Dexamethasone Intravitreal Implant Is Active at the Molecular Level Eight Weeks after Implantation in Experimental Central Retinal Vein Occlusion

Abstract

Central retinal vein occlusion (CRVO) is a visually disabling condition resulting from a thrombus in the major outflow vessel of the eye. The inflammatory response in CRVO is effectively treated with a dexamethasone (DEX) intravitreal implant. Uncovering the proteome changes following DEX implant intervention in CRVO may identify key proteins that mediate the beneficial effects of DEX. In six Göttingen minipigs, CRVO was induced in both eyes with an argon laser using a well-established experimental model. The right eyes were treated with a DEX intravitreal implant (Ozurdex, Allergan), while the left control eyes received a sham injection. Eight weeks after DEX intervention, retinal samples were collected and analyzed with tandem mass tag-based mass spectrometry. DEX implant intervention resulted in the upregulation of peptidyl-prolyl cis–trans isomerase FKBP5 (FKBP5) and ubiquilin-4. Immunohistochemistry showed expression of FKBP5 in the nuclei in all cellular layers of the retina. Cell adhesion molecule 3, tumor necrosis factor receptor superfamily member 16, and trans-1,2-dihydrobenzene-1,2-diol dehydrogenase were downregulated following DEX intervention. The upregulation of the corticosteroid-sensitive protein FKBP5 suggests that the implant remained active at the molecular level after eight weeks of treatment. Future studies may investigate if FKBP5 regulates the efficacy and duration of the DEX implant.

Laser-Induced Porcine Model of Experimental Retinal Vein Occlusion: An Optimized Reproducible Approach

“…We reviewed animal studies of experimental RVO performed in our laboratories in a 10-year period [ 25 , 26 , 28 , 29 , 30 , 31 , 32 , 33 ]. A total of 130 eyes from 65 animals were used ( Table 1 ), which included 65 eyes with experimental BRVO from 38 animals and 43 eyes with experimental CRVO from 27 animals.…”

“…Furthermore, we find enucleation more difficult in Göttingen minipigs due to their fairly deep orbital cavities. For studies of experimental RVO, we recommend female Danish Landrace pigs of approximately 20 kg [ 28 , 34 ]. Animals were housed under a 12 h light/dark cycle.…”

“…Proteomic analysis of retinal control samples and retinal tissue exposed to RVO is effectively performed using both label-free and labeling techniques [ 26 , 28 , 29 ]. For labeling approaches, we find the 10-plex tandem mass tag (TMT) kit (Thermo Scientific, Waltham, MA, USA) useful for the study of experimental RVO.…”

“…The peptides are separated by liquid chromatography and detected in the mass spectrometer using synchronous precursor selection. The reporter ions are detected by MS 3 in the Orbitrap as described previously [ 28 , 29 , 30 , 33 ].…”

“…Raw files obtained with mass spectrometry are searched against relevant databases using MaxQuant software (Max Planck Institute of Biochemistry, Martinsried, Germany; the software can be downloaded from the following site: ). In our experience, optimal proteome coverage is achieved by searching against the UniProt Sus scrofa database as well as the UniProt Homo sapiens database [ 28 ].…”