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Gayathri Tummala, Adam Crain, Sarah John, Jessica Rowlan, Maureen Neitz, Russell Van Gelder, Kathryn Pepple; Characterization of intraocular inflammation following intravitreal AAV injection in a mouse model. Invest. Ophthalmol. Vis. Sci. 2020;61(7):443.
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© ARVO (1962-2015); The Authors (2016-present)
Adeno-associated virus (AAV) is the primary vector used for human intraocular gene therapy. Although AAV has low immunogenic potential, some studies have reported an inflammatory response that has not been well characterized. A better understanding of this immune response would help guide preventative strategies. In this study, we characterize the timing and composition of the cellular infiltrate in mouse eyes following AAV intravitreal injection.
Two AAV capsid types (2 and 7m8) carrying a photoreceptor specific expression cassette (PR2.1 GFP) were injected intravitreally into C57Bl/6 mice at doses between 109 to 1011 genomes. Clinical inflammation was assessed on serial optical coherence tomography (OCT) images. Cell infiltrates were characterized by flow cytometry using an LSRII cytometer. Results were analyzed using FlowJo and statistical analysis was performed using Prism v8.0.
A total of 20/21 AAV injected mice developed signs of inflammation compared to 0/5 sham injected mice. The majority of mice developed inflammation starting on day 7 after injection (n=15), although there was a range from day 6 (n=3) to day 15 (n=2). In 16/20 eyes, inflammation peaked within 15 days after injection: day 7 (n=7), day 10 (n=3), day 14 (n=4), and day 15 (n=2). Clinical inflammation resolved within one month of injection in all mice. Flow performed on day 7 eyes (n=3) identified a total of 22,032 CD45+ cells/injected eye (fellow eyes: 1,114 CD45+ cells/eye). The populations identified were T cells (56.8%), Natural Killer (NK) cells (20.2%), Ly6C+ monocytes (4.7%), B cells (2.7%), and neutrophils (2.1%). Flow performed on day 29 eyes (n=4) identified a total of 5,742 CD45+ cells/injected eye (fellow eyes: 764 CD45+ cells/eye). The populations identified were T cells (56.5%), B cells (7.6%), NK cells (4.2%), Ly6C+ monocytes (3.3%), and neutrophils (1%).
Intravitreal AAV generates inflammation in a majority of mice. Clinically evident inflammation is mild and resolves spontaneously. Even in quiet eyes, a large number and range of inflammatory cells are present. Future experiments will be needed to determine the underlying mechanisms recruiting these cells and the functional significance of their presence to the long-term outcomes of AAV intravitreal gene therapy.
This is a 2020 ARVO Annual Meeting abstract.
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