Method Article
Oxygen-induced retinopathy (OIR) can be used to model ischemic retinal diseases such as retinopathy of prematurity and proliferative diabetic retinopathy and to serve as a model for proof-of-concept studies in evaluating antiangiogenic drugs for neovascular diseases. OIR induces robust and reproducible neovascularization in the retina that can be quantified.
One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR model induction and its readouts in both mice and rats. Retinal neovascularization is induced in OIR by exposing rodent pups either to hyperoxia (mice) or alternating levels of hyperoxia and hypoxia (rats). The primary readouts of these models are the size of neovascular (NV) and avascular (AVA) areas in the retina. This preclinical in vivo model can be used to evaluate the efficacy of potential anti-angiogenic drugs or to address the role of specific genes in the retinal angiogenesis by using genetically manipulated animals. The model has some strain and vendor specific variation in the OIR induction which should be taken into consideration when designing the experiments.
Reliable and reproducible experimental models are needed to study the pathology behind angiogenic eye diseases and to develop novel therapeutics to these devastating diseases. Pathological angiogenesis is the hallmark for wet age-related macular degeneration (AMD) and for many ischemic retinal diseases among them retinopathy of prematurity (ROP), proliferative diabetic retinopathy (PDR) and retinal vein occlusion (RVO)1,2,3,4. Human and rodent retinas follow a similar pattern of development, as both human and rodent retina are among the last....
Access restricted. Please log in or start a trial to view this content.
The protocol described here has been approved by the National Animal Ethics Committee of Finland (protocol number ESAVI/9520/2020 and ESAVI/6421/04.10.07/2017).
1. Experimental animals and mouse OIR model induction
NOTE: Use time-mated animals, e.g., commonly used C57BL/6J mice, to get pups born on the same day. Use fostering dams, e.g., 129 strain (129S1/SvImJ or 129S3/SvIM) lactating dams, to nurse the pups during and after the induction of hyperoxia. Alternatively, make sure that there are extra lactating dams available in case the nursing dams need to be replaced due to exhaustion. Restrict the litter size to 6....
Access restricted. Please log in or start a trial to view this content.
The main outcome of the model is the vascular phenotype: the size of AVAs and the amount of NV. In the mouse OIR model, the vaso-obliteration occurs in the central retina (Figure 2A), while in the rat model it develops in the periphery, i.e., similar to human ROP22 (Figure 3A). This is because the superficial vascular plexus has already developed when mice are exposed to hyperoxia, whereas in the rat model the retina is avascular at the t.......
Access restricted. Please log in or start a trial to view this content.
The severity of disease phenotype is dependent on both the strain and even vendor in both mouse and rat OIR models23. This suggests that there is a wide genotypic variability in the pathology development. In general, pigmented rodents develop more severe phenotype than the albino ones. For example, the retinal vasculature of albino BALB/c revascularizes rapidly after hyperoxia and does not develop NV at all24. Similarly, in rats, pigmented Brown Norway rats show more severe.......
Access restricted. Please log in or start a trial to view this content.
The authors Maria Vähätupa, PhD, Niina Jääskeläinen, Marc Cerrada-Gimenez, PhD, and Rubina Thapa are employees of Experimentica Ltd.
The author Giedrius Kalesnykas, PhD, is an employee (President and Chief Executive Officer) and shareholder of Experimentica Ltd. that offers contract research services employing preclinical OIR models used in this Article.
Tero Järvinen, M.D., PhD, and Hannele Uusitalo-Järvinen, M.D., PhD, have nothing to disclose.
We thank Marianne Karlsberg, Anne Mari Haapaniemi, Päivi Partanen and Anne Kankkunen for excellent technical support. This work was funded by the Academy of Finland, Päivikki and Sakari Sohlberg Foundation, Tampere Tuberculosis Foundation, Finnish Medical Foundation, Pirkanmaa Hospital District Research Foundation and the Tampere University Hospital Research Fund.
....Access restricted. Please log in or start a trial to view this content.
| Name | Company | Catalog Number | Comments |
|---|---|---|---|
| 33 gauge, Small Hub RN Needle | Hamilton Company | 7803-05, 10mm, 25°, PS4 | For intravitreal injection |
| Adobe Photoshop | Adobe Inc. | For image analysis | |
| Air pump air100 | Eheim GmbH & Co. KG. | 143207 | For inhalation anaesthesia |
| Anaesthesia unit 410 AP | Univentor Ltd. | 2360309 | For inhalation anaesthesia |
| AnalaR NORMAPUR Soda lime | VWR International Ltd | 22666.362 | For CO2 control during model induction |
| Attane Vet 1000 mg/g | VET MEDIC ANIMAL HEALTH OY | vnr 17 05 79 | For inhalation anaesthesia |
| Brush | For preparation of flat mounts | ||
| Carbon dioxide gas | For sacrifice | ||
| Celeris D430 ERG system | Diagnosys LLC | 121 | For in vivo ERG |
| Cell culture dishes | Greiner Bio-One International GmbH | 664 160 | For preparation of flat mounts |
| Cepetor Vet 1 mg/mL | VET MEDIC ANIMAL HEALTH OY | vnr 08 78 96 | For anaesthesia |
| Cover slips | Thermo Fisher Scientific | 15165452 | For preparation of flat mounts |
| O2 Controlled InVivo Cabinet, Aninal Filtrarion System and Dehumidifier | Coy Laboratory Products | Closed system for disease model induction, optional for semi-closed system | |
| E702 O2 sensor | BioSphenix, Ltd. | E207, 1801901 | For oxygen level measurement |
| Envisu R2200 Spectral Domain Optical Coherence Tomograph (SD-OCT) | Bioptigen, Inc. | BPN000668 | For in vivo imaging |
| Eye spears | Beaver-Visitec International, Inc. | 0008685 | For intravitreal injection and in vivo imaging |
| Flexilux 600LL Cold light source | Mikron | 11140 | For intravitreal injection or tissue collection |
| Fluorescein sodium salt | Merck KGaA | F6377-100G | For in vivo imaging |
| Gas Exhaust unit (+Double 3-way valve, mouse and rat face masks, UNOsorb filter) | UNO Roestvaststaal BV | GEX 17015249 | For inhalation anaesthesia |
| Glass syringe, Model 65 RN | Hamilton Company | 7633-01 | For intravitreal injection |
| HRA2 Retina angiograph (FA) | Heidelberg Engineering GmbH | Spec-KT-05488 | For in vivo imaging |
| Isolectin GS-IB4, Alexa Fluor 488 Conjugate | Thermo Fisher Scientific | I21411 | For labeling retinal vasculature on flat mounts |
| Ketaminol Vet 50 mg/mL | Intervet International B.V. | vnr 51 14 85 | For anaesthesia |
| Medicinal Oxygen gas | For disease model induction | ||
| Mice C57BL/6JRj | Janvier Labs | Also other strains possible | |
| Microscope slides | Thermo Fisher Scientific | J1800AMNZ | For preparation of flat mounts |
| Minims Povidone Iodine 5% (unit) | Bausch & Lomb U.K Limited | vnr 24 11 304 | For intravitreal injection |
| Nitrogen gas | For disease model induction (rat) | ||
| Oftan Chlora 10 mg/g | Santen Pharmaceutical Co., Ltd. | vnr 55 01 11 | For intravitreal injection |
| Oftan Metaoksedrin 100 mg/ml | Santen Pharmaceutical Co., Ltd. | vnr 55 03 43 | For in vivo ERG |
| Oftan Obucain 4 mg/ml | Santen Pharmaceutical Co., Ltd. | vnr 55 03 50 | For intravitreal injection |
| Oftan Tropicamid 5 mg/ml | Santen Pharmaceutical Co., Ltd. | vnr 04 12 36 | For in vivo imaging |
| ProOx Model 110 O2 controller and animal chamber | BioSphenix, Ltd. | 803 | For disease model induction, semi-closed system, optional for closed system |
| ProOx Model P360 O2 controller and animal chamber | BioSphenix, Ltd. | 538 | For disease model induction, semi-closed system, optional for closed system |
| Rats CD(SD) | Charles River Laboratories | Also other strains possible | |
| Revertor 5 mg/mL | VET MEDIC ANIMAL HEALTH OY | vnr 13 04 97 | For anaesthesia reversal |
| Silica gel | For humidity control during model induction | ||
| Systane Ultra 10ml | Alcon | Tamro 2050250 | For hydration of the eye |
| Systane Ultra unit 0.7ml | Alcon | Tamro 2064871 | For hydration of the eye |
| Transfer pipette | Thermo Fisher Scientific | 1343-9108 | For preparation of flat mounts |
| VENTI-Line VL 180 PRIME Drying oven | VWR | VL180S 170301 | For drying silica gel |
| VisiScope SZT350 Stereomicroscope | VWR | 481067 | For intravitreal injection or tissue collection |
Access restricted. Please log in or start a trial to view this content.
Request permission to reuse the text or figures of this JoVE article
Request Permission