Ophthalmology
Abstract
Vision loss in age-related macular degeneration (AMD) stems from disruption of photoreceptor cells in the macula, the central retinal area required for high-acuity vision. Mice and rats have no macula, but surgical insertion of a subretinal implant can induce localized photoreceptor degeneration due to chronic separation from retinal pigment epithelium, simulating a key aspect of AMD. We find that the implant-induced loss of photoreceptors in rat retina leads to local changes in the physiology of downstream retinal ganglion cells (RGCs), similar to changes in RGCs of rodent models of retinitis pigmentosa (RP), an inherited disease causing retina-wide photoreceptor degeneration. The local implant-induced changes in RGCs include enhanced intrinsic excitability leading to accelerated spontaneous firing, increased membrane permeability to fluorescent dyes, and enhanced photosensitization by azobenzene photoswitches. The local physiological changes are correlated with an increase in Retinoic Acid Receptor (RAR)-induced gene transcription, the key process underlying retinal remodeling in mouse models of RP. Hence the loss of photoreceptors, whether by local physical perturbation or by inherited mutation, leads to a stereotypical set of pathophysiological consequences in RGCs. These findings implicate RAR as a possible common therapeutic target for reversing the signal-corrupting effects of retinal remodeling in both RP and AMD.
Authors
Bristol Denlinger, Zachary Helft, Michael Telias, Henri Lorach, Daniel Palanker, Richard H. Kramer
LXRs regulate features of age-related macular degeneration and may be a potential therapeutic target
Abstract
Effective treatments and animal models for the most prevalent neurodegenerative form of blindness in the elderly, called age-related macular degeneration (AMD), are lacking. Genome-wide association studies have identified lipid metabolism and inflammation as AMD-associated pathogenic pathways. Given liver x receptors, encoded by NR1H3 and NR1H2, are master regulators of these pathways, herein we investigated the role of LXR in human and mouse eyes as a function of age and disease, and tested the therapeutic potential of targeting LXR. We identified immunopositive LXR fragments in human extracellular early dry AMD lesions and a decrease in LXR expression within the retinal pigment epithelium (RPE) as a function of age. Aged mice, lacking LXR presented with isoform dependent ocular pathologies. Specifically, loss of the Nr1h3 isoform results in pathobiologies aligned with AMD, supported by compromised visual function, accumulation of native and oxidized lipids in the outer retina, and upregulation of ocular inflammatory cytokines, while absence of Nr1h2 is associated with ocular lipoidal degeneration. Therapeutically, LXR activation, ameliorated lipid accumulation and oxidant-induced injury in RPE cells in vitro, and decreased ocular inflammatory markers and lipid deposition in a mouse model, in vivo, providing translational support for pursuing LXR-active pharmaceuticals as potential therapies for dry AMD.
Authors
Mayur Choudhary, Ebraheim N. Ismail, Pei-Li Yao, Faryan Tayyari, Roxana A. Radu, Steven Nusinowitz, Michael E. Boulton, Rajendra S. Apte, Jeffrey W. Ruberti, James T. Handa, Peter Tontonoz, Goldis Malek
Abstract
Vascular endothelial growth factor A (VEGF) induces angiogenesis and vascular hyperpermeability in ocular tissues and is therefore a key therapeutic target for eye conditions in which these processes are dysregulated. In contrast, the therapeutic potential of VEGF’s neurotrophic roles in the eye has remained unexploited. In particular, it is not known whether modulating levels of any of the 3 major alternatively spliced VEGF isoforms might provide a therapeutic approach to promote neural health in the eye without inducing vascular pathology. Here, we have used a variety of mouse models to demonstrate differences in overall VEGF levels and VEGF isoform ratios across tissues in the healthy eye. We further show that VEGF isoform expression was differentially regulated in retinal versus corneal disease models. Among the 3 major isoforms — termed VEGF120, VEGF164, and VEGF188 — VEGF188 was upregulated to the greatest extent in injured cornea, where it was both necessary and sufficient for corneal nerve regeneration. Moreover, topical VEGF188 application further promoted corneal nerve regeneration without inducing pathological neovascularization. VEGF isoform modulation should therefore be explored further for its potential in promoting neural health in the eye.
Authors
James T. Brash, Laura Denti, Christiana Ruhrberg, Franziska Bucher
Abstract
Mitochondrial quality control (MQC) is crucial for regulating central nervous system homeostasis and its disruption has been implicated in the pathogenesis of some of the most common neurodegenerative diseases. In healthy tissues, the maintenance of MQC depends upon an exquisite balance between mitophagy (removal of damaged mitochondria by autophagy) and biogenesis (de-novo synthesis of mitochondria). Here, we show that mitophagy is disrupted in diabetic retinopathy (DR) and decoupled from mitochondrial biogenesis during the progression of the disease. Diabetic retinas from human post-mortem donors and experimental mice exhibit a net loss of mitochondrial contents during the early stages of the disease process. Using novel diabetic mitophagy-reporter mice (mitoQC-Ins2Akita) alongside pMitoTimer (a molecular clock to address mitochondrial-age dynamics), we demonstrate that mitochondrial loss arose due to an inability of mitochondrial biogenesis to compensate for diabetes-exacerbated mitophagy. However, as diabetes duration increases, Pink1-dependent mitophagy deteriorates, leading to the build-up of mitochondria primed for degradation in DR. Impairment of mitophagy during prolonged diabetes is linked with the development of retinal senescence, a phenotype that blunted hyperglycaemia-induced mitophagy in mitoQC primary Müller cells. Our findings suggest that normalizing mitochondrial turnover may preserve MQC and provide novel therapeutic options for the management of DR-associated complications.
Authors
Jose R. Hombrebueno, Lauren Cairns, Louise R. Dutton, Timothy J. Lyons, Derek P. Brazil, Paul Moynagh, Tim M. Curtis, Heping Xu
Abstract
Von Hippel–Lindau (Vhl) protein inhibits hypoxia-inducible factor (Hif), yet its deletion in murine retina does not cause the extensive angiogenesis expected with Hif induction. The mechanism is unclear. Here we show that retinoblastoma tumor suppressor (Rb1) constrains expression of Hif target genes in the Vhl-/- retina. Deleting Rb1 induced extensive retinal neovascularization and autophagic ablation of photoreceptors in the Vhl-/- retina. RNA sequencing, ChIP and reporter assays showed Rb1 recruitment to and repression of certain Hif target genes. Activating Rb1 by deleting cyclin D1 induced a partial defect in the retinal superficial vascular plexus (SVP). Unexpectedly, removing Vhl suppressed retinoblastoma formation in murine Rb1/Rbl1-deficient retina, but generated subretinal vascular growths resembling retinal angiomatous proliferation (RAP), and retinal capillary hemangioblastoma (RCH). Most stromal cells in the RAP/RCH-like lesions were Sox9+, suggesting a Müller glia origin, and expressed Lgals3, a marker of human brain hemangioblastoma. Thus, the Rb family limit Hif target gene expression in the Vhl-/- retina, and removing this inhibitory signal generates new models for RAP and RCH.
Authors
Ran Wei, Xiang Ren, Hongyu Kong, Zhongping Lv, Yongjiang Chen, Yunjing Tang, Yujiao Wang, Lirong Xiao, Sabiha Hacibekiroglu, Chen Liang, Andras Nagy, Rod Bremner, Danian Chen
Abstract
Age-related macular degeneration (AMD) is the leading cause of central retinal vision loss worldwide, with an estimated 1 in 10 people over the age of 55 showing early signs of the condition. There are currently no forms of therapy available for the end stage of dry AMD, geographic atrophy (GA). Here, we show that the inner blood-retina barrier (iBRB) is highly dynamic and may play a contributory role in GA development. We have discovered that the gene CLDN5, which encodes claudin-5, a tight junction protein abundantly expressed at the iBRB, is regulated by BMAL1 and the circadian clock. Persistent suppression of claudin-5 expression in mice exposed to a cholesterol-enriched diet induced striking retinal pigment epithelium (RPE) cell atrophy, and persistent targeted suppression of claudin-5 in the macular region of nonhuman primates induced RPE cell atrophy. Moreover, fundus fluorescein angiography in human and nonhuman primate subjects showed increased retinal vascular permeability in the evening compared with the morning. These findings implicate an inner retina–derived component in the early pathophysiological changes observed in AMD, and we suggest that restoring the integrity of the iBRB may represent a novel therapeutic target for the prevention and treatment of GA secondary to dry AMD.
Authors
Natalie Hudson, Lucia Celkova, Alan Hopkins, Chris Greene, Federica Storti, Ema Ozaki, Erin Fahey, Sofia Theodoropoulou, Paul F. Kenna, Marian M. Humphries, Annie M. Curtis, Eleanor Demmons, Akeem Browne, Shervin Liddie, Matthew S. Lawrence, Christian Grimm, Mark T. Cahill, Pete Humphries, Sarah L. Doyle, Matthew Campbell
Abstract
The heterogeneity of individual cells in a tissue has been well characterized, largely using ex vivo approaches that do not permit longitudinal assessments of the same tissue over long periods of time. We demonstrate a potentially novel application of adaptive optics fluorescence microscopy to visualize and track the in situ mosaicism of retinal pigment epithelial (RPE) cells directly in the human eye. After a short, dynamic period during which RPE cells take up i.v.-administered indocyanine green (ICG) dye, we observed a remarkably stable heterogeneity in the fluorescent pattern that gradually disappeared over a period of days. This pattern could be robustly reproduced with a new injection and follow-up imaging in the same eye out to at least 12 months, which enabled longitudinal tracking of RPE cells. Investigation of ICG uptake in primary human RPE cells and in a mouse model of ICG uptake alongside human imaging corroborated our findings that the observed mosaicism is an intrinsic property of the RPE tissue. We demonstrate a potentially novel application of fluorescence microscopy to detect subclinical changes to the RPE, a technical advance that has direct implications for improving our understanding of diseases such as oculocutaneous albinism, late-onset retinal degeneration, and Bietti crystalline dystrophy.
Authors
HaeWon Jung, Jianfei Liu, Tao Liu, Aman George, Margery G. Smelkinson, Sarah Cohen, Ruchi Sharma, Owen Schwartz, Arvydas Maminishkis, Kapil Bharti, Catherine Cukras, Laryssa A. Huryn, Brian P. Brooks, Robert Fariss, Johnny Tam
Abstract
Changes in neuronal activity alter blood flow to match energy demand with the supply of oxygen and nutrients. This functional hyperemia is maintained by interactions between neurons, vascular cells, and glia. However, how changing neuronal activity prevalent at the onset of neurodegenerative disease affects neurovascular elements is unclear. Here, in mice with photoreceptor degeneration, a model of neuron-specific dysfunction, we combined assessment of visual function, neurovascular unit structure, and the blood-retina barrier permeability. We found that the rod loss paralleled remodeling of the neurovascular unit, comprised of photoreceptors, retinal pigment epithelium, and Muller glia. When significant visual function was still present, blood flow became disrupted and blood-retina barrier began to fail, facilitating cone loss and vision decline. Thus, in contrast to the established view, vascular deficit in neuronal degeneration is not a late consequence of neuronal dysfunction, but is present early in the course of disease. These findings further establish the importance of vascular deficit and blood retina barrier function in neuron-specific loss, and highlight it as a target for early therapeutic intervention.
Authors
Elena Ivanova, Nazia M. Alam, Glen T. Prusky, Botir T. Sagdullaev
Abstract
While anti-VEGF drugs are commonly used to inhibit pathological retinal and choroidal neovascularization, not all patients respond in an optimal manner. Mechanisms underpinning resistance to anti-VEGF therapy include the upregulation of other pro-angiogenic factors. Therefore, therapeutic strategies that simultaneously target multiple growth factor signalling pathways would have significant value. Here, we show that Ca2+/calmodulin-dependent kinase II (CAMKII) mediates the angiogenic actions of a range of growth factors in human retinal endothelial cells and that this kinase acts as a key nodal point for the activation of several signal transduction cascades that are known to play a critical role in growth factor-induced angiogenesis. We also demonstrate that endothelial CAMKIIγ and δ isoforms differentially regulate the angiogenic effects of different growth factors and that genetic deletion of these isoforms suppresses pathological retinal and choroidal neovascularisation in vivo. Our studies suggest that CAMKII could provide a novel and efficacious target to inhibit multiple angiogenic signalling pathways for the treatment of vasoproliferative diseases of the eye. CAMKIIγ represents a particularly promising target, as deletion of this isoform inhibited pathological neovascularisation, whilst enhancing reparative angiogenesis in the ischemic retina.
Authors
Sadaf Ashraf, Samuel Bell, Caitriona O'Leary, Paul Canning, Ileana Micu, Jose A. Fernandez, Michael O'Hare, Peter Barabas, Hannah McCauley, Derek P. Brazil, Alan W. Stitt, J. Graham McGeown, Tim M. Curtis
Abstract
BACKGROUND. Oculocutaneous albinism (OCA) results in reduced melanin synthesis, skin hypopigmentation, increased risk of UV-induced malignancy, and developmental eye abnormalities affecting vision. No treatments exist. We have shown that oral nitisinone increases ocular and fur pigmentation in a mouse model of one form of albinism, OCA-1B, due to hypomorphic mutations in the Tyrosinase gene. METHODS. In this open-label pilot study, 5 adult patients with OCA-1B established baseline measurements of iris, skin, and hair pigmentation and were treated over 12 months with 2 mg/d oral nitisinone. Changes in pigmentation and visual function were evaluated at 3-month intervals. RESULTS. The mean change in iris transillumination, a marker of melanin, from baseline was 1.0 ± 1.54 points, representing no change. The method of iris transillumination grading showed a high intergrader reliability (intraclass correlation coefficient ≥ 0.88 at each visit). The number of letters read (visual acuity) improved significantly at month 12 for both eyes (right eye, OD, mean 4.2 [95% CI, 0.3, 8.1], P = 0.04) and left eye (OS, 5 [1.0, 9.1], P = 0.003). Skin pigmentation on the inner bicep increased (M index increase = 1.72 [0.03, 3.41], P = 0.047). Finally, hair pigmentation increased by both reflectometry (M index [17.3 {4.4, 30.2}, P = 0.01]) and biochemically. CONCLUSION. Nitisinone did not result in an increase in iris melanin content but may increase hair and skin pigmentation in patients with OCA-1B. The iris transillumination grading scale used in this study proved robust, with potential for use in future clinical trials. TRIAL REGISTRATION. ClinicalTrials.gov NCT01838655. FUNDING. Intramural program of the National Eye Institute.
Authors
David R. Adams, Supriya Menezes, Ramon Jauregui, Zaheer M. Valivullah, Bradley Power, Maria Abraham, Brett G. Jeffrey, Angel Garced, Ramakrishna P. Alur, Denise Cunningham, Edythe Wiggs, Melissa A. Merideth, Pei-Wen Chiang, Shanna Bernstein, Shosuke Ito, Kazumasa Wakamatsu, Rhona M. Jack, Wendy J. Introne, William A. Gahl, Brian P. Brooks
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