Jacklyn H. Salmon

Treatment of Acute Posterior Uveitis in a Porcine Model by Injection of Triamcinolone Acetonide Into the Suprachoroidal Space Using Microneedles

PURPOSE To evaluate the effect of triamcinolone acetonide (TA) administered into the suprachoroidal space (SCS) using a microneedle and compare it with intravitreal (IVT) TA injections in a porcine model of acute posterior segment inflammation. MATERIALS An IVT injection of balanced salt solution (BSS) or lipopolysaccharide (LPS) was followed 24 hours later with an injection of 0.2 mg or 2.0 mg of TA into the SCS or IVT. The SCS was accessed using microneedles in a minimally invasive procedure. Ocular inflammatory scores and IOP measurements were collected daily, whereas electroretinography, optical coherence tomography, and wide-field ocular fundus photography was performed on -1, 0, and 3 days after treatment. Aqueous and vitreous humor cell counts and protein levels and histopathology were also compared. RESULTS Delivery of TA to the SCS using microneedles was simple, effective, and not associated with adverse effects or toxicity. SCS injection of low (0.2 mg) and high doses (2.0 mg) of TA was as effective in reducing acute inflammation in the ocular posterior segment as high-dose IVT injection. Low-dose SCS TA was also effective in reducing inflammation; however, low-dose IVT TA was not. CONCLUSIONS Results from this study suggest that 0.2 mg and 2.0 mg of SCS TA was as effective in reducing inflammation as 2.0 mg IVT TA injection in a model of acute posterior segment inflammation. There were no adverse effects, increased IOP, or evidence of procedural or drug toxicity following injection of TA into the SCS in porcine eyes.

Role of bacteria in the pathogenesis of recurrent uveitis in horses from the southeastern United States

OBJECTIVE To determine the role of intraocular bacteria in the pathogenesis of equine recurrent uveitis (ERU) in horses from the southeastern United States by evaluating affected eyes of horses with ERU for bacterial DNA and intraocular production of antibodies against Leptospira spp. SAMPLE POPULATION Aqueous humor, vitreous humor, and serum samples of 24 clinically normal horses, 52 horses with ERU, and 17 horses with ocular inflammation not associated with ERU (ie, non-ERU inflammation). PROCEDURES Ribosomal RNA quantitative PCR (real-time PCR) assay was used to detect bacterial DNA in aqueous humor and vitreous humor from clinically normal horses (n = 12) and horses with chronic (> 3-month) ERU (28). Aqueous humor and serum were also evaluated for anti-Leptospira antibody titers from clinically normal horses (n = 12), horses with non-ERU inflammation (17), and horses with confirmed chronic ERU (24). RESULTS Bacterial DNA was not detected in aqueous humor or vitreous humor of horses with ERU or clinically normal horses. No significant difference was found in titers of anti-Leptospira antibodies in serum or aqueous humor among these 3 groups. Only 2 horses, 1 horse with ERU and 1 horse with non-ERU inflammation, had definitive intraocular production of antibodies against Leptospira organisms. CONCLUSIONS AND CLINICAL RELEVANCE In horses from the southeastern United States, Leptospira organisms may have helped initiate ERU in some, but the continued presence of the organisms did not play a direct role in the pathogenesis of this recurrent disease.

Ocular toxicity and distribution of subconjunctival and intravitreal rapamycin in horses

In vitro photosensitivity of rapamycin (RAPA) and ocular toxicity and distribution of intravitreal and subconjunctival RAPA was evaluated in normal horses. RAPA (2.5 mg, 5 mg, and 10 mg) was placed in 10 mL of PBS and maintained in a water bath at 37 degrees C, kept in the dark or subjected to room light, and sampled for up to 3 months for RAPA levels. Six normal adult horses received either 5 mg (n = 2) or 10 mg (n = 2) of RAPA intravitreally or 10 mg (n = 2) subconjunctivally. Ophthalmic exams and electroretinography (ERG) were performed prior to injection and on days 1, 7, 14, and 21 post-injection. Eyes were enucleated and samples were collected for RAPA concentrations and histopathology. No difference in light vs. dark RAPA concentrations was observed, suggesting a lack of RAPA phototoxicity. No evidence of ocular toxicity was noted on ophthalmic examination or histopathology. RAPA was not detected intraocularly 7 days post-injection in eyes receiving subconjunctival RAPA, but was detected in the vitreous at 21 days post-injection. Drug could be detected in both the aqueous and vitreous humor after intravitreal injection. Further study is needed to determine the efficacy of intravitreal RAPA.

Evaluation of 30- and 25-diopter intraocular lens implants in equine eyes after surgical extraction of the lens

OBJECTIVE To determine appropriate intraocular lens (IOL) implant strength to approximate emmetropia in horses. SAMPLE POPULATION 16 enucleated globes and 4 adult horses. PROCEDURES Lens diameter of 10 enucleated globes was measured. Results were used to determine the appropriate-sized IOL implant for insertion in 6 enucleated globes and 4 eyes of adult horses. Streak retinoscopy and ocular ultrasonography were performed before and after insertion of 30-diopter (D) IOL implants (enucleated globes) and insertion of 25-D IOL implants (adult horses). RESULTS In enucleated globes, mean +/- SD lens diameter was 20.14 +/- 0.75 mm. Preoperative and postoperative refractive state of enucleated globes with 30-D IOL implants was -0.46 +/- 1.03 D and -2.47 +/- 1.03 D, respectively; preoperative and postoperative difference in refraction was 2.96 +/- 0.84 D. Preoperative anterior chamber (AC) depth, crystalline lens thickness (CLT), and axial globe length (AxL) were 712 +/- 0.82 mm, 11.32 +/- 0.81 mm, and 40.52 +/- 1.26 mm, respectively; postoperative AC depth was 10.76 +/- 1.16 mm. Mean ratio of preoperative to postoperative AC depth was 0.68. In eyes receiving 25-D IOL implants, preoperative and postoperative mean refractive error was 0.08 +/- 0.68 D and -3.94 +/- 1.88 D, respectively. Preoperative AC depth, CLT, and AxL were 6.36 +/- 0.22 mm, 10.92 +/- 1.92 mm, and 38.64 +/- 2.59 mm, respectively. Postoperative AC depth was 8.99 +/- 1.68 mm. Mean ratio of preoperative to postoperative AC depth was 0.73. CONCLUSIONS AND CLINICAL RELEVANCE Insertion of 30-D (enucleated globes) and 25-D IOL implants (adult horses) resulted in overcorrection of refractive error.

Ocular distribution and toxicity of intravitreal injection of triamcinolone acetonide in normal equine eyes

OBJECTIVE To determine ocular distribution and toxicity of a single injection of intravitreal triamcinolone acetonide (TA) in normal horses. ANIMALS STUDIED Six adult horses, donated to North Carolina State University. PROCEDURES Six horses were injected intravitreally with either 10, 20, or 40 mg (n = 2 each) of TA. The opposite eye of each horse was injected with balanced salt solution (BSS). Ocular toxicity was assessed by biomicroscopy, tonometry, indirect ophthalmoscopy, and electroretinogram. Aqueous humor (AH), vitreous humor (VH), and plasma samples were collected. Horses were euthanized 7 or 21 days after injection and eyes enucleated for histopathology. TA concentrations in AH, VH, and plasma were measured by HPLC. RESULTS Three control eyes and one TA eye developed inflammation after injection or collection of AH. Positive bacterial cultures (Corynebacterium spp., Staphylococcus spp., and Streptococcus spp.) were obtained from three of these eyes. Other than transient corneal edema in TA injected eyes, which resolved by 7 days after injection, no other changes were observed. TA crystals were visible within the vitreous body. No evidence of TA toxic effect was noted on histopathology. TA was detected in all AH and VH samples from treated eyes following injection. Drug was not detected in the plasma. CONCLUSIONS There was no evidence of overt toxicity from intravitreal TA in normal horses and a single intravitreal injection resulted in TA ocular levels for 21 days. However, the risk for bacterial infections with intravitreal injection or anterior chamber aspirations in horses is high. Use of topical and systemic antibiotics after injection is recommended.

Immunohistochemical and immunopathologic characterization of superficial stromal immune-mediated keratitis in horses

OBJECTIVE To describe the immunopathologic characteristics of superficial stromal immune-mediated keratitis (IMMK) immunopathologically by characterizing cellular infiltrate in affected corneas of horses. ANIMALS 10 client-owned horses with IMMK. PROCEDURES Immunohistochemical staining was performed on keratectomy samples with equine antibodies against the T-cell marker CD3 and B-cell marker CD79a (10 eyes) and the T-helper cytotoxic marker CD4 and T-cell cytotoxic marker CD8 (6 eyes). Percentage of positively stained cells was scored on a scale from 0 (no cells stained) to 4 (> 75% of cells stained). Equine IgG, IgM, and IgA antibodies were used to detect corneal immunoglobulin via direct immunofluorescence (10 eyes). Serum and aqueous humor (AH) samples from 3 horses with IMMK were used to detect circulating and intraocular IgG against corneal antigens via indirect immunofluorescence on unaffected equine cornea. RESULTS Percentage scores (scale, 0 to 4) of cells expressing CD3 (median, 2.35 [range, 0.2 to 3.7]; mean ± SD, 2.36 ± 1.08) were significantly greater than scores of cells expressing CD79a (median, 0.55 [range, 0 to 1.5]; mean, 0.69 ± 0.72). All samples stained positively for CD4- and CD8-expressing cells, with no significant difference in scoring. All samples stained positively for IgG, IgM, and IgA. No serum or AH samples collected from horses with IMMK reacted with unaffected equine cornea. CONCLUSIONS AND CLINICAL RELEVANCE Pathogenesis of superficial stromal IMMK included cell-mediated inflammation governed by both cytotoxic and helper T cells. Local immunoglobulins were present in affected corneas; however, corneal-binding immunoglobulins were not detected in the serum or AH from horses with IMMK.

Treatment of immune‐mediated keratitis in horses with episcleral silicone matrix cyclosporine delivery devices

PURPOSE To describe the use of episcleral silicone matrix cyclosporine (ESMC) drug delivery devices in horses with immune-mediated keratitis (IMMK) with evaluation of tolerability and efficacy in long-term control of inflammation. METHODS Retrospective study. ESMC implants (1.2 cm length, 30% wt/wt cyclosporine (CsA) in silicone; with approximately 2 μg/day steady-state release for at least 400 days) were used. RESULTS Nineteen horses (20 eyes) received two or more ESMC implants for superficial stromal (n = 9), midstromal (n = 3), or endothelial (n = 5) IMMK. Three additional horses received two or more ESMC implants for pigmentary keratouveitis (PK). Nine eyes of eight horses with superficial and five eyes of five horses with endothelial IMMK were well controlled after placement of ESMC implants (mean follow-up 176.8 and 207.2 days, respectively). Horses with midstromal IMMK and PK were not controlled with ESMC implants alone, but instead required frequent use of other medications or surgery to control the disease. The mean duration of disease prior to ESMC implantation of horses with midstromal IMMK was 495 ± 203.9 days, compared with 121.6 ± 92.7 days with superficial IMMK. ESMC implants were well tolerated by all horses without documented loss of the device. CONCLUSIONS Results from this preliminary retrospective study suggest that the ESMC implants were well tolerated and associated with treatment success with superficial and endothelial IMMK, especially if placed early in the disease process. Further study is needed to determine the duration of efficacy, number of implants required, and better therapies for chronic midstromal IMMK and pigmentary keratouveitis.

Sustained-Release Celecoxib from Incubated Acrylic Intraocular Lenses Suppresses Lens Epithelial Cell Growth in an Ex Vivo Model of Posterior Capsule Opacity

PURPOSE To determine whether celecoxib (CXB) can be released from incubated intraocular lenses (IOLs) sufficiently to inhibit lens epithelial cell (LEC) growth in an ex vivo model of posterior capsule opacification (PCO). MATERIALS LEC growth was evaluated for 14 days in canine lens capsules (LCs) that had been exposed to media containing 20 μM CXB for 1-5 days. After the incubation of hydrophilic and hydrophobic IOLs in CXB solution, the determination of the in vitro release of CXB from the IOLs was performed for up to 28 days. The incubated and nonincubated IOLs were evaluated in the ex vivo model of PCO, and the rate of LEC growth was evaluated over 28 days. RESULTS The treatment of LCs with 20 μM CXB for 4 and 5 days completely inhibited LEC growth. LEC repopulation did not occur after the removal of CXB. IOLs incubated in CXB for 24 h resulted in a sustained release of CXB in vitro at levels theoretically sufficient to inhibit PCO. LCs in the ex vivo model of PCO treated with acrylic IOLs incubated in CXB had significantly suppressed LEC ingrowth compared with untreated and IOL-only LCs. CONCLUSIONS A 4-day treatment of LCs with a concentration of 20 μM CXB may effectively prevent PCO. IOLs incubated in CXB for 24 h resulted in a sustained release of CXB in vitro at levels sufficient to inhibit LEC growth in the ex vivo model of PCO. Further studies are needed to determine whether CXB-incubated IOLs can effectively prevent the development of PCO in vivo.

Aqueous humor and plasma concentrations of a compounded 0.2% solution of terbinafine following topical ocular administration to normal equine eyes

OBJECTIVE To determine the transcorneal penetration and systemic absorption of a compounded 0.2% terbinafine solution following repeated topical administration to normal equine eyes. Sample population  Six healthy adult horses with normal ocular examinations. PROCEDURES One eye of each horse received 0.2 mL of a compounded 0.2% terbinafine solution every 4 h for seven doses. During the 1 h following administration of the final dose, multiple peripheral blood samples were obtained, and a single aqueous humor (AH) sample was collected at the end of the hour. AH and plasma concentrations of terbinafine were determined using high pressure liquid chromatography (HPLC). Stability of the formulation was assessed with HPLC analysis over a 14-day time period. RESULTS Terbinafine was not detected in the AH or plasma of any horse at any time point. No signs of ocular irritation or systemic toxicity were noted in any horse at any time point. The solution was stable over 14 days. CONCLUSION Topical ocular administration of compounded 0.2% terbinafine solution does not result in detectable AH or plasma levels following administration to normal equine eyes, suggesting its use for deep corneal or intraocular fungal infections in equine ophthalmology may be limited.

Effect of Choroidal Perfusion on Ocular Tissue Distribution After Intravitreal or Suprachoroidal Injection in an Arterially Perfused Ex Vivo Pig Eye Model

PURPOSE To compare tissue distribution of dye-drug surrogates after intravitreal (IVT) and suprachoroidal (SCS) delivery to determine the influence of drug lipophilicity and choroidal circulation. METHODS Thirty-two pig eyes were collected immediately after euthanasia. Sixteen eyes were perfused for 30 min through one long posterior ciliary artery with nondye containing nutrient media. An IVT or SCS injection was performed with either a 100 μL balanced salt solution (BSS, n=8), 1% sodium fluorescein (NaF, n=12) or 0.12% lipophilic carbocyanine dye (DiI, n=12). Globes were maintained at 37°C for 15 min, and then snap-frozen and dissected. Aqueous extraction and measurement of NaF or DiI concentration was performed using spectrophotometry and spectrofluorometry, respectively. RESULTS After SCS delivery of NaF scleral, iris-ciliary body, choroidal and vitreous dye levels were higher in nonperfused eyes compared to perfused eyes. After DiI SCS or IVT delivery, no significant differences were found in dye tissue concentrations in perfused eyes compared to nonperfused eyes. Following perfusion, a better and even drug distribution was found in the retinal pigmented epithelium (RPE)-choroid following IVT and SCS delivery of the hydrophilic drug and after IVT injection of the lipophilic drug compared to nonperfused eyes. CONCLUSIONS Choroidal circulation reduces the tissue drug concentration of the hydrophilic drug suggesting an early clearance mechanism after SCS delivery. SCS injections of lipid and hydrophilic drugs allowed direct drug delivery to the retina and RPE-choroid with limited exposition to the anterior segment.