Standardizing the Quantification of Neovascularization and Vessel Loss in the Mouse Model of Oxygen-Induced Retinopathy

Abstract

Purpose: : Angiogenic research relies heavily upon the utilization of rodent models to gain insight into developmental and pathological angiogensis. The mouse model of oxygen-induced retinopathy (OIR) is in the forefront of in vivo research. However, assessing neovascularization (NV) and vessel loss (VO) in the OIR model has been time-consuming, variable, and difficult to standardize. Here we describe a computer-aided quantification method that aims to improve efficiency and reproducibility of assessing NV in the OIR model.

Methods: : Vessel loss and subsequent pathological neovascularization were induced by exposing nursing mouse mothers and their pups to 75% oxygen from P7 to P12. Mice were then returned to room air until sacrifice on P17. VO was quantified in Adobe Photoshop by comparing the VO area to total retinal area. For quantification of retinal NV, a semi-automated, computer-aided quantification method, SWIFT_NV, was developed and compared to the current manual quantification technique on retinal flatmounts.

Results: : Comparing measurements from SWIFT_NV vs. manual quantifications revealed an R2 value of 0.9372 and comparison of the mean values resulted in almost exact congruence (7.76±0.40 vs. 7.51±0.41% NV per total retina). Intra-individual reproducibility of SWIFT_NV over time was assessed by repeating quantifications by the same investigator on the same data set 3 months apart, revealing strong correlation between original and repeated measures (R2=0.937). Inter-individual reproducibility was analyzed through independent quantifications by two investigators, revealing high correlation between users (R2=0.942).

Conclusions: : Computer-aided quantification of retinal neovascularization using SWIFT_NV can provide time-efficient and accurate results that correlate well with manual NV measurements. Furthermore, given the high reproducibility between users, SWIFT_NV can be valuable to further standardize NV quantification between individual investigators and laboratories using the OIR mouse model.

Keywords: retinopathy of prematurity • retinal neovascularization • imaging/image analysis: non-clinical