In Vivo Vascular Injury Readouts in Mouse Retina to Promote Reproducibility

Summary

Here, we present three data analysis protocols for fluorescein angiography (FA) and optical coherence tomography (OCT) images in the study of Retinal Vein Occlusion (RVO).

Abstract

Advancements in ophthalmic imaging tools offer an unprecedented level of access to researchers working with animal models of neurovascular injury. To properly leverage this greater translatability, there is a need to devise reproducible methods of drawing quantitative data from these images. Optical coherence tomography (OCT) imaging can resolve retinal histology at micrometer resolution and reveal functional differences in vascular blood flow. Here, we delineate noninvasive vascular readouts that we use to characterize pathological damage post vascular insult in an optimized mouse model of retinal vein occlusion (RVO). These readouts include live imaging analysis of retinal morphology, disorganization of retinal inner layers (DRIL) measure of capillary ischemia, and fluorescein angiography measures of retinal edema and vascular density. These techniques correspond directly to those used to examine patients with retinal disease in the clinic. Standardizing these methods enables direct and reproducible comparison of animal models with clinical phenotypes of ophthalmic disease, increasing the translational power of vascular injury models.

Introduction

Neurovascular disease is a major healthcare problem responsible for ischemic strokes, a leading cause of mortality and morbidity, and retinal vascular diseases that lead to vision loss^1,2. To model neurovascular disease, we employ a mouse model of retinal vein occlusion (RVO). This model is noninvasive and utilizes similar in vivo imaging techniques to those used to examine people with retinal vascular disease in a clinical setting. The use of this model thus increases the translational potential of studies utilizing this model. As with all mouse models, it is critical to maximize reproducibility of t....

Protocol

This protocol follows the Association for Research in Vision and Ophthalmology (ARVO) statement for the use of animals in ophthalmic and vision research. Rodent experiments were approved and monitored by the Institutional Animal Care and Use Committee (IACUC) of Columbia University.

NOTE: Imaging was done on 2 month old C57BL/6J male mice that weighed approximately 23 g.

1. Preparation of reagents for retinal imaging

1. Preparation of injec.......

Discussion

Noninvasive rodent retinal imaging presents an avenue to study pathology and develop interventions. Previous studies have developed and optimized a mouse model of RVO, limiting variability and allowing for reliable translation of common clinical pathologies in the murine retina^5,7,13. Developments in ophthalmic imaging technology further allow for the use of clinical in vivo imaging techniques such as FA and OCT in expe.......

Materials

Name	Company	Catalog Number	Comments
AK-Fluor 10%	Akorn	NDC: 17478-253-10	light-sensitive
Carprofen	Rimadyl	NADA #141-199	keep at 4 °C
GenTeal	Alcon	00658 06401
Image J	NIH
InSight 2D	Phoenix Technology Group		OCT analysis software
Ketamine Hydrochloride	Henry Schein	NDC: 11695-0702-1
Phenylephrine	Akorn	NDCL174478-201-15
Phoenix Micron IV	Phoenix Technology Group		Retinal imaging microscope
Phoenix Micron Meridian Module	Phoenix Technology Group		Laser photocoagulator software
Phoenix Micron Optical Coherence Tomography Module	Phoenix Technology Group		OCT imaging software
Phoenix Micron StreamPix Module	Phoenix Technology Group		Fundus imaging and acquisition targeting
Photoshop	Adobe
Refresh	Allergan	94170
Tropicamide	Akorn	NDC: 174478-102-12
Xylazine	Akorn	NDCL 59399-110-20