The many faces of SIRT6 in the retina and retinal pigment epithelium.

Sirtuin 6 (SIRT6) is a member of the mammalian sirtuin family of NAD+-dependent protein deacylases, homologues of the yeast silent information regulator 2 (Sir2). SIRT6 has remarkably diverse functions and plays a key role in a variety of biological processes for maintaining cellular and organismal homeostasis. In this review, our primary aim is to summarize recent progress in understanding SIRT6's functions in the retina and retinal pigment epithelium (RPE), with the hope of further drawing interests in SIRT6 to increase efforts in exploring the therapeutic potential of this unique protein in the vision field. Before describing SIRT6's role in the eye, we first discuss SIRT6's general functions in a wide range of biological contexts. SIRT6 plays an important role in gene silencing, metabolism, DNA repair, antioxidant defense, inflammation, aging and longevity, early development, and stress response. In addition, recent studies have revealed SIRT6's role in macrophage polarization and mitochondrial homeostasis. Despite being initially understudied in the context of the eye, recent efforts have begun to elucidate the critical functions of SIRT6 in the retina and RPE. In the retina, SIRT6 is essential for adult retinal function, regulates energy metabolism by suppressing glycolysis that affects photoreceptor cell survival, protects retinal ganglion cells from oxidative stress, and plays a role in Müller cells during early neurodegenerative events in diabetic retinopathy. In the RPE, SIRT6 activates autophagy in culture and protects against oxidative stress in mice. Taken together, this review demonstrates that better understanding of SIRT6's functions and their mechanisms, both in and out of the context of the eye, holds great promise for the development of SIRT6-targeted strategies for prevention and treatment of blinding eye diseases.

Regional Gene Expression in the Retina, Optic Nerve Head, and Optic Nerve of Mice with Optic Nerve Crush and Experimental Glaucoma.

A major risk factor for glaucomatous optic neuropathy is the level of intraocular pressure (IOP), which can lead to retinal ganglion cell axon injury and cell death. The optic nerve has a rostral unmyelinated portion at the optic nerve head followed by a caudal myelinated region. The unmyelinated region is differentially susceptible to IOP-induced damage in rodent models and human glaucoma. While several studies have analyzed gene expression changes in the mouse optic nerve following optic nerve injury, few were designed to consider the regional gene expression differences that exist between these distinct areas. We performed bulk RNA-sequencing on the retina and separately micro-dissected unmyelinated and myelinated optic nerve regions from naïve C57BL/6 mice, mice after optic nerve crush, and mice with microbead-induced experimental glaucoma (total = 36). Gene expression patterns in the naïve unmyelinated optic nerve showed significant enrichment of the Wnt, Hippo, PI3K-Akt, and transforming growth factor ß pathways, as well as extracellular matrix-receptor and cell membrane signaling pathways, compared to the myelinated optic nerve and retina. Gene expression changes induced by both injuries were more extensive in the myelinated optic nerve than the unmyelinated region, and greater after nerve crush than glaucoma. Changes present three and fourteen days after injury largely subsided by six weeks. Gene markers of reactive astrocytes did not consistently differ between injury states. Overall, the transcriptomic phenotype of the mouse unmyelinated optic nerve was significantly different from immediately adjacent tissues, likely dominated by expression in astrocytes, whose junctional complexes are inherently important in responding to IOP elevation.

Regional Gene Expression in the Retina, Optic Nerve Head, and Optic Nerve of Mice with Experimental Glaucoma and Optic Nerve Crush.

A major risk factor for glaucomatous optic neuropathy is the level of intraocular pressure (IOP), which can lead to retinal ganglion cell axon injury and cell death. The optic nerve has a rostral unmyelinated portion at the optic nerve head followed by a caudal myelinated region. The unmyelinated region is differentially susceptible to IOP-induced damage in rodent models and in human glaucoma. While several studies have analyzed gene expression changes in the mouse optic nerve following optic nerve injury, few were designed to consider the regional gene expression differences that exist between these distinct areas. We performed bulk RNA-sequencing on the retina and on separately micro-dissected unmyelinated and myelinated optic nerve regions from naïve C57BL/6 mice, mice after optic nerve crush, and mice with microbead-induced experimental glaucoma (total = 36). Gene expression patterns in the naïve unmyelinated optic nerve showed significant enrichment of the Wnt, Hippo, PI3K-Akt, and transforming growth factor ß pathways, as well as extracellular matrix-receptor and cell membrane signaling pathways, compared to the myelinated optic nerve and retina. Gene expression changes induced by both injuries were more extensive in the myelinated optic nerve than the unmyelinated region, and greater after nerve crush than glaucoma. Changes three and fourteen days after injury largely subsided by six weeks. Gene markers of reactive astrocytes did not consistently differ between injury states. Overall, the transcriptomic phenotype of the mouse unmyelinated optic nerve was significantly different from immediately adjacent tissues, likely dominated by expression in astrocytes, whose junctional complexes are inherently important in responding to IOP elevation.

Alternative RNA Splicing in the Retina: Insights and Perspectives.

Alternative splicing is a fundamental and highly regulated post-transcriptional process that enhances transcriptome and proteome diversity. This process is particularly important in neuronal tissues, such as the retina, which exhibit some of the highest levels of differentially spliced genes in the body. Alternative splicing is regulated both temporally and spatially during neuronal development, can be cell-type-specific, and when altered can cause a number of pathologies, including retinal degeneration. Advancements in high-throughput sequencing technologies have facilitated investigations of the alternative splicing landscape of the retina in both healthy and disease states. Additionally, innovations in human stem cell engineering, specifically in the generation of 3D retinal organoids, which recapitulate many aspects of the in vivo retinal microenvironment, have aided studies of the role of alternative splicing in human retinal development and degeneration. Here we review these advances and discuss the ongoing development of strategies for the treatment of alternative splicing-related retinal disease.

Fatality assessment and variant risk monitoring for COVID-19 using three new hospital occupancy related metrics.

BACKGROUND: Though case fatality rate (CFR) is widely used to reflect COVID-19 fatality risk, its use is limited by large temporal and spatial variation. Hospital mortality rate (HMR) is also used to assess the severity of COVID-19, but HMR data is not directly available globally. Alternative metrics are needed for COVID-19 severity and fatality assessment. METHODS: We introduce new metrics for COVID-19 fatality risk measurements/monitoring and a new mathematical model to estimate average hospital length of stay for deaths (Ldead) and discharges (Ldis). Multiple data sources were used for our analyses. FINDINGS: We propose three, new metrics: hospital occupancy mortality rate (HOMR), ratio of total deaths to hospital occupancy (TDHOR), and ratio of hospital occupancy to cases (HOCR), for dynamic assessment of COVID-19 fatality risk. Estimated Ldead and Ldis for 501,079 COVID-19 hospitalizations in 34 US states between 7 August 2020 and 1 March 2021 were 18·2(95%CI:17·9-18·5) and 14·0(95%CI:13·9-14·0) days, respectively. We found the dramatic changes in COVID-19 CFR observed in 27 countries during early stages of the pandemic were mostly caused by undiagnosed cases. Compared to the first week of November 2021, the week mean HOCRs (mimics hospitalization-to-case ratio) for Omicron variant (58·6% of US new cases as of 25 December 2021) decreased 65·16% in the US as of 16 January 2022. INTERPRETATION: The new and reliable measurements described here could be useful for COVID-19 fatality risk and variant-associated risk monitoring. FUNDING: No specific funding was associated with the present study.

Retinal homeostasis and metformin-induced protection are not affected by retina-specific Pparδ knockout.

Peroxisome proliferator-activated receptors (PPARs) are a family of three nuclear hormone receptors (PPARα, PPARδ, and PPARγ) that are known to regulate expression of lipid metabolism and oxidative stress genes. Given their role in reducing oxidative stress in a variety of tissues, these genes are likely important for retinal homeostasis. This hypothesis has been further supported by recent studies suggesting that PPAR-activating drugs are protective against retinal degenerations. The objective of the present study was to determine the role of PPARδ in the neuroretina. RNA-seq data show that Pparα and Pparδ are both expressed in the retina, but that Pparδ is expressed at 4-fold higher levels. Single-cell RNAseq data show that Pparδ is broadly expressed in all retinal cell types. To determine the importance of Pparδ to the retina, we generated retina-specific Pparδ knockout mice. We found that deletion of Pparδ had a minimal effect on retinal function or morphology out to 12 months of age and did not increase retinal sensitivity to oxidative stress induced by exposure to bright light. While data show that PPARδ levels were increased by the drug metformin, PPARδ was not necessary for metformin-induced protection from light damage. These data suggest that Pparδ either has a redundant function with Pparα or is not essential for normal neuroretina function or resistance to oxidative stress.

A Drug-Tunable Gene Therapy for Broad-Spectrum Protection against Retinal Degeneration.

Retinal degenerations are a large cluster of diseases characterized by the irreversible loss of light-sensitive photoreceptors that impairs the vision of 9.1 million people in the US. An attractive treatment option is to use gene therapy to deliver broad-spectrum neuroprotective factors. However, this approach has had limited clinical translation because of the inability to control transgene expression. To address this problem, we generated an adeno-associated virus vector named RPF2 that was engineered to express domains of leukemia inhibitory factor fused to the destabilization domain of bacterial dihydrofolate reductase. Fusion proteins containing the destabilization domain are degraded in mammalian cells but can be stabilized with the binding of the drug trimethoprim. Our data show that expression levels of RPF2 are tightly regulated by the dose of trimethoprim and can be reversed by trimethoprim withdrawal. We further show that stabilized RPF2 can protect photoreceptors and prevent blindness in treated mice.