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Diabetologia Jul 2021Homo sapiens evolved under conditions of intermittent food availability and prolonged fasting between meals. Periods of fasting are important for recovery from...
AIMS/HYPOTHESIS
Homo sapiens evolved under conditions of intermittent food availability and prolonged fasting between meals. Periods of fasting are important for recovery from meal-induced oxidative and metabolic stress, and tissue repair. Constant high energy-density food availability in present-day society contributes to the pathogenesis of chronic diseases, including diabetes and its complications, with intermittent fasting (IF) and energy restriction shown to improve metabolic health. We have previously demonstrated that IF prevents the development of diabetic retinopathy in a mouse model of type 2 diabetes (db/db); however the mechanisms of fasting-induced health benefits and fasting-induced risks for individuals with diabetes remain largely unknown. Sirtuin 1 (SIRT1), a nutrient-sensing deacetylase, is downregulated in diabetes. In this study, the effect of SIRT1 stimulation by IF, fasting-mimicking cell culture conditions (FMC) or pharmacological treatment using SRT1720 was evaluated on systemic and retinal metabolism, systemic and retinal inflammation and vascular and bone marrow damage.
METHODS
The effects of IF were modelled in vivo using db/db mice and in vitro using bovine retinal endothelial cells or rat retinal neuroglial/precursor R28 cell line serum starved for 24 h. mRNA expression was analysed by quantitative PCR (qPCR). SIRT1 activity was measured via histone deacetylase activity assay. NR1H3 (also known as liver X receptor alpha [LXRα]) acetylation was measured via western blot analysis.
RESULTS
IF increased Sirt1 mRNA expression in mouse liver and retina when compared with non-fasted animals. IF also increased SIRT1 activity eightfold in mouse retina while FMC increased SIRT1 activity and expression in retinal endothelial cells when compared with control. Sirt1 expression was also increased twofold in neuronal retina progenitor cells (R28) after FMC treatment. Moreover, FMC led to SIRT1-mediated LXRα deacetylation and subsequent 2.4-fold increase in activity, as measured by increased mRNA expression of the genes encoding ATP-binding cassette transporter (Abca1 and Abcg1). These changes were reduced when retinal endothelial cells expressing a constitutively acetylated LXRα mutant were tested. Increased SIRT1/LXR/ABC-mediated cholesterol export resulted in decreased retinal endothelial cell cholesterol levels. Direct activation of SIRT1 by SRT1720 in db/db mice led to a twofold reduction of diabetes-induced inflammation in the retina and improved diabetes-induced visual function impairment, as measured by electroretinogram and optokinetic response. In the bone marrow, there was prevention of diabetes-induced myeloidosis and decreased inflammatory cytokine expression.
CONCLUSIONS/INTERPRETATION
Taken together, activation of SIRT1 signalling by IF or through pharmacological activation represents an effective therapeutic strategy that provides a mechanistic link between the advantageous effects associated with fasting regimens and prevention of microvascular and bone marrow dysfunction in diabetes.
Topics: Animals; Cattle; Cell Death; Cells, Cultured; Diabetes Mellitus, Experimental; Diabetic Angiopathies; Fasting; Gene Expression; Heterocyclic Compounds, 4 or More Rings; Hypoglycemic Agents; Liver X Receptors; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Rats; Retina; Retinal Neurons; Retinal Vessels; Signal Transduction; Sirtuin 1
PubMed: 33770194
DOI: 10.1007/s00125-021-05431-5 -
Proceedings of the National Academy of... May 2023Age-related macular degeneration, Stargardt disease, and their mouse model are characterized by accelerated accumulation of the pigment lipofuscin, derived from...
Age-related macular degeneration, Stargardt disease, and their mouse model are characterized by accelerated accumulation of the pigment lipofuscin, derived from photoreceptor disc turnover in the retinal pigment epithelium (RPE); lipofuscin accumulation and retinal degeneration both occur earlier in albino mice. Intravitreal injection of superoxide (O) generators reverses lipofuscin accumulation and rescues retinal pathology, but neither the target nor mechanism is known. Here we show that RPE contains thin multi-lamellar membranes (TLMs) resembling photoreceptor discs, which associate with melanolipofuscin granules in pigmented mice but in albinos are 10-fold more abundant and reside in vacuoles. Genetically over-expressing tyrosinase in albinos generates melanosomes and decreases TLM-related lipofuscin. Intravitreal injection of generators of O or nitric oxide (NO) decreases TLM-related lipofuscin in melanolipofuscin granules of pigmented mice by ~50% in 2 d, but not in albinos. Prompted by evidence that O plus NO creates a dioxetane on melanin that excites its electrons to a high-energy state (termed "chemiexcitation"), we show that exciting electrons directly using a synthetic dioxetane reverses TLM-related lipofuscin even in albinos; quenching the excited-electron energy blocks this reversal. Melanin chemiexcitation assists in safe photoreceptor disc turnover.
Topics: Mice; Animals; Melanins; Lipofuscin; Macular Degeneration; Retina; Retinal Pigment Epithelium; ATP-Binding Cassette Transporters
PubMed: 37155898
DOI: 10.1073/pnas.2216935120 -
International Journal of Molecular... Feb 2022Fluid and solute transporters of the retinal pigment epithelium (RPE) are core components of the outer blood-retinal barrier. Characterizing these transporters and their...
Fluid and solute transporters of the retinal pigment epithelium (RPE) are core components of the outer blood-retinal barrier. Characterizing these transporters and their role in retinal homeostasis may provide insights into ocular function and disease. Here, we describe RPE defects in mice, which exhibit hypopigmented patches in the central retina. Mapping and nucleotide sequencing of mice revealed a disrupted 5' splice donor sequence in , a sodium bicarbonate cotransporter gene. expression was reduced 19.7-fold in RPE relative to controls, and alternative splice variants were detected. SLC4A5 was localized to the Golgi apparatus of cultured human RPE cells and in apical and basal membranes. Fundus imaging, optical coherence tomography, microscopy, and electroretinography (ERG) of mice revealed retinal detachment, hypopigmented patches corresponding to neovascular lesions, and retinal folds. Detachment worsened and outer nuclear layer thickness decreased with age. ERG a- and b-wave response amplitudes were initially normal but declined in older mice. The direct current ERG fast oscillation and light peak were reduced in amplitude at all ages, whereas other RPE-associated responses were unaffected. These results link a new mutation to subretinal fluid accumulation and altered light-evoked RPE electrophysiological responses, suggesting that SLC4A5 functions at the outer blood-retinal barrier.
Topics: Animals; Cells, Cultured; Disease Models, Animal; Female; Humans; Male; Mice; Mice, Inbred C57BL; Mutation; RNA Splicing; Retina; Retinal Detachment; Retinal Pigment Epithelium; Sodium-Bicarbonate Symporters; Tomography, Optical Coherence
PubMed: 35216333
DOI: 10.3390/ijms23042220 -
Clinical & Experimental Optometry Sep 2020Posterior microphthalmos (PM) is a rare developmental disorder characterised by high hyperopia, short axial length, presence of retinal papillomacular fold and... (Comparative Study)
Comparative Study
BACKGROUND
Posterior microphthalmos (PM) is a rare developmental disorder characterised by high hyperopia, short axial length, presence of retinal papillomacular fold and relatively normal anterior segment findings. The study objective is to describe the retinal structural and vascular changes in eyes with PM with spectral domain optical coherence tomography, optical coherence tomography angiography and multicolour imaging.
METHODS
In this retrospective, comparative case series, 10 eyes of five patients with PM as cases and 10 eyes of five age- and sex-matched controls were included. Structural changes, namely inner and outer retinal layer thicknesses, were measured using optical coherence tomography. Multicolour imaging findings were noted. Perifoveal vascular changes with qualitative and quantitative assessments were analysed using optical coherence tomography angiography.
RESULTS
The foveal dip was absent in all 10 eyes (100 per cent) with PM. There was an elevated retinal papillomacular fold in six eyes (60 per cent) and intraretinal cystoid spaces in two eyes (20 per cent) with PM. The inner retinal layers were thicker in PM. On multicolour imaging, foveal avascular zone and retinal wrinkles were identified in eyes with retinal papillomacular fold in blue and green reflectance images. Perifoveal vascular changes in optical coherence tomography angiography included foveal area size reduction in superficial and deep vascular networks. The foveal capillary vessel densities were higher in PM compared to the controls in both superficial (46.3 ± 3.7 per cent; p = 0.000) and deep (54.7 ± 3.5 per cent; p = 0.000) capillary plexuses. Flow areas in superficial (p = 0.693) and deep (p = 0.088) capillary plexuses were not statistically relevant.
CONCLUSION
The study suggests that in PM, retinal foreshortening occurs in one meridian leading to secondary changes such as loss of foveal dip, posterior bowing of the outer layers of the eye, thickening of the inner retina and ultimately, retinal papillomacular fold formation. The findings of the study need further validation in a larger series of patients with PM.
Topics: Adolescent; Adult; Biometry; Female; Fluorescein Angiography; Follow-Up Studies; Fovea Centralis; Fundus Oculi; Humans; Male; Microphthalmos; Retinal Vessels; Retrospective Studies; Tomography, Optical Coherence; Visual Acuity; Young Adult
PubMed: 31489720
DOI: 10.1111/cxo.12966 -
Stem Cell Research & Therapy Jan 2021Adult mammalian retinal stem cells (RSCs) readily proliferate, self-renew, and generate progeny that differentiate into all retinal cell types in vitro. RSC-derived...
BACKGROUND
Adult mammalian retinal stem cells (RSCs) readily proliferate, self-renew, and generate progeny that differentiate into all retinal cell types in vitro. RSC-derived progeny can be induced to differentiate into photoreceptors, making them a potential source for retinal cell transplant therapies. Despite their proliferative propensity in vitro, RSCs in the adult mammalian eye do not proliferate and do not have a regenerative response to injury. Thus, identifying and modulating the mechanisms that regulate RSC proliferation may enhance the capacity to produce RSC-derived progeny in vitro and enable RSC activation in vivo.
METHODS
Here, we used medium-throughput screening to identify small molecules that can expand the number of RSCs and their progeny in culture. In vitro differentiation assays were used to assess the effects of synthetic glucocorticoid agonist dexamethasone on RSC-derived progenitor cell fate. Intravitreal injections of dexamethasone into adult mouse eyes were used to investigate the effects on endogenous RSCs.
RESULTS
We discovered that high-affinity synthetic glucocorticoid agonists increase RSC self-renewal and increase retinal progenitor proliferation up to 6-fold without influencing their differentiation in vitro. Intravitreal injection of synthetic glucocorticoid agonist dexamethasone induced in vivo proliferation in the ciliary epithelium-the niche in which adult RSCs reside.
CONCLUSIONS
Together, our results identify glucocorticoids as novel regulators of retinal stem and progenitor cell proliferation in culture and provide evidence that GCs may activate endogenous RSCs.
Topics: Animals; Cell Differentiation; Cell Proliferation; Cell Self Renewal; Cells, Cultured; Glucocorticoids; Mice; Retina
PubMed: 33494791
DOI: 10.1186/s13287-021-02136-9 -
Investigative Ophthalmology & Visual... Sep 2015To determine the frequency, patterns, associations, and biomechanical implications of retinal and choroidal folds in papilledema due to idiopathic intracranial...
PURPOSE
To determine the frequency, patterns, associations, and biomechanical implications of retinal and choroidal folds in papilledema due to idiopathic intracranial hypertension (IIH).
METHODS
Retinal and choroidal folds were studied in patients enrolled in the IIH Treatment Trial using fundus photography (n = 165 study eyes) and spectral-domain optical coherence tomography (SD-OCT; n = 125). We examined the association between folds and peripapillary shape, retinal nerve fiber layer (RNFL) thickness, disc volume, Frisén grade, acuity, perimetric mean deviation, intraocular pressure, intracranial pressure, and refractive error.
RESULTS
We identified three types of folds in IIH patients with papilledema: peripapillary wrinkles (PPW), retinal folds (RF), and choroidal folds (CF). Frequency, with photos, was 26%, 19%, and 1%, respectively; SD-OCT frequency was 46%, 47%, and 10%. At least one type of fold was present in 41% of patients with photos and 73% with SD-OCT. Spectral-domain OCT was more sensitive. Structural parameters related to the severity of papilledema were associated with PPW and RF, whereas anterior deformation of the peripapillary RPE/basement membrane layer was associated with CF and RF. Folds were not associated with vision loss at baseline.
CONCLUSIONS
Folds in papilledema are biomechanical signs of stress/strain on the optic nerve head and load-bearing structures induced by intracranial hypertension. Folds are best imaged with SD-OCT. The patterns of retinal and choroidal folds are the products of a complex interplay between the degree of papilledema and anterior deformation of the load-bearing structures (sclera and possibly the lamina cribrosa), both modulated by structural geometry and material properties of the optic nerve head. (ClinicalTrials.gov number, NCT01003639.).
Topics: Adult; Aged; Biomechanical Phenomena; Choroid; Female; Humans; Intracranial Hypertension; Intracranial Pressure; Intraocular Pressure; Male; Middle Aged; Nerve Fibers; Optic Disk; Papilledema; Refractive Errors; Retina; Tomography, Optical Coherence; Visual Acuity
PubMed: 26335066
DOI: 10.1167/iovs.15-17459 -
Graefe's Archive For Clinical and... Oct 2022In many retinal pathological conditions, rod and cone degeneration differs. For example, the early-onset maculopathy Stargardts disease type 1 (STGD1) is typified by... (Review)
Review
PURPOSE
In many retinal pathological conditions, rod and cone degeneration differs. For example, the early-onset maculopathy Stargardts disease type 1 (STGD1) is typified by loss of cones while rods are often less affected. We wanted to examine whether there exist intrinsic membrane differences between rods and cones that might explain such features.
METHODS
Abca4 mRNA and protein levels were quantified in rod- and cone-enriched samples from wild-type and Nrl mice retinas; rod- and cone-enriched outer segments (ROS and COS respectively) were prepared from pig retinas, and total lipids were analyzed by flame ionization, chromatography, and tandem mass spectrometry. Immunohistochemical staining of cone-rich rodent Arvicanthis ansorgei retinas was conducted, and ultra-high performance liquid chromatography of lipid species in porcine ROS and COS was performed.
RESULTS
Abca4 mRNA and Abca4 protein content was significantly higher (50-300%) in cone compared to rod-enriched samples. ROS and COS displayed dramatic differences in several lipids, including very long chain poly-unsaturated fatty acids (VLC-PUFAs), especially docosahexaenoic acid (DHA, 22:6n-3): ROS 20.6% DHA, COS 3.3% (p < 0.001). VLC-PUFAs (> 50 total carbons) were virtually absent from COS. COS were impoverished (> 6× less) in phosphatidylethanolamine compared to ROS. ELOVL4 ("ELOngation of Very Long chain fatty acids 4") antibody labelled Arvicanthis cones only very weakly compared to rods. Finally, there were large amounts (905 a.u.) of the bisretinoid A2PE in ROS, whereas it was much lower (121 a.u., ~ 7.5-fold less) in COS fractions. In contrast, COS contained fivefold higher amounts of all-trans-retinal dimer (115 a.u. compared to 22 a.u. in rods).
CONCLUSIONS
Compared to rods, cones expressed higher levels of Abca4 mRNA and Abca4 protein, were highly impoverished in PUFA (especially DHA) and phosphatidylethanolamine, and contained significant amounts of all-trans-retinal dimer. Based on these and other data, we propose that in contrast to rods, cones are preferentially vulnerable to stress and may die through direct cellular toxicity in pathologies such as STGD1.
Topics: Animals; Docosahexaenoic Acids; Murinae; Phosphatidylethanolamines; RNA, Messenger; Reactive Oxygen Species; Retinal Cone Photoreceptor Cells; Retinal Degeneration; Retinaldehyde; Swine
PubMed: 35524799
DOI: 10.1007/s00417-022-05684-9 -
Investigative Ophthalmology & Visual... Feb 2020Vigabatrin (VGB) is an effective antiepileptic that increases concentrations of inhibitory γ-aminobutyric acid (GABA) by inhibiting GABA transaminase. Reports of...
PURPOSE
Vigabatrin (VGB) is an effective antiepileptic that increases concentrations of inhibitory γ-aminobutyric acid (GABA) by inhibiting GABA transaminase. Reports of VGB-associated visual field loss limit its clinical usefulness, and retinal toxicity studies in laboratory animals have yielded conflicting results.
METHODS
We examined the functional and morphologic effects of VGB in C57BL/6J mice that received either VGB or saline IP from 10 to 18 weeks of age. Retinal structure and function were assessed in vivo by optical coherence tomography (OCT), ERG, and optomotor response. After euthanasia, retinas were processed for immunohistochemistry, and retinal GABA, and VGB quantified by mass spectrometry.
RESULTS
No significant differences in visual acuity or total retinal thickness were identified between groups by optomotor response or optical coherence tomography, respectively. After 4 weeks of VGB treatment, ERG b-wave amplitude was enhanced, and amplitudes of oscillatory potentials were reduced. Dramatic rod and cone bipolar and horizontal cell remodeling, with extension of dendrites into the outer nuclear layer, was observed in retinas of VGB-treated mice. VGB treatment resulted in a mean 3.3-fold increase in retinal GABA concentration relative to controls and retinal VGB concentrations that were 20-fold greater than brain.
CONCLUSIONS
No evidence of significant retinal thinning or ERG a- or b-wave deficits were apparent, although we describe significant alterations in ERG b-wave and oscillatory potentials and in retinal cell morphology in VGB-treated C57BL/6J mice. The dramatic concentration of VGB in retina relative to the target tissue (brain), with a corresponding increase in retinal GABA, offers insight into the pathophysiology of VGB-associated visual field loss.
Topics: Animals; Anticonvulsants; GABA Agents; Male; Mice, Inbred C57BL; Neuronal Plasticity; Oculomotor Muscles; Random Allocation; Retina; Retinal Diseases; Tomography, Optical Coherence; Vigabatrin; Visual Fields
PubMed: 32053727
DOI: 10.1167/iovs.61.2.17 -
International Journal of Molecular... Aug 2021To investigate the retinal proteins associated with primary and secondary retinal ganglion cell (RGC) degeneration and explore their molecular pathways, SWATH label-free...
To investigate the retinal proteins associated with primary and secondary retinal ganglion cell (RGC) degeneration and explore their molecular pathways, SWATH label-free and target-based mass spectrometry was employed to identify the proteomes in various retinal locations in response to localized optic nerve injury. Unilateral partial optic nerve transection (pONT) was performed on adult Wistar rats and their retinas were harvested 2 weeks later. To confirm the separation of primary and secondary RGC degeneration, immunohistochemistry of RNA binding protein with multiple splicing (RBPMS) and glial fibrillary acidic protein (GFAP) was performed on retinal whole-mounts. Retinal proteomes in the temporal and nasal quadrants were evaluated with high resolution hybrid quadrupole time-of-flight mass spectrometry (QTOF-MS), and SWATH-based acquisition, and their expression was compared to the corresponding retinal quadrant in contralateral control eyes and further validated by multiple reaction monitoring mass spectrometry (MRM-MS). A total of 3641 proteins (FDR < 1%) were identified using QTOF-MS. The raw data are available via ProteomeXchange with the identifier PXD026783. Bioinformatics data analysis showed that there were 37 upregulated and 25 downregulated proteins in the temporal quadrant, whereas 20 and five proteins were upregulated and downregulated, respectively, in the nasal quadrant, respectively ( = 4, < 0.05; fold change ≥ 1.4-fold or ≤0.7). Six proteins were regulated in both the temporal and the nasal quadrants, including CLU, GFAP, GNG5, IRF2BPL, L1CAM, and CPLX1. Linear regression analysis indicated a strong association between the data obtained by means of SWATH-MS and MRM-MS (temporal, R = 0.97; nasal, R = 0.96). Gene ontology analysis revealed statistically significant changes in the biological processes and cellular components of primary RGC degeneration. The majority of the significant changes in structural, signaling, and cell death proteins were associated with the loss of RGCs in the area of primary RGC degeneration. The combined use of SWATH-MS and MRM-MS methods detects and quantifies regional changes of retinal protein expressions after localized injury. Future investigation with this integrated approach will significantly increase the understanding of diverse processes of progressive RGC degeneration from a proteomic prospective.
Topics: Animals; Eye Proteins; Mass Spectrometry; Nerve Degeneration; Optic Nerve Injuries; Proteome; Proteomics; Rats; Rats, Wistar; Retina; Retinal Degeneration; Retinal Ganglion Cells
PubMed: 34445296
DOI: 10.3390/ijms22168592 -
Molecular Vision 2021Collagen is a key player contributing to vitreoelasticity and vitreoretinal adhesions. Molecular reorganization causes spontaneous weakening of these adhesions with age,...
PURPOSE
Collagen is a key player contributing to vitreoelasticity and vitreoretinal adhesions. Molecular reorganization causes spontaneous weakening of these adhesions with age, resulting in the separation of the posterior hyaloid membrane (PHM) from the retina in what is called complete posterior vitreous detachment (PVD). Incomplete separation of the posterior hyaloid or tight adherence or both can lead to retinal detachment, vitreomacular traction syndrome, or epiretinal membrane formation, which requires surgical intervention. Pharmacological vitrectomy has the potential of avoiding surgical vitrectomy; it is also useful as an adjunct during retinal surgery to induce PVD. Previously studied enzymatic reagents, such as collagenase derived from are nonspecific and potentially toxic. We studied a novel collagenase from (VMC) which remains active (VMA), even after deletion of 51 C-terminal amino acids. To limit the activity of VMA to the vitreous cavity, a fusion construct (inhibitor of hyaluronic acid-VMA [iHA-VMA]) was made in which a 12-mer peptide (iHA, which binds to HA) was fused to the N-terminus of VMA. The construct was evaluated in the context of PVD.
METHODS
VMA and iHA-VMA were expressed in purified, and characterized with gelatin zymography, collagen degradation assay, fluorescamine-based assay, and cell-based assays. Two sets of experiments were performed in New Zealand albino rabbits. Group A (n = 10) received iHA-VMA, while group B (n = 5) received the equivalent dose of VMA. In both groups, saline was injected as a control in the contralateral eyes. Animals were monitored with indirect ophthalmoscopy, optical coherence tomography (OCT), and B-scan ultrasonography. Retinal toxicity was assessed with hematoxylin and eosin (H&E) staining of retinal tissue.
RESULTS
The activity of iHA-VMA and VMA was comparable and 65-fold lower than that of collagenase Type IV. In the iHA-VMA group, all the rabbits (n = 10) developed PVD, with complete PVD seen in six animals. No statistically significant histomorphological changes were seen. In the VMA group, four of the five rabbits developed complete PVD; however, retinal morphological changes were seen in two animals.
CONCLUSIONS
iHA-VMA displays targeted action confined to the vitreous and shows potential for safe pharmacologic vitreolysis.
Topics: Animals; Cell Survival; Collagenases; Electrophoresis, Polyacrylamide Gel; Flow Cytometry; Goats; Hyaluronic Acid; Intravitreal Injections; Microscopy, Electron, Scanning; Ophthalmoscopy; Rabbits; Recombinant Fusion Proteins; Retina; Vibrio mimicus; Vitrectomy; Vitreous Body; Vitreous Detachment
PubMed: 33907368
DOI: No ID Found