Dark Rearing (DR) as a means of mimicking ‘Physiological Hypoxia’: A rationale for non-invasive treatment of Retinopathy of Prematurity

Abstract

Purpose: : The initiating event in the pathogenesis of ROP is hyperoxia-induced regression of the retinal vasculature via VEGF down regulation. We hypothesize that neonates in a high oxygen environment should be subject to total darkness as it causes a metabolic sump, driven by continuous ion currents in the photoreceptor outer segments.

Methods: : Sprague-Dawley rat pups were DR from birth to P30, and under hyperoxic conditions (55-60 or 70-75% O2) with and without DR, and sacrificed at regular intervals. Retinas were examined for vascular density index; pericyte and astrocyte ensheathment, TEM; and VEGF, & HIF-1a expression. Retinal function was assessed using fERG in P18 & P30 DR rats vs rats in 12hr light/dark cycle. P30 was selected to represent the likely upper limit of DR required for a premature infant. Recovery group rats were returned to cyclic light at P30 and recorded at P60 & P90.

Results: : Retinas from DR rats raised in room air had significantly higher blood vessel density compared to age-matched controls (39.4±4.2 vs. 50.8±2.4, P0.05). When DR was combined with 70-75% O2, arterial oxygen resulted in significant vaso-obliteration (vascular density 33.8±1.6, P<0.05). Retinas from neonates returned to room air, and dark reared in 55-60% O2, showed near-normal mural cell ensheathment; at 70-75% O2, abnormal preretinal neovascular formations were seen. At P8 in animals raised in cyclic light and 70-75% O2 there was significant vasoproliferation, and VEGF mRNA levels were significantly greater than controls (P<0.01). However, in DR hyperoxic animals (70-75% O2) at P8, there was no statistical difference in VEGF expression compared to controls (P<0.01) Average b-wave amplitudes from DR versus dark/light rats were not significantly different for all groups (n=6).

Conclusions: : DR protects vessels from oxygen-induced vaso-obliteration without harmful effects on normal microvascular cell interactions, retinal morphology and function. DR also attenuates increased VEGF that can lead to pathological vasoproliferation after hyperoxia. We suggest that DR precludes the initiation of ROP, offering a viable, noninvasive treatment for prevention of ROP. This is timely, given the push for clinical adoption of anti-VEGF therapy for ROP, where possible systemic effects are not fully investigated.

Keywords: retinopathy of prematurity