Abstract
Purpose:
The lack of regeneration found in the adult mammalian retina following optic nerve injury has led to the study of cell and tissue engineered cell delivery methods. However when designing these delivery devices, it is necessary to orient axon growth and cellular organization to mimic that of the nerve fiber layer as well as stimulate dendrite growth and synapse formation with the cell binding partners in the host retina.
Methods:
RGC layer scaffolds were produced by electrospinning medical grade PLA using a radial collector which orients fiber formation from the outer edge to a central point. Fibrous scaffolds were reacted using a gradient filter and UV initiated photo crosslinker to immobilize Netrin-1, polarizing RGC growth. RGC-seeded scaffolds were then cultured in a 3D model with amacrine cells or transplanted to explanted rat retinas. Following co-culture and transplantation models, samples were fixed and evaluated for synapse formation by immunostaining as a measure of integration.
Results:
RGCs seeded on radial scaffolds mimicked the organization of the mammalian retina with 81% of axons orienting radially. When scaffolds were immobilized with a radial-in netrin gradient, the seeded cells were further polarized to >50% growing axons towards the scaffold center. RGCs formed synaptic connections to amacrine cells in in vitro models and to explanted rat retinas.
Conclusions:
Using this scaffold in both a 3D model and ex vivo transplantation systems we have observed the formation of synapses by immunostaining, a step towards the goal of RGC transplantation.