Very Low-Pressure Subretinal Injection with Internal Limiting Membrane Removal Preserves Retinal Microstructure in Macaque Eyes

Abstract

Purpose : Recently, we reported a new technique of subretinal injection after removal of internal limiting membrane (ILM) at the injection site (RETINA 2016). It allows for injection at a very low pressure (6 psi), but the effect of this technique on the retina is not clear. We investigated the microstructure of the retina after injections with this technique.

Methods : Subretinal injections were performed in 6 eyes of 3 Macaca fascicularises. After vitrectomy, the ILM (1/4 disc area) was removed at the injection site and balanced salt solution was injected with a 38-gauge cannula (MedOne) connected to the Viscous Fluid Control system (Alcon) by placing the cannula tip in contact with the retinal nerve fiber layer. The injection was performed at 6 sites per eye at the injection pressure of either 6 or 20 psi. Optical coherence tomography (OCT, Spectralis, Heidelberg Engineering) was done preoperatively and every 1 week after the injection. Eyes were enucleated at 1 and 6 weeks and processed for light and electron microscopy and TdT-mediated dUTP nick end labeling.

Results : After the injection at 6 psi, the ellipsoid zone (Ez) was continuous on OCT for the entire 6 weeks after the injection, and this correlated with the microscopic findings that the structure of the retina was preserved. After the injection at 20 psi, the Ez was disrupted on OCT at 1 week. Electron microscopy revealed damage of photoreceptor outer segments (OS) and stratification of retinal pigment epithelium (RPE). Ez recovered gradually from 2 weeks on OCT and became continuous at 6 weeks although the alignment did not recover as completely as with 6 psi injection. The recovery of OS was confirmed with electron microscopy and the RPE was restored to a monolayer. Apoptosis of photoreceptor cells was not present after injection at either pressure.

Conclusions : The subretinal injection with the pressure of 6 psi appears safe and causes minimal damage to the microstructure of the retina. This technique may be useful in subretinal injections of gene therapy vectors and regenerative cells.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.