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Medicina (Kaunas, Lithuania) May 2024Hydroxychloroquine sulfate (HCQ) is a lysosomotropic agent administered in systemic lupus erythematosus and rheumatoid arthritis that has fewer toxic effects than...
Hydroxychloroquine sulfate (HCQ) is a lysosomotropic agent administered in systemic lupus erythematosus and rheumatoid arthritis that has fewer toxic effects than chloroquine. However, HCQ may still be responsible for retinal toxicity. In this study, we observed structural changes in the retinas of experimental rats after prolonged exposure to HCQ. We investigated several aspects regarding retinal changes, at both the histopathological and ultrastructural levels. We used 96 male albino Wistar rats distributed into four equal groups (n = 24 per group): the first three groups were treated with different doses of HCQ (50, 100, and 200 mg/kg HCQ, injected intraperitoneally in a single dose daily), and the last group (the control group, n = 24) was treated with saline solution administered in the same way (0.4 mL of saline solution). The treated groups received HCQ daily for 4 months, and every month, six animals from each group were sacrificed to assess retinal changes. The eyes were examined via optical (OM) and electronic microscopy (EM). Statistical analysis was deployed, and results regarding retinal morpho-photometry were acquired. We observed structural retinal changes in both high and low doses of HCQ; while high doses determined a significant thinning of the retina, lower doses caused retinal thickening. Morphological retinal changes upon exposure to HCQ are believed to be caused by accumulated HCQ in lysosomes found in retinal ganglion cells and in the inner nuclear and photoreceptor cell layers. Such changes were most evident in the group receiving HCQ intraperitoneally in doses of 100 mg/kg for a longer period (4 months). The present study highlights histopathological and ultrastructural retinal changes induced by chronic HCQ administration, which were strongly connected to the dosage and period of exposure.
Topics: Hydroxychloroquine; Animals; Rats, Wistar; Rats; Retina; Male; Antirheumatic Agents
PubMed: 38929463
DOI: 10.3390/medicina60060846 -
Antioxidants (Basel, Switzerland) May 2024This study aims to investigate the role of microRNA let-7f in the dysfunction and degeneration of retinal pigment epithelium (RPE) cells through the induction of...
This study aims to investigate the role of microRNA let-7f in the dysfunction and degeneration of retinal pigment epithelium (RPE) cells through the induction of senescence and oxidative stress. Furthermore, we explore whether let-7f inhibition can protect these cells against sodium iodate (SI)-induced oxidative stress. Oxidative stress and let-7f expression are reciprocally regulated in retinal pigment epithelial cells. Overexpression of let-7f in ARPE-19 cells induced oxidative stress as demonstrated by increased reactive oxygen species (ROS) production as well as senescence. Inhibition of let-7f successfully protected RPE cells from the detrimental effects induced by SI. In addition, let-7f overexpression induced RPE cellular dysfunction by diminishing their migratory capabilities and reducing the phagocytosis of porcine photoreceptor outer segments (POS). Results were further confirmed in vivo by intravitreal injections of SI and let-7f antagomir in C57BL/6 mice. Our results provide strong evidence that let-7f is implicated in the dysfunction of RPE cells through the induction of senescence and oxidative injury. These findings may help to uncover novel and relevant processes in the pathogenesis of dry AMD.
PubMed: 38929085
DOI: 10.3390/antiox13060646 -
International Journal of Molecular... Jun 2024Among the myriad of existing tyrosine kinase receptors, the TAM family-abbreviated from Tyro3, Axl, and Mer tyrosine kinase (MerTK)-has been extensively studied with an...
Among the myriad of existing tyrosine kinase receptors, the TAM family-abbreviated from Tyro3, Axl, and Mer tyrosine kinase (MerTK)-has been extensively studied with an outstanding contribution from the team of Prof. Greg Lemke. MerTK activity is implicated in a wide variety of functions involving the elimination of apoptotic cells and has recently been linked to cancers, auto-immune diseases, and atherosclerosis/stroke. In the retina, MerTK is required for the circadian phagocytosis of oxidized photoreceptor outer segments by the retinal-pigment epithelial cells, a function crucial for the long-term maintenance of vision. We previously showed that MerTK ligands carry the opposite role in vitro, with Gas6 inhibiting the internalization of photoreceptor outer segments while Protein S acts conversely. Using site-directed mutagenesis and ligand-stimulated phagocytosis assays on transfected cells, we presently demonstrate, for the first time, that Gas6 and Protein S recognize different amino acids on MerTK Ig-like domains. In addition, MerTK's function in retinal-pigment epithelial cells is rhythmic and might thus rely on the respective stoichiometry of both ligands at different times of the day. Accordingly, we show that ligand bioavailability varies during the circadian cycle using RT-qPCR and immunoblots on retinal and retinal-pigment epithelial samples from control and beta5 integrin knockout mice where retinal phagocytosis is arrhythmic. Taken together, our results suggest that Gas6 and Protein S might both contribute to refine the acute regulation of MerTK in time for the daily phagocytic peak.
Topics: c-Mer Tyrosine Kinase; Phagocytosis; Animals; Intercellular Signaling Peptides and Proteins; Protein S; Humans; Retina; Mice; Circadian Rhythm; Ligands; Retinal Pigment Epithelium; Receptor Protein-Tyrosine Kinases
PubMed: 38928335
DOI: 10.3390/ijms25126630 -
Genes Jun 2024Inherited cone disorders (ICDs) are a heterogeneous sub-group of inherited retinal disorders (IRDs), the leading cause of sight loss in children and working-age adults.... (Review)
Review
Inherited cone disorders (ICDs) are a heterogeneous sub-group of inherited retinal disorders (IRDs), the leading cause of sight loss in children and working-age adults. ICDs result from the dysfunction of the cone photoreceptors in the macula and manifest as the loss of colour vision and reduced visual acuity. Currently, 37 genes are associated with varying forms of ICD; however, almost half of all patients receive no molecular diagnosis. This review will discuss the known ICD genes, their molecular function, and the diseases they cause, with a focus on the most common forms of ICDs, including achromatopsia, progressive cone dystrophies (CODs), and cone-rod dystrophies (CORDs). It will discuss the gene-specific therapies that have emerged in recent years in order to treat patients with some of the more common ICDs.
Topics: Humans; Color Vision Defects; Cone-Rod Dystrophies; Retinal Cone Photoreceptor Cells; Cone Dystrophy; Blindness; Animals; Genetic Therapy
PubMed: 38927662
DOI: 10.3390/genes15060727 -
Translational Vision Science &... Jun 2024To assess longitudinal reproducibility of metrics of foveal density (peak cone density [PCD], cone density centroid [CDC], and 80th percentile centroid area) in...
PURPOSE
To assess longitudinal reproducibility of metrics of foveal density (peak cone density [PCD], cone density centroid [CDC], and 80th percentile centroid area) in participants with normal vision.
METHODS
Participants (n = 19; five male and 14 female) were imaged at two time points (average interval of 3.2 years) using an adaptive optics scanning light ophthalmoscope (AOSLO). Foveally centered regions of interest (ROIs) were extracted from AOSLO montages. Cone coordinate matrices were semiautomatically derived for each ROI, and cone mosaic metrics were calculated.
RESULTS
On average, there were no significant changes in cone mosaic metrics between visits. The average ± SD PCD was 187,000 ± 20,000 cones/mm2 and 189,000 ± 21,700 cones/mm2 for visits 1 and 2, respectively (P = 0.52). The average ± SD density at the CDC was 183,000 ± 19,000 cones/mm2 and 184,000 ± 20,800 cones/mm2 for visits 1 and 2, respectively (P = 0.78). The average ± SD 80th percentile isodensity contour area was 15,400 ± 1800 µm2 and 15,600 ± 1910 µm2 for visits 1 and 2, respectively (P = 0.57).
CONCLUSIONS
Foveal cone mosaic density metrics were highly reproducible in the cohort examined here, although further study is required in more diverse populations.
TRANSLATIONAL RELEVANCE
Determination of the normative longitudinal changes in foveal cone topography is key for evaluating longitudinal measures of foveal cone topography in patients with progressive retinal dystrophies.
Topics: Humans; Retinal Cone Photoreceptor Cells; Male; Fovea Centralis; Female; Adult; Reproducibility of Results; Middle Aged; Cell Count; Young Adult; Ophthalmoscopy; Tomography, Optical Coherence; Visual Acuity
PubMed: 38913007
DOI: 10.1167/tvst.13.6.18 -
Frontiers in Cellular Neuroscience 2024Mechanical sensitive channels expressed in mammalian retinas are effectors of elevated pressure stresses, but it is unclear how their activation affects visual function...
INTRODUCTION
Mechanical sensitive channels expressed in mammalian retinas are effectors of elevated pressure stresses, but it is unclear how their activation affects visual function in pressure-related retinal disorders.
METHODS
This study investigated the role of the transient potential channel vanilloid TRPV4 in photoreceptors and rod bipolar cells (RBCs) with immunohistochemistry, confocal microscopy, electroretinography (ERG), and patch-clamp techniques.
RESULTS
TRPV4 immunoreactivity (IR) was found in the outer segments of photoreceptors, dendrites and somas of PKCα-positive RBCs and other BCs, plexiform layers, and retinal ganglion cells (RGCs) in wild-type mice. TRPV4-IR was largely diminished in the retinas of homozygous TRPV4 transgenic mice. Genetically suppressing TRPV4 expression moderately but significantly enhanced the amplitude of ERG a- and b-waves evoked by scotopic and mesopic lights (0.55 to 200 Rh*rod s) and photopic lights (10-10 Rh*rod s) compared to wild-type mice in fully dark-adapted conditions. The implicit time evoked by dim lights (0.55 to 200 Rh*rod s) was significantly decreased for b-waves and elongated for a-waves in the transgenic mice. ERG b-wave evoked by dim lights is primarily mediated by RBCs, and under voltage-clamp conditions, the latency of the light-evoked cation current in RBCs of the transgenic mice was significantly shorter compared to wild-type mice. About 10% of the transgenic mice had one eye undeveloped, and the percentage was significantly higher than in wild-type mice.
CONCLUSIONS
The data indicates that TRPV4 involves ocular development and is expressed and active in outer retinal neurons, and interventions of TRPV4 can variably affect visual signals in rods, cones, RBCs, and cone ON BCs.
PubMed: 38903773
DOI: 10.3389/fncel.2024.1404929 -
BioRxiv : the Preprint Server For... May 2024The neurovascular unit (NVU), comprising vascular, glial and neural elements, supports the energetic demands of neural computation, but this aspect of the retina's...
The neurovascular unit (NVU), comprising vascular, glial and neural elements, supports the energetic demands of neural computation, but this aspect of the retina's trilaminar vessel network is poorly understood. Only the innermost vessel layer - the superficial vascular plexus (SVP) - is ensheathed by astrocytes, like brain capillaries, whereas glial ensheathment in other layers derives from radial Müller glia. Using serial electron microscopy reconstructions from mouse and primate retina, we find that Müller processes cover capillaries in a tessellating pattern, mirroring the tiled astrocytic endfeet wrapping brain capillaries. However, gaps in the Müller sheath, found mainly in the intermediate vascular plexus (IVP), permit different neuron types to contact pericytes and the endothelial cells directly. Pericyte somata are a favored target, often at spine-like structures with a reduced or absent vascular basement lamina. Focal application of adenosine triphosphate (ATP) to the vitreal surface evoked Ca signals in Müller sheaths in all three vascular layers. Pharmacological experiments confirmed that Müller sheaths express purinergic receptors that, when activated, trigger intracellular Ca signals that are amplified by IP -controlled intracellular Ca stores. When rod photoreceptors die in a mouse model of retinitis pigmentosa ( ), Müller sheaths dissociate from the deep vascular plexus (DVP) but are largely unchanged within the IVP or SVP. Thus, Müller glia interact with retinal vessels in a laminar, compartmentalized manner: glial sheathes are virtually complete in the SVP but fenestrated in the IVP, permitting direct neural-to-vascular contacts. In the DVP, the glial sheath is only modestly fenestrated and is vulnerable to photoreceptor degeneration.
PubMed: 38903067
DOI: 10.1101/2024.04.30.591885 -
Nature Communications Jun 2024Phototransduction involves changes in concentration of ions and other solutes within photoreceptors and in subretinal space, which affect osmotic pressure and the...
Phototransduction involves changes in concentration of ions and other solutes within photoreceptors and in subretinal space, which affect osmotic pressure and the associated water flow. Corresponding expansion and contraction of cellular layers can be imaged using optoretinography (ORG), based on phase-resolved optical coherence tomography (OCT). Until now, ORG could reliably detect only photoisomerization and phototransduction in photoreceptors, primarily in cones under bright stimuli. Here, by employing a phase-restoring subpixel motion correction algorithm, which enables imaging of the nanometer-scale tissue dynamics during minute-long recordings, and unsupervised learning of spatiotemporal patterns, we discover optical signatures of the other retinal structures' response to visual stimuli. These include inner and outer segments of rod photoreceptors, retinal pigment epithelium, and subretinal space in general. The high sensitivity of our technique enables detection of the retinal responses to dim stimuli: down to 0.01% bleach level, corresponding to natural levels of scotopic illumination. We also demonstrate that with a single flash, the optoretinogram can map retinal responses across a 12° field of view, potentially replacing multifocal electroretinography. This technique expands the diagnostic capabilities and practical applicability of optoretinography, providing an alternative to electroretinography, while combining structural and functional retinal imaging in the same OCT machine.
Topics: Tomography, Optical Coherence; Animals; Retinal Pigment Epithelium; Retinal Rod Photoreceptor Cells; Retina; Light; Photic Stimulation; Algorithms; Male
PubMed: 38898002
DOI: 10.1038/s41467-024-49014-5 -
BioRxiv : the Preprint Server For... Jun 2024CLN3 Batten disease (also known as Juvenile Neuronal Ceroid Lipofuscinosis; JNCL) is a lysosomal storage disorder that typically initiates with retinal degeneration but...
PURPOSE
CLN3 Batten disease (also known as Juvenile Neuronal Ceroid Lipofuscinosis; JNCL) is a lysosomal storage disorder that typically initiates with retinal degeneration but is followed by seizure onset, motor decline and premature death. Patient-derived CLN3 disease iPSC-RPE cells show defective phagocytosis of photoreceptor outer segments (POSs). Because modifier genes are implicated in CLN3 disease, our goal here was to investigate a direct link between mutation and POS phagocytosis defect.
METHODS
Isogenic control and mutant stem cell lines were generated by CRISPR-Cas9-mediated biallelic deletion of exons 7 and 8. A transgenic ( ) Yucatan miniswine was also used to study the impact of mutation on POS phagocytosis. POS phagocytosis by cultured RPE cells was analyzed by Western blotting and immunohistochemistry. Electroretinogram, optical coherence tomography and histological analysis of and wild-type miniswine eyes were carried out at 6-, 36-, or 48-month age.
RESULTS
RPE ( RPE) displayed reduced POS binding and consequently decreased uptake of POS compared to isogenic control RPE cells. Furthermore, wild-type miniswine RPE cells phagocytosed POS less efficiently than wild-type POS. Consistent with decreased POS phagocytosis, lipofuscin/autofluorescence was decreased in miniswine RPE at 36 months-of-age and was followed by almost complete loss of photoreceptors at 48 months of age.
CONCLUSIONS
mutation (that affects up to 85% patients) affects both RPE and POSs and leads to photoreceptor cell loss in CLN3 disease. Furthermore, both primary RPE dysfunction and mutant POS independently contribute to impaired POS phagocytosis in CLN3 disease.
PubMed: 38895469
DOI: 10.1101/2024.06.09.597388 -
BioRxiv : the Preprint Server For... Jun 2024Rod photoreceptor formation in the postnatal mouse is a widely used model system for studying mammalian photoreceptor development. This experimental paradigm provides...
Rod photoreceptor formation in the postnatal mouse is a widely used model system for studying mammalian photoreceptor development. This experimental paradigm provides opportunities for both gain and loss-of-function studies which can be accomplished through in vivo plasmid delivery and electroporation. However, the cis-regulatory elements used to implement this approach have not been fully evaluated or optimized for the unique transcriptional environment of photoreceptors. Here we report that the use of a photoreceptor cis-regulatory element from the Crx gene in combination with broadly active promoter elements can increase the targeting of developing rod photoreceptors in the mouse. This can lead to greater reporter expression, as well as enhanced misexpression and loss-of-function phenotypes in these cells. This study also highlights the importance of identifying and testing relevant cis-regulatory elements when planning cell subtype specific experiments. The use of the specific hybrid elements in this study will provide a more efficacious gene delivery system to study mammalian photoreceptor formation.
PubMed: 38895286
DOI: 10.1101/2024.06.06.597220