial was performed. For each dog, conjunctival samples of the right eye were obtained with the 2 brushes (ie, SCB or MCB) at 2 time points that were 5 to 11 days apart. The left eye was used as a control. Cytologic quality of conjunctival samples was scored on the basis of cellularity, clearness of background, uniformity of distribution of cells on the cytology slide, artifacts, cellular overlapping, cell preservation, presence of mucus on the cytology slide, and number of RBCs. RESULTS: On cytologic evaluation, conjunctival samples collected with an SCB scored significantly better in terms of higher cellularity, less background debris, and more uniformity in the distribution of cells, compared with conjunctival samples collected with an MCB. Conjunctival samples collected with an MCB scored significantly better in terms of less cellular overlapping and less mucus in the background, compared with conjunctival samples collected with an SCB. CONCLUSIONS AND CLINICAL RELEVANCE: Overall conjunctival samples obtained with an SCB for cytologic evaluation had better diagnostic quality, compared with conjunctival samples obtained with an MCB. Use of an MCB, however, was advantageous to access localized conjunctival areas as well as collect conjunctival samples from patients with small palpebral fissures.

Gene therapy reforms photoreceptor structure and restores vision in NPHP5-associated Leber congenital amaurosis.

The inherited childhood blindness caused by mutations in NPHP5, a form of Leber congenital amaurosis, results in abnormal development, dysfunction, and degeneration of photoreceptors. A naturally occurring NPHP5 mutation in dogs leads to a phenotype that very nearly duplicates the human retinopathy in terms of the photoreceptors involved, spatial distribution of degeneration, and the natural history of vision loss. We show that adeno-associated virus (AAV)-mediated NPHP5 gene augmentation of mutant canine retinas at the time of active degeneration and peak cell death stably restores photoreceptor structure, function, and vision with either the canine or human NPHP5 transgenes. Mutant cone photoreceptors, which failed to form outer segments during development, reform this structure after treatment. Degenerating rod photoreceptor outer segments are stabilized and develop normal structure. This process begins within 8 weeks after treatment and remains stable throughout the 6-month posttreatment period. In both photoreceptor cell classes mislocalization of rod and cone opsins is minimized or reversed. Retinal function and functional vision are restored. Efficacy of gene therapy in this large animal ciliopathy model of Leber congenital amaurosis provides a path for translation to human treatment.