In-vivo and ex-vivo characterization of laser-induced choroidal neovascularization variability in mice

Abstract

Retinal argon laser coagulation is an established procedure for induction of choroidal neovascularization (CNV) in rodents. This study aimed to evaluate the in-vivo and ex-vivo morphology and variability of laser-induced CNV spots over time. Female C57/Bl/6 mice, 3-6months of age, were treated with five spots of retinal argon laser coagulation per eye (150mW, 100ms, 50μm). In-vivo fluorescein angiography (FA) and standard high-resolution spectral-domain optical coherence tomography (SD-OCT) were performed on day (d) 0, d1, d4, d7, d14 and d21. Ex-vivo histology, CD31 immunostaining, flatmount and confocal microscopy were also conducted. CNV size in the retinal and choroidal focus, CNV morphology, central retinal thickness (CRT) and FA CNV activity grading were assessed in-vivo at all times and compared to the ex-vivo assessments. SD-OCT revealed sub-retinal and intra-retinal fluid, and permitted evaluation of longitudinal morphologic changes of the induced CNV. Laser spot area in FA and CRT in SD-OCT did not differ in longitudinal evaluation. CNV could not be consistently outlined on SD-OCT images, and CNV volume as assessed on SD-OCT did not change over time. Significant CNV activity changes were only found in FA CNV activity grading, peaking on d4 and decreasing by d7. Non-invasive SD-OCT provides additional morphological information on laser-induced CNV. However, reliable evaluation of CNV requires FA. Spontaneous regression of CNV activity within 14days after induction has to be taken into account when utilizing this model for testing the efficacies of potential future treatments.