Biology subjects

Ail, D.

Publications and source records attributed to Ail, D..

4 recordsLinked to original sources

bioRxiv2025

AAV-delivered CRISPR-Cas9 elicits persistent retinal immune responses compared with transient responses to RNP

CRISPR-Cas9 is a powerful gene-editing tool with great potential for treating genetic diseases, including inherited retinal disorders. However, its bacterial origin can induce immune responses that may eliminate transduced cells, threatening editing efficiency. A deeper understanding of CRISPR-Cas9 immunogenicity is therefore needed. Previous studies have shown that systemic delivery via Cas9 induces an immune response, but the detailed inflammation and the impact of the vector remain unclear, especially in immune-privileged organs like the eye. In this study, we found that Cas9 delivered to the retina using adeno-associated virus (AAV) induced persistent inflammation, whereas delivery as naked ribonucleoprotein (RNP) complexes resulted in acute inflammation that faded three weeks post-injection. Inflammation was more severe in the rd10 mouse model of inherited retinal degeneration, which exhibits basal inflammation. These findings provide new insights into vector-dependent immune responses to Cas9 in the eye and highlight potential risks associated with its clinical application. TEASERUnderstanding immune reactions to CRISPR-Cas9 and linking these to their delivery methodology increases their safety.

bioengineering↗

bioRxiv2022

Inducible non-human primate models of retinal degeneration for testing end stage therapies and understanding disease mechanisms

The anatomical differences between the retinas of humans and most animal models pose a challenge for testing novel therapies. Non-human primate (NHP) retina is anatomically closest to the human retina with the presence of a high acuity region called the fovea. However, there is a lack of relevant NHP models for retinal degeneration that can be used for preclinical studies of vision restoration. To address this unmet need we aimed to generate inducible NHP models of photoreceptor degeneration. We generated three cynomolgus macaque models using distinct strategies. We used two genetically targeted strategies using optogenetics and Crispr-Cas9 to ablate specifically rods to mimic rod-cone dystrophy. Additionally, we created an acute model by physical separation of the photoreceptors and retinal pigment epithelium using a polymer patch. Retinal degeneration was evaluated in all three models by in-life exams such as fundus imaging, optical coherence tomography, adaptive optics and electroretinography. In the genetic models we observed punctuate areas of degeneration in the injected area marked by disorganization of outer segments, loss of rod photoreceptors and thinning of the outer nuclear layer. In the acute model, the degeneration was faster and involved both rods and cones. Among the three distinct NHP models, the Crispr-Cas9 based approach was the most advantageous model in view of recapitulating disease specific features and its ease of implementation. The acute model however resulted in the fastest degeneration making it the most relevant model for testing end-stage vision restoration therapies such as stem cell transplantation.

bioengineering↗

bioRxiv2021

Systemic and local immune response to intraocular AAV vector administration in non-human primates

The positive clinical outcomes in adeno-associated virus (AAV)-mediated retinal gene therapy have often been attributed to the low immunogenicity of AAVs along with the immune-privilege of the eye. However, several recent preclinical studies and clinical trials have shown potential for inflammatory responses to AAV mediated gene therapy. Our current understanding of the factors contributing to intraocular inflammation such as the existence of serum antibodies against AAVs prior to injection and their contribution to increases in antibody levels post-injection is incomplete. The parameters that regulate the generation of new antibodies in response to the AAV capsid or transgene post-injection after intraocular administration are also insufficiently described. In this study we carried out a retrospective analysis of the pre-existing serum antibodies in correlation with changes in antibody levels after intraocular injections of AAV in non-human primates (NHPs). We analyzed NHP serums for the presence of both Binding Antibodies (BABs), as well as a subset of these called Neutralizing Antibodies (NABs) that impede AAV transduction upon binding. We observed significantly higher pre-existing serum BABs against AAV8 compared to other serotypes. We observed a dose-dependent increase in both BABs and NABs in the serums collected post-injection, irrespective of the serotype or the mode of injection. Lastly, we were able to demonstrate a co-relation between the serum BAB levels with clinical grading of inflammation and levels of transgene expression.

immunology↗

bioRxiv2018

Linking YAP to Müller glia quiescence exit in the degenerative retina

Contrasting with fish or amphibian, retinal regeneration from Muller glial cells is largely limited in mammals. In our quest towards the identification of molecular cues that may boost their stemness potential, we investigated the involvement of the Hippo pathway effector YAP, which we previously found to be upregulated in Muller cells following retinal injury. We report that conditional Yap deletion in Muller cells prevents the upregulation of cell cycle genes that normally accompanies reactive gliosis upon photoreceptor cell death. This occurs as a consequence of defective EGFR signaling. Consistent with a function of YAP in triggering Muller glia cell cycle re-entry, we further show that in Xenopus, a species endowed with efficient regenerative capacity, YAP is required for their injury-dependent proliferative response. Finally, and noteworthy, we reveal that YAP overactivation in mouse Muller cells is sufficient to induce their reprogramming into highly proliferative cells. Overall, we unravel a pivotal role for YAP in tuning Muller cell response to injury and highlight a novel YAP-EGFR axis by which Muller cells exit their quiescence state, a critical step towards regeneration.

developmental biology↗