Stem cell therapy enhancement with gold nanoparticles functionalized with nerve growth factor in an animal model of laser damaged retina

Abstract

Purpose : Currently, there is no cure for the permanent visual loss caused by degenerative retinal diseases. We aimed to develop a novel cell therapy by combining mezenchymal stem cells (MSCs) with nanotechnology to deliver nerve growth factor (NGF) in an animal model of retinal injury.

Methods : MSCs isolated from mice bone marrow were characterized for the expression of specific markers (immuno-fenotyping and RT-PCR). Gold spherical 20 nm nanoparticles (GNPs) were synthesized by citrate reduction of gold salt, functionalized with NGF-beta and characterized by UV-VIS absorption spectroscopy, dynamic light scattering and zeta potential. Retina was injured by indirect diode laser photocoagulation in Bl 57 mice. Subsequently, PKH26 fluorescent MSCs loaded with GNPs conjugated with NGF were injected intravitreally. The GNP-NGF internalization was tested by dark-field microscopy. MSCs grafting and retinal morphology were assessed by confocal laser scanning microscopy (CLSM) and histopathology at 1 and 2 weeks after the administration of MSCs.

Results : Isolated cells for transplant were adult type MSCs. Dark-field microscopy and confocal microscopy are presented in picture 1 and picture 2 respectively. Histopathological analysis indicated the lack of an inflammatory mediated process for GNPs and of a cell mediated rejection process of the transplanted stem cells. Retinal repair processes were identified in all the examined lots, being primarily characterized by the adherence of the transplanted cells to the damaged tissue and the onset of on-site repair processes.

Conclusions : Intravitreal administration of MSCs loaded with GNP-NGF has an acceptable invasiveness and a low risk of infection. GNPs conjugated with NGF offer a proper microenvironment for MSCs grafting and enhance the differentiation into retinal cells. Stem cells can be used as neurotrophic agents and represent an alternative delivery system to viral vectors.

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

 

Dark-field microscopy of MSCs: A-control cells. B- MSCs loaded with GNPs. C- MSCs loaded with GNP-NGF

 

CLSM: migration of MSCs (red) in the retina. A, B – MSCs in the superficial layers of the retina. C – 2.5D reconstruction the MSCs fluorescence intensity compared to the background. D, E - fluorescence peaks of MSCs (white arrows) and their localization within the retina. Draq5-stained nuclei of the retinal cells are shown by the blue fluorescent channel.