# En-Face Optical Coherence Tomography Angiography for Longitudinal Monitoring of Retinal Injury
## 3 Results
Comparing the baseline to 1-hour lesion response using the combination of B-scan and OCTA visualization, the damage of the acute injury lesions is confined to the photoreceptor layer and has not disrupted inner retinal vasculature (Figure 8) or the choroid (Figure 9). The acute injury lesions showed no gross changes in the capillary network in the OCTA scans. The acute injury lesion and scarring in the ONL slab were apparent and did not penetrate the choroid slab. To confirm the RPE rupture and capillary non-perfusion, a repeat experimental OCTA group was set with time points 1h, 24h, and 72h. With the short time points, the choroid disruption was visible immediately and inner retina capillary non-perfusion apparent at 24h (Figure 8). The inner retina at 1 h in CNV shows displacement of layers, and at 24h the signal from the vasculature is undetectable. The acute injury samples and surrounding areas do not lose optical signal, demonstrating this is a localized disruption (Figure 9). Importantly, the integrity of the choroid slab can be immediately assessed for RPE rupture. At 1h, the acute injury lesions distort the RPE but do not penetrate through and protrude into the choroid. At 24h, the initial distortion from the localized inflammation is minimized showing choroid as a continuous layer. In L-CNV, clear rupture of the RPE and Choroid were observed at 1 h while the retinal layers are not continuous and the damage from the vaporization bubble is apparent. The CNV lesions affected all retinal layers by displacing the inner layers and disrupting the continuity of the outer retina. The en-face view of the OCTA highlighted several key features: the rupture of the choroid, the infiltration into the ONL, and the non-perfusion of capillaries in the IPL/INL. The choroid rupture was directly correlated to the presence of leakage in the FA. None of the acute injury lesions had choroidal disruption, while the CNV lesions had disruptions proportional to the FA leakage. Figure 8: Color-coded depth-resolved projections of the SVP and DVP in the initial 24 hours following lesion induction. For each condition, the lesions were labeled in order of induction (counterclockwise) for reference. The acute injury response (**a–c**) does not indicate a loss of capillaries. Throughout the initial 24 hours, there was no net loss of SVP or DVP capillaries. The transient response of the inner retina vasculature in the CNV lesions (**d–f**) shows that non-perfusion is immediate. The CNV conditions show that the DVP capillaries are immediately disrupted at 1H (**e**), and the displacement area expands with increased swelling at 24 hours (**f**). Visualizing the CNV model through longitudinal en-face OCTA revealed that the lesions exhibited early microvascular changes before a prolonged lesion growth phase that indicates the development of chronic inflammation can be observed (Figure 10), as the lesions' dimensions failed to reach a steady state at day 21 (Figure 5 Panel B). While most rodent CNV studies focus on days 7-21, relying on histology to confirm neovascularization [48], our results indicate there is an early dynamic process on a scale of hours to 3 days that influences both layer morphology and microvasculature. The implication is that photoreceptor layer swelling causes a deflection of the inner retina during a critical period of 1h to 3d. In this early vascular change, it is important to study how the severity of early inflammation affects the transition from acute to chronic phase 3-7 days. It should be noted that the critical periods for CNV are the early acute phase 1h-3d, 3-7 days for the transition to the chronic phase, and 7-21 days of the chronic phase neovascularization. Figure 10: Alternative processing of the OCTA data to show the changes in the Deep Vascular Plexus over time. A single lesion was isolated in the center of the scan. (**a**) pretreatment, the entire field has clear vessels. (**b**) 1 hour, vessels in center are not well visualized.