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Vision Research Nov 2021Following photopigment bleaching, the rhodopsin and cone-opsins show a characteristic exponential regeneration in the dark with a photocycle dependent on the retinal...
Following photopigment bleaching, the rhodopsin and cone-opsins show a characteristic exponential regeneration in the dark with a photocycle dependent on the retinal pigment epithelium. Melanopsin pigment regeneration in animal models requires different pathways to rods and cones. To quantify melanopsin-mediated light adaptation in humans, we first estimated its photopigment regeneration kinetics through the photo-bleach recovery of the intrinsic melanopsin pupil light response (PLR). An intense broadband light (~120,000 Td) bleached 43% of melanopsin compared to 86% of the cone-opsins. Recovery from a 43% bleach was 3.4X slower for the melanopsin than cone-opsin. Post-bleach melanopsin regeneration followed an exponential growth with a 2.5 min time-constant (τ) that required 11.2 min for complete recovery; the half-bleaching level (I) was ~ 4.47 log melanopic Td (16.10 log melanopsin effective photons.cm.s; 8.25 log photoisomerisations.photoreceptor.s). The effect on the cone-directed PLR of the level of the melanopsin excitation during continuous light adaptation was then determined. We observed that cone-directed pupil constriction amplitudes increased by ~ 10% when adapting lights had a higher melanopic excitation but the same mean photometric luminance. Our findings suggest that melanopsin light adaptation enhances cone signalling along the non-visual retina-brain axis. Parameters τ and I will allow estimation of the level of melanopsin bleaching in any light units; the data have implications for quantifying the relative contributions of putative melanopsin pathways to regulate the post-bleach photopigment regeneration and adaptation.
Topics: Adaptation, Ocular; Dark Adaptation; Humans; Light; Photobleaching; Retinal Cone Photoreceptor Cells; Retinal Rod Photoreceptor Cells; Rod Opsins
PubMed: 34315092
DOI: 10.1016/j.visres.2021.07.005 -
Aging Jul 2023Long noncoding RNAs (lncRNAs) play important roles in the development of age-related macular degeneration (AMD). However, the effect of long non-coding RNA activated by...
Long noncoding RNAs (lncRNAs) play important roles in the development of age-related macular degeneration (AMD). However, the effect of long non-coding RNA activated by DNA damage (NORAD) on AMD remains unknown. This study aimed to investigate the effect of NORAD on RPE cell senescence and degeneration. Irradiated adult retinal pigment epithelial cell line-19 (ARPE-19) and sodium iodate-treated mice were used as and AMD models. Results showed that irradiation-induced AMD characteristics of ARPE-19 and NORAD-knockdown aggravated cell cycle arrest in the G2/M phase, cell apoptosis and cell senescence along with the increased expression of phosphorylated P53 (p-P53) and P21. AMD factors C3, ICAM-1, APP, APOE, and VEGF-A were also increased by NORAD-knockdown. Moreover, NORAD-knockdown increased irradiation-induced reduction of mitochondrial homeostasis factors, (i.e., TFAM and POLG) and mitochondrial respiratory chain complex genes (i.e., ND1 and ND5) along with mitochondrial reactive oxygen species (ROS). We also identified a strong interaction of NORAD and PGC-1α and sirtuin 1 (SIRT1) in ARPE-19; that is, NORAD knockdown increases the acetylation of PGC-1α. In NORAD knockout mice, NORAD-knockout accelerated the sodium iodate-reduced retinal thickness reduction, function impairment and loss of retinal pigment in the fundus. Therefore, NORAD-knockdown accelerates retinal cell senescence, apoptosis, and AMD markers via PGC-1α acetylation, mitochondrial ROS, and the p-P53-P21signaling pathway, in which NORAD-mediated effect on PGC-1α acetylation might occur through the direct interaction with PGC-1α and SIRT1.
Topics: Mice; Animals; Reactive Oxygen Species; Oxidative Stress; RNA, Long Noncoding; Tumor Suppressor Protein p53; Sirtuin 1; Macular Degeneration; Retinal Pigments; Retinal Pigment Epithelium
PubMed: 37517088
DOI: 10.18632/aging.204917 -
The Journal of Comparative Neurology Jun 2022Obtaining a parts list of the sensory components of the retina is vital to understand the effects of light in behavior, health, and disease. Rods, cones, and...
Obtaining a parts list of the sensory components of the retina is vital to understand the effects of light in behavior, health, and disease. Rods, cones, and intrinsically photosensitive retinal ganglion cells (ipRGCs) are the best described photoreceptors in the mammalian retina, but recent functional roles have been proposed for retinal neuropsin (Opn5)-an atypical opsin. However, little is known about the pattern of Opn5 expression in the retina. Using cre (Opn5 ) and cre-dependent reporters, we uncover patterns of Opn5 expression and find that Opn5 is restricted to retinal ganglion cells (RGCs). Opn5-RGCs are nonhomogenously distributed through the retina, with greater densities of cells located in the dorsotemporal quadrant. In addition to the local topology of these cells, using cre-dependent AAV viral tracing, we surveyed their central targets and found that they are biased towards image-forming and image-stabilizing regions. Finally, molecular and electrophysiological profiling reveal that Opn5-RGCs comprise previously defined RGC types that respond optimally to edges and object-motion (F-mini-ONs, HD2, HD1, LEDs, ooDSRGCs, etc.). Together, these data describe the second collection of RGCs that express atypical opsins in the mouse, and expand the roles of image-forming cells in retinal physiology and function.
Topics: Animals; Mammals; Membrane Proteins; Mice; Opsins; Retina; Retinal Cone Photoreceptor Cells; Retinal Ganglion Cells; Rod Opsins
PubMed: 34743323
DOI: 10.1002/cne.25272 -
Nature Communications Oct 2022The retinal pigment epithelium (RPE) plays an important role in the development of diabetic retinopathy (DR), a leading cause of blindness worldwide. Here we set out to...
The retinal pigment epithelium (RPE) plays an important role in the development of diabetic retinopathy (DR), a leading cause of blindness worldwide. Here we set out to explore the role of Akt2 signaling-integral to both RPE homeostasis and glucose metabolism-to DR. Using human tissue and genetically manipulated mice (including RPE-specific conditional knockout (cKO) and knock-in (KI) mice), we investigate whether Akts in the RPE influences DR in models of diabetic eye disease. We found that Akt1 and Akt2 activities were reciprocally regulated in the RPE of DR donor tissue and diabetic mice. Akt2 cKO attenuated diabetes-induced retinal abnormalities through a compensatory upregulation of phospho-Akt1 leading to an inhibition of vascular injury, inflammatory cytokine release, and infiltration of immune cells mediated by the GSK3β/NF-κB signaling pathway; overexpression of Akt2 has no effect. We propose that targeting Akt1 activity in the RPE may be a novel therapy for treating DR.
Topics: Animals; Cytokines; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Epithelial Cells; Glucose; Glycogen Synthase Kinase 3 beta; Humans; Mice; NF-kappa B; Proto-Oncogene Proteins c-akt; Retinal Pigment Epithelium; Retinal Pigments
PubMed: 36229454
DOI: 10.1038/s41467-022-33773-0 -
International Journal of Molecular... Jun 2023Diabetic retinopathy (DR) is the leading cause of vision loss and a critical complication of diabetes with a very complex etiology. The build-up of reactive oxygen...
Diabetic retinopathy (DR) is the leading cause of vision loss and a critical complication of diabetes with a very complex etiology. The build-up of reactive oxygen species (ROS) due to hyperglycemia is recognized as a primary risk factor for DR. Although spermidine, a naturally occurring polyamine, has been reported to have antioxidant effects, its effectiveness in DR has not yet been examined. Therefore, in this study, we investigated whether spermidine could inhibit high glucose (HG)-promoted oxidative stress in human retinal pigment epithelial (RPE) cells. The results demonstrated that spermidine notably attenuated cytotoxicity and apoptosis in HG-treated RPE ARPE-19 cells, which was related to the inhibition of mitochondrial ROS production. Under HG conditions, interleukin (IL)-1β and IL-18's release levels were markedly increased, coupled with nuclear factor kappa B (NF-κB) signaling activation. However, spermidine counteracted the HG-induced effects. Moreover, the expression of nucleotide-binding oligomerization domain-like receptor (NLR) protein 3 (NLRP3) inflammasome multiprotein complex molecules, including TXNIP, NLRP3, ASC, and caspase-1, increased in hyperglycemic ARPE-19 cells, but spermidine reversed these molecular changes. Collectively, our findings demonstrate that spermidine can protect RPE cells from HG-caused injury by reducing ROS and NF-κB/NLRP3 inflammasome pathway activation, indicating that spermidine could be a potential therapeutic compound for DR treatment.
Topics: Humans; Inflammasomes; NF-kappa B; NLR Family, Pyrin Domain-Containing 3 Protein; Reactive Oxygen Species; Spermidine; Oxidative Stress; Diabetic Retinopathy; Glucose; Epithelial Cells; Retinal Pigments
PubMed: 37445726
DOI: 10.3390/ijms241310550 -
Investigative Ophthalmology & Visual... Aug 2023To model the in vivo effects of chloroquine on the retinal pigment epithelium in experimentally tractable cell culture systems and determine the effects of mild...
PURPOSE
To model the in vivo effects of chloroquine on the retinal pigment epithelium in experimentally tractable cell culture systems and determine the effects of mild chloroquine treatment on lysosome function and turnover.
METHODS
Effects of low-dose chloroquine treatment on lysosomal function and accessibility to newly endocytosed cargo were investigated in primary and embryonic stem cell-derived RPE cells and ARPE19 cells using fluorescence and electron microscopy of fluorescent and gold-labeled probes. Lysosomal protein expression and accumulation were measured by quantitative PCR and Western blotting.
RESULTS
Initial chloroquine-induced lysosome neutralization was followed by partial recovery, lysosomal expansion, and accumulation of undegraded endocytic, phagocytic, and autophagic cargo and inhibition of cathepsin D processing. Accumulation of enlarged lysosomes was accompanied by a gradual loss of accessibility of these structures to the endocytic pathway, implying impaired lysosome reformation. Chloroquine-induced accumulation of pro-cathepsin D, as well as the lysosomal membrane protein, LAMP1, was reproduced by treatment with protease inhibitors and preceded changes in lysosomal gene expression.
CONCLUSIONS
Low-dose chloroquine treatment inhibits lysosome reformation, causing a gradual depletion of lysosomes able to interact with cargo-carrying vacuoles and degrade their content. The resulting accumulation of newly synthesized pro-cathepsin D and LAMP1 reflects inhibition of normal turnover of lysosomal constituents and possibly lysosomes themselves. A better understanding of the mechanisms underlying lysosome reformation may reveal new targets for the treatment of chloroquine-induced retinopathy.
Topics: Humans; Chloroquine; Lysosomes; Phagocytosis; Autophagy; Retinal Diseases; Epithelial Cells; Retinal Pigments
PubMed: 37548963
DOI: 10.1167/iovs.64.11.10 -
Cells May 2022Multinucleated retinal pigment epithelium (RPE) cells have been reported in humans and other mammals. Rodents have an extremely high percentage of multinucleated cells...
Multinucleated retinal pigment epithelium (RPE) cells have been reported in humans and other mammals. Rodents have an extremely high percentage of multinucleated cells (more than 80%). Both mouse and human multinucleated RPE cells exhibit specific regional distributions that are potentially correlated with photoreceptor density. However, detailed investigations of multinucleated RPE in different species and their behavior after DNA damage are missing. Here, we compared the composition of multinucleated RPE cells in nocturnal and diurnal animals that possess distinct rod and cone proportions. We further investigated the reactive oxygen species (ROS) production and DNA damage response in mouse mononucleated and multinucleated RPE cells and determined the effect of p53 dosage on the DNA damage response in these cells. Our results revealed an unrealized association between multinucleated RPE cells and nocturnal vision. In addition, we found multinucleated RPE cells exhibited increased ROS production and DNA damage after X-ray irradiation. Furthermore, haploinsufficiency of p53 led to increased DNA damage frequency after irradiation, and mononucleated RPE cells were more sensitive to a change in p53 dosage. In conclusion, this study provides novel information on in vivo PRE topography and the DNA damage response, which may reflect specific requirements for vision adaption and macular function.
Topics: Animals; DNA Damage; Epithelial Cells; Mammals; Mice; Reactive Oxygen Species; Retinal Pigment Epithelium; Retinal Pigments; Tumor Suppressor Protein p53
PubMed: 35563857
DOI: 10.3390/cells11091552 -
Proceedings of the National Academy of... Jun 2023Transplantation of stem cell-derived retinal pigment epithelial (RPE) cells is considered a viable therapeutic option for age-related macular degeneration (AMD). Several...
Transplantation of stem cell-derived retinal pigment epithelial (RPE) cells is considered a viable therapeutic option for age-related macular degeneration (AMD). Several landmark Phase I/II clinical trials have demonstrated safety and tolerability of RPE transplants in AMD patients, albeit with limited efficacy. Currently, there is limited understanding of how the recipient retina regulates the survival, maturation, and fate specification of transplanted RPE cells. To address this, we transplanted stem cell-derived RPE into the subretinal space of immunocompetent rabbits for 1 mo and conducted single-cell RNA sequencing analyses on the explanted RPE monolayers, compared to their age-matched in vitro counterparts. We observed an unequivocal retention of RPE identity, and a trajectory-inferred survival of all in vitro RPE populations after transplantation. Furthermore, there was a unidirectional maturation toward the native adult human RPE state in all transplanted RPE, regardless of stem cell resource. Gene regulatory network analysis suggests that tripartite transcription factors (, , and ) may be specifically activated in posttransplanted RPE cells, to regulate canonical RPE signature gene expression crucial for supporting host photoreceptor function, and to regulate prosurvival genes required for transplanted RPE's adaptation to the host subretinal microenvironment. These findings shed insights into the transcriptional landscape of RPE cells after subretinal transplantation, with important implications for cell-based therapy for AMD.
Topics: Adult; Animals; Humans; Rabbits; Transcriptome; Macular Degeneration; Stem Cells; Epithelial Cells; Retinal Pigments
PubMed: 37339216
DOI: 10.1073/pnas.2214842120 -
Scientific Reports Oct 2020Drosophila melanogaster has long been a popular model insect species, due in large part to the availability of genetic tools and is fast becoming the model for insect...
Drosophila melanogaster has long been a popular model insect species, due in large part to the availability of genetic tools and is fast becoming the model for insect colour vision. Key to understanding colour reception in Drosophila is in-depth knowledge of spectral inputs and downstream neural processing. While recent studies have sparked renewed interest in colour processing in Drosophila, photoreceptor spectral sensitivity measurements have yet to be carried out in vivo. We have fully characterised the spectral input to the motion and colour vision pathways, and directly measured the effects of spectral modulating factors, screening pigment density and carotenoid-based ocular pigments. All receptor sensitivities had significant shifts in spectral sensitivity compared to previous measurements. Notably, the spectral range of the Rh6 visual pigment is substantially broadened and its peak sensitivity is shifted by 92 nm from 508 to 600 nm. We show that this deviation can be explained by transmission of long wavelengths through the red screening pigment and by the presence of the blue-absorbing filter in the R7y receptors. Further, we tested direct interactions between inner and outer photoreceptors using selective recovery of activity in photoreceptor pairs.
Topics: Animals; Color Perception; Drosophila melanogaster; Eye; Photoreceptor Cells, Invertebrate; Retinal Pigments; Vision, Ocular
PubMed: 33106518
DOI: 10.1038/s41598-020-74742-1 -
Molecular and Cellular Endocrinology Nov 2023Pyroptosis of retinal pigment epithelium (RPE) cells is associated with the etiology of diabetic retinopathy (DR). In this study, we investigated the effect of DNMT1 on...
BACKGROUND
Pyroptosis of retinal pigment epithelium (RPE) cells is associated with the etiology of diabetic retinopathy (DR). In this study, we investigated the effect of DNMT1 on RPE cell pyroptosis by regulating miR-20a/TXNIP expression through DNA methylation.
METHODS
High glucose (HG)-induced ARPE-19 cells and mice were injected with streptozotocin (STZ) to generate DR cells and animal models. RT‒qPCR was used to detect the expression of miR-20a, and methylation-specific PCR (MS-PCR) was used to determine the occurrence of methylation of miR-20a. The expression of pyroptosis-related proteins (caspase-1 and NLRP3) and DNA methyltransferase (DNMT1) was detected by western blotting, and the expression of inflammatory factors (IL-1β and IL-18) was detected by ELISA. Apoptosis was detected by flow cytometry and TUNEL. HE staining was used to observe the pathological changes in retinal tissue in mice.
RESULTS
In HG-induced DR cell models, the expression of miR-20a was significantly downregulated, while the expression of inflammatory factors (IL-1β, IL-18) and pyroptosis-associated proteins (caspase-1, NLRP3) was significantly upregulated. Transfection of miR-20a mimic can effectively reverse HG-induced pyroptosis and release of inflammatory factors. DNMT1 promotes miR-20a methylation and inhibits the expression of miR-20a. DNMT1-mediated methylation is involved in the pyroptosis process of high glucose-induced RPE cells, and silencing DNMT1 can promote the expression of miR-20a, thereby inhibiting the release of IL-1β and IL-18 and reducing the occurrence of cell pyroptosis. miR-20a targets negative regulation of TXNIP expression, and overexpression of TXNIP can effectively reverse the inhibitory effect of miR-20a on pyroptosis. The methylation inhibitor 5-AZ can inhibit the occurrence of pyroptosis and DR processes, while treatment with a miR-20a inhibitor or OE-TXNIP can reverse the effect of 5-AZ.
CONCLUSION
DNMT1 promotes DNA methylation, decreases the expression of miR-20a and increases the expression of TXNIP, which ultimately leads to the occurrence of pyroptosis in RPE cells.
Topics: Mice; Animals; Pyroptosis; MicroRNAs; Interleukin-18; NLR Family, Pyrin Domain-Containing 3 Protein; DNA Methylation; Caspase 1; Diabetic Retinopathy; Glucose; Epithelial Cells; Retinal Pigments
PubMed: 37506869
DOI: 10.1016/j.mce.2023.112012