transconjunctival-approach-for-injection-into-the-rat-optic-nerve

Transconjunctival Approach for Injection into the Rat Optic Nerve

The rat serves as an important model for studying disorders of the optic nerve, including heritable, traumatic, neoplastic, and autoimmune conditions. ...

University of Iowa

Adeno-associated virus (AAV)-mediated gene therapy has great potential for treating a wide range of retinal degenerative diseases. However, some initial enthusiasm for gene therapy has been tempered by emerging evidence of AAV-associated inflammation, which in several instances has contributed to clinical trial discontinuation. Currently, there is a paucity of data describing the variable immune responses to different AAV serotypes, and similarly, little is known regarding how these responses differ depending on route of ocular delivery, including in animal models of disease. In this study, we characterize the severity and retinal distribution of AAV-associated inflammation in rats triggered by delivery of five different AAV vectors (AAV1, AAV2, AAV6, AAV8, and AAV9), each of which contained enhanced green fluorescent protein (eGFP) driven under control of the constitutively active cytomegalovirus promoter. We further compare the inflammation across three different potential routes (intravitreal, subretinal, and suprachoroidal) of ocular delivery. Compared to buffer-injected controls for each route of delivery, AAV2 and AAV6 induced the most inflammation across all routes of delivery of vectors tested, with AAV6 inducing the highest levels of inflammation when delivered suprachoroidally. AAV1-induced inflammation was highest when delivered suprachoroidally, whereas minimal inflammation was seen with intravitreal delivery. In addition, AAV1, AAV2, and AAV6 each induce infiltration of adaptive immune cells like T cells and B cells into the neural retina, suggesting an innate adaptive response to a single dose of virus. AAV8 and AAV9 induced minimal inflammation across all routes of delivery. Importantly, the degree of inflammation was not correlated with vector-mediated transduction and expression of eGFP. These data emphasize the importance of considering ocular inflammation when selecting AAV serotypes and ocular delivery routes for the development of gene therapy strategies.

The Degree of Adeno-Associated Virus-Induced Retinal Inflammation Varies Based on Serotype and Route of Delivery: Intravitreal, Subretinal, or Suprachoroidal.

In retinal diseases such as age-related macular degeneration (AMD) and choroideremia, a key pathophysiologic step is loss of endothelial cells of the choriocapillaris. Repopulation of choroidal vasculature early in the disease process may halt disease progression. Prior studies have shown that injection of donor cells in suspension results in significant cellular efflux and poor cell survival. As such, the goal of this study was to develop a hydrogel system designed to support choroidal endothelial cell transplantation. A library of hydrogels was synthesized using laminin (i.e., LN111, LN121, and LN421), carboxy methyl chitosan, and oxidized dextran via reversible Schiff base chemistry. Each of the developed self-healing hydrogels was readily injectable into the suprachoroidal space, with ideal gelation, mechanical, and degradation properties. While all hydrogels were found to be compatible with choroidal endothelial cell survival in vitro, only LN111 and LN121 gels were well-tolerated in vivo. To determine if hydrogel mediated cell delivery enhances donor cell retention and survival in vivo, iPSC-derived choroidal endothelial cell laden hydrogels were injected into the suprachoroidal space of an immunocompromised choroidal cell injury rat model. Significantly more donor cells were retained and survived in eyes that received cell laden hydrogels versus contralateral hydrogel free controls. Furthermore, donor cells positive for human nuclear antigen were identified in the choroid of hydrogel eyes only. These findings pave the way for future cell replacement studies in large animal models of choroidal cell dropout focused on evaluating functional integration of donor cells within decellularized vascular tubes. STATEMENT OF SIGNIFICANCE: Age related macular degeneration (AMD) is a leading cause of untreatable blindness in the industrial world. A key pathologic step in AMD is loss of the choriocapillaris endothelial cells, which provide vascular support to the overlying retina. Choroidal cell replacement early in disease may prevent retinal cell death and subsequent vision loss. In this study, we present a strategy for repopulating the choriocapillaris using choroidal endothelial cell laden hydrogels. Specifically, we demonstrate the synthesis and characterization of 3 different laminin-based hydrogel systems. LN111 and LN121 hydrogels were found to have excellent biocompatibility both in vitro and in vivo. Hydrogel mediated delivery of iPSC-derived choroidal endothelial cells enhanced donor cell retention and survival, paving the way for functional large animal studies.

Development of self-healing hydrogels to support choroidal endothelial cell transplantation for the treatment of early age related macular degeneration.

Emily Meyering

Institute for Vision Research

University of Iowa

Transconjunctival Approach for Injection into the Rat Optic Nerve

Emily Meyering

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Institute for Vision Research

University of Iowa

Transconjunctival Approach for Injection into the Rat Optic Nerve

Institute for Vision Research