Transient RORβ Knockdown by Cell-penetrating siRNA Alleviates Retinal Degeneration Caused by Proteotoxicity

ABSTRACT: Retinitis pigmentosa (RP) is characterized by rod photoreceptor degeneration-driven apoptosis, which secondarily triggers cone photoreceptor loss and eventual blindness. Preserving rod function remains a primary therapeutic objective. Inspired by prior studies demonstrating that neural retina leucine zipper (Nrl) knockdown conferred resistance to rod mutations and induced in vivo rod-to-cone conversion with therapeutic effects in inherited retinal degeneration (IRD) models, we hypothesized that knockdown of retinoid-related orphan receptor beta (RORβ), an upstream regulator of Nrl, could yield similar or superior outcomes. To test this, we developed a cell-penetrating asymmetric siRNA (cp-asiRNA) targeting RORβ (cp-asiRORB), designed to overcome conventional siRNA limitations, including inefficient cellular delivery, off-target effects, and immune activation. Intravitreal administration of cp-asiRORB in RhoP23H mice—an autosomal dominant RP model harboring the P23H rhodopsin mutation—successfully reduced RORβ expression in rod photoreceptors. Contrary to our initial expectation of observing an in vivo reprogramming effect, RORβ knockdown did not seem to induce rod-to-cone conversion. Instead, transient RORβ suppression effectively mitigated apoptosis by enhancing proteostasis, leading to improved rod survival, reduced degeneration, and preserved visual function. Furthermore, single-cell RNA sequencing of retinas treated with cp-asiRORB in RhoP23H mice revealed significant upregulation of proteasomal subunits in RORβ-reduced rod photoreceptors. Consistent with these findings, in HEK293T cells under proteotoxic stress, RORβ knockdown reduced apoptosis, improved cell viability, and diminished aggresome formation. These results demonstrate that transient RORβ suppression alleviates RP progression by augmenting proteasomal activity to resolve proteotoxicity, offering a novel therapeutic avenue for RP patients.

ORGANISM(S): Mus musculus

PROVIDER: GSE291775 | GEO | 2025/09/26

REPOSITORIES: GEO