Related Experiment Video
Updated: Jun 4, 2026
Limbal Approach-Subretinal Injection of Viral Vectors for Gene Therapy in Mice Retinal Pigment Epithelium
Published on: August 7, 2015
Adenovirus-mediated delivery of CD46 attenuates the alternative complement pathway on RPE: implications for
J H Sweigard1, S M Cashman, R Kumar-Singh
1Department of Ophthalmology and Neuroscience, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Boston, MA 02111, USA.
Abstract:
Activation of the alternative pathway of the complement system has been implicated in the pathogenesis of age-related macular degeneration. Membrane attack complex (MAC) has been identified mainly on the Bruch's membrane and drusen underlying the retinal pigment epithelium (RPE). Membrane cofactor protein (CD46) preferentially regulates the alternative pathway of complement. The aim of this study was to evaluate the potential of increasing CD46 expression on RPE cells using an adenovirus as a gene therapy approach to reduce alternative pathway-mediated damage to RPE cells. We generated a recombinant adenovirus vector expressing human CD46 (hCD46) and delivered the vector to murine hepatocytes and RPE cells in vitro. After incubation in human serum in conditions in which the classical pathway of complement was blocked, we measured alternative pathway-mediated damage of these cells by quantifying lysis and MAC formation. Adenovirus expressing hCD46 was delivered to the subretinal space of adult mice, and 1 week later, ocular flat mounts were challenged with human serum and the levels of complement-mediated damage was quantified. Adenovirus-mediated delivery of hCD46 localizes to the basal and lateral surfaces of RPE cells where it offers protection from alternative pathway-mediated damage, but not classical, allowing the classical pathway to function unhindered.
Insights
Gene therapy using adenovirus to increase membrane cofactor protein (CD46) on retinal cells protects against complement-mediated damage implicated in age-related macular degeneration.
Area of Science:
- Ophthalmology
- Immunology
- Genetics
Background:
- Age-related macular degeneration (AMD) pathogenesis involves complement system activation.
- Membrane attack complex (MAC) deposition is observed in AMD lesions.
- Membrane cofactor protein (CD46) regulates the complement alternative pathway.
Purpose of the Study:
- To investigate gene therapy with adenovirus to enhance CD46 expression on retinal pigment epithelium (RPE) cells.
- To assess the protective effect of increased CD46 against complement-mediated RPE damage.
Main Methods:
- Constructed a recombinant adenovirus vector expressing human CD46 (hCD46).
- Delivered the vector to murine hepatocytes and RPE cells in vitro.
- Administered the adenovirus to the subretinal space of adult mice.
- Assessed complement-mediated cell lysis and MAC formation.
Main Results:
- Adenovirus-mediated hCD46 delivery to RPE cells reduced alternative pathway-mediated damage.
- hCD46 localized to basal and lateral RPE surfaces, providing protection.
- The classical complement pathway remained functional.
Conclusions:
- Adenovirus-mediated CD46 gene therapy shows potential for treating AMD.
- Targeting CD46 offers a protective strategy against alternative pathway-driven RPE damage.
- This approach preserves classical pathway function, crucial for immune defense.
