Inflammasome Activation in Retinal Pigment Epithelium from Human Donors with Age-Related Macular Degeneration.
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ABSTRACT:
SUBMITTER: Ebeling MC
PROVIDER: S-EPMC9265516 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:To identify disease-specific transcriptional programs in retinal pigment epithelium (RPE) cells, fibroblasts from 43 patients with geographic atrophy (GA) were reprogrammed into induced pluripotent stem cells (iPSCs) before being differentiated into RPE and compared to those from 36 healthy individuals. 127,659 RPE cells were profiled via single cell RNA-sequencing (scRNA-seq) and cell classification identified 7 cellular states related to RPE maturation.
2022-06-20 | E-MTAB-11642 | biostudies-arrayexpress
Project description:Inflammation and oxidative stress are involved in age-related macular degeneration (AMD) and possibly associated with an activation of neuronal apoptosis inhibitor protein/class II transcription activator of the Major Histocompatibility Complex (MHC)/heterokaryon incompatibility/telomerase-associated protein 1, leucine-rich repeat or nucleotide-binding domain, leucine-rich repeat-containing family, and pyrin domain-containing 3 (NLRP3) inflammasome. In the present study, we used a translational approach to address this hypothesis. In patients with AMD, we observed increased mRNA levels of NLRP3, pro-interleukin-1 beta (IL-1β) and pro-IL-18 in AMD lesions of the retinal pigment epithelium (RPE) and photoreceptor. In vitro, a similar increase was evoked by oxidative stress or lipopolysaccharide (LPS) stimulation in the adult retinal pigment epithelium (ARPE-19) cell line, and the increase was reduced in siRNA transfected cells to knockdown NLRP3. Ultrastructural studies of ARPE-19 cells showed a swelling of the cytoplasm, mitochondrial damage, and occurrence of autophagosome-like structures. NLRP3 positive dots were detected within autophagosome-like structures or in the extracellular space. Next, we used a mouse model of AMD, Ccl2/Cx3cr1 double knockout on rd8 background (DKO rd8) to ascertain the in vivo relevance. Ultrastructural studies of the RPE of these mice showed damaged mitochondria, autophagosome-like structures, and cytoplasmic vacuoles, which are reminiscent of the pathology seen in stressed ARPE-19 cells. The data suggest that the NLRP3 inflammasome may contribute in AMD pathogenesis.
| S-EPMC4730317 | biostudies-literature
Project description:Age-related macular degeneration (AMD) is the leading cause of irreversible blindness in developed countries, characterized by the death of retinal pigment epithelial (RPE) cells and photoreceptors. Previous studies report an accumulation of damaged and dysfunctional mitochondria in RPE of human donors with AMD. Understanding how damaged mitochondria accumulate in AMD is an important step in discovering disease mechanisms and identifying therapeutic targets. In this report, we assessed mitochondrial fission and fusion by quantifying proteins and measured mitochondrial autophagy (mitophagy) via protein analysis and advanced imaging techniques using mitochondrial targeted mKeima in primary human RPE from donors with or without AMD. We report disease-specific differences in mitochondrial proteins that regulate fission, fusion, and mitophagy that were present at baseline and with treatments to stimulate these pathways. Data suggest AMD RPE utilize receptor-mediated mitophagy as a compensatory mechanism for deficits in the ubiquitin-mediated mitophagy pathway. These changes in mitochondrial homeostasis could lead to the buildup of damaged and dysfunctional mitochondria observed in the RPE of AMD donors.
| S-EPMC9758189 | biostudies-literature
Project description:Induced pluripotent stem cells generated from patients with geographic atrophy as well as healthy individuals were differentiated to retinal pigment epithelium (RPE) cells. By integrating transcriptional profiles of 127,659 RPE cells generated from 43 individuals with geographic atrophy and 36 controls with genotype data, we identified 439 expression Quantitative Trait (eQTL) loci in cis that were associated with disease status and specific to subpopulations of RPE cells. We identified loci linked to two genes with known associations with geographic atrophy - PILRB and PRPH2, in addition to 43 genes with significant genotype x disease interactions that are candidates for novel genetic associations for geographic atrophy. On a transcriptome-only level, we identified molecular pathways significantly upregulated in geographic atrophy-RPE including in mitochondrial functions, metabolic pathways, and extracellular cellular matrix reorganisation. We subsequently implemented a large-scale proteomics analysis, confirming modification in proteins associated with these pathways. We also identified six significant protein (p) QTL that regulate protein expression in the RPE cells and in geographic atrophy - two of which share variants with cis-eQTL, including proteins involve in mitochondrial biology and neuodegeneration. Investigation of mitochondrial functions in the two cohorts confirmed a modification of respiration etc…. This study provides strong proof of concept of the validity of using iPSC for the modeling of complex diseases. It is the first to use a large scale patient -derived iPSC cohort to uncover important differences in RPE homeostasis associated with geographic atrophy. It clearly identifies mitochondrial activity as a core constitutive difference of the RPE from patients with geographic atrophy, and could be a target of potential therapies for this condition (STACEY/MATT).
2022-06-01 | PXD029501 | Pride
Project description:There are currently no treatments for geographic atrophy, the advanced form of age-related macular degeneration. Hence, innovative studies are needed to model this condition and prevent or delay its progression. Induced pluripotent stem cells generated from patients with geographic atrophy and healthy individuals were differentiated to retinal pigment epithelium. Integrating transcriptional profiles of 127,659 retinal pigment epithelium cells generated from 43 individuals with geographic atrophy and 36 controls with genotype data, we identify 445 expression quantitative trait loci in cis that are asssociated with disease status and specific to retinal pigment epithelium subpopulations. Transcriptomics and proteomics approaches identify molecular pathways significantly upregulated in geographic atrophy, including in mitochondrial functions, metabolic pathways and extracellular cellular matrix reorganization. Five significant protein quantitative trait loci that regulate protein expression in the retinal pigment epithelium and in geographic atrophy are identified - two of which share variants with cis- expression quantitative trait loci, including proteins involved in mitochondrial biology and neurodegeneration. Investigation of mitochondrial metabolism confirms mitochondrial dysfunction as a core constitutive difference of the retinal pigment epithelium from patients with geographic atrophy. This study uncovers important differences in retinal pigment epithelium homeostasis associated with geographic atrophy.
| S-EPMC9325891 | biostudies-literature
Project description:PurposeTo test whether increased light transmission (hypertransmission) through subretinal drusenoid deposits (SDD) into the choroid in age-related macular degeneration (AMD) represented retinal pigment epithelium (RPE) degeneration.DesignCross-sectional study.MethodsNineteen eyes of 12 patients with early- to intermediate-stage AMD and 18 eyes of 12 normal subjects were evaluated with color fundus photography, optical coherence tomography (OCT), and high-resolution adaptive optics scanning laser ophthalmoscopy (AOSLO) at baseline and 24 months later. SDD were classified using an OCT-based 3-stage grading system. Hypertransmission beneath SDD into the choroid was examined in OCT. SDD microstructure was assessed with AOSLO. To characterize the hypertransmission-associated chorioretinal degeneration, choroidal thickness and photoreceptor length were measured in OCT at 1 mm and 2 mm superior, inferior, temporal, and nasal to the foveal center.ResultsOCT disclosed hypertransmission beneath stage 3 SDD in 8 eyes. These lesions showed a distinctive regressing structure in AOSLO, compared with stage 3 lesions without hypertransmission. The phenomenon persisted at follow-up, and new hypertransmission developed as SDD advanced. In eyes with hypertransmission, choroids were thinner than those of normal eyes at all sites (by 44%-56%, P ≤ .0028) and those of eyes with SDD but without hypertransmission at superior and temporal sites (by 31%-46%, P ≤ .039). Photoreceptors were significantly shorter than those in normal eyes (by 6%-26%, P ≤ .0379).ConclusionsHypertransmission into the choroid, accompanied with SDD regression and thinning of choroid and photoreceptor layers, indicates RPE degeneration associated with advanced stages in the SDD life cycle.
| S-EPMC5337135 | biostudies-literature
Project description:Retinal cell therapy can have the objectives of rescue (i.e., modulation of metabolic abnormalities primarily for sight preservation) as well as replacement (i.e., replace cells lost due to injury or disease for sight restoration as well as preservation). The first clinical trials of retinal pigment epithelium (RPE) transplantation for vision-threatening complications of age-related macular degeneration (AMD) have begun with some preliminary signs of success (e.g., improvement in vision in some patients, anatomic evidence of transplant-host integration with some evidence of host photoreceptor recovery, long-term survival of autologous induced pluripotent stem cell-derived RPE transplants without immune suppression) as well as limitations (e.g., limited RPE suspension survival in the AMD eye, limited tolerance for long-term systemic immune suppression in elderly patients, suggestion of uncontrolled cell proliferation in the vitreous cavity). RPE survival on aged and AMD Bruch's membrane can be improved with chemical treatment, which may enhance the efficacy of RPE suspension transplants in AMD patients. Retinal detachment, currently used to deliver transplanted RPE cells to the subretinal space, induces disjunction of the first synapse in the visual pathway: the photoreceptor-bipolar synapse. This synaptic change occurs even in areas of attached retina near the locus of detachment. Synaptic disjunction and photoreceptor apoptosis associated with retinal detachment can be reduced with Rho kinase inhibitors. Addition of Rho kinase inhibitors may improve retinal function and photoreceptor survival after subretinal delivery of cells either in suspension or on scaffolds.
| S-EPMC6477002 | biostudies-literature
Project description:The purpose of this study was to quantitatively evaluate retinal pigment epithelium (RPE) changes in serous pigment epithelial detachment (PED) among patients with age-related macular degeneration by means of prototype multi-contrast optical coherence tomography (OCT), which is capable of simultaneous collection of OCT angiography, polarization-sensitive OCT, and standard OCT images. We evaluated 26 eyes of 21 patients with serous PED. RPE-melanin OCT images were calculated from the multi-contrast OCT dataset and compared with near-infrared autofluorescence images. An active RPE lesion was defined as an area of thickened RPE-melanin (≥ 70 μm; RPE70) on RPE-melanin OCT. Each PED area was divided into peak and slope regions. RPE70 area ratios were compared with the maximum PED height, PED area, PED volume, and slope area ratio (area of slope region/area of whole PED). RPE-melanin OCT images were consistent with near-infrared autofluorescence images. The RPE70 area ratio in the slope region was significantly negatively correlated with the slope area ratio. Development of active RPE lesions in the slope region was correlated with the PED configuration. Multi-contrast OCT is useful for objective evaluation of changes in the RPE in patients with age-related macular degeneration.
| S-EPMC7854605 | biostudies-literature
Project description:Age-related macular degeneration (AMD), the leading cause of blindness in elderly populations, involves the loss of central vision due to progressive dysfunction of the retinal pigment epithelium (RPE) and subsequent loss of light-sensing photoreceptors. While age is a key risk factor, not every aged individual develops AMD. Thus, the critical question is what specific cellular changes tip the balance from healthy aging to disease. To distinguish between changes associated with aging and AMD, we compared the RPE proteome in human eye bank tissue from nondiseased donors during aging (n = 50, 29-91 years) and in donors with AMD (n = 36) compared to age-matched donors without disease (n = 28). Proteins from RPE cells were separated on two-dimensional gels, analyzed for content, and identified using mass spectrometry. A total of 58 proteins displayed significantly altered content with either aging or AMD. Proteins involved in metabolism, protein turnover, stress response, and cell death were altered with both aging and AMD. However, the direction of change was predominantly opposite. With aging, we detected an overall decrease in metabolism and reductions in stress-associated proteins, proteases, and chaperones. With AMD, we observed upregulation of metabolic proteins involved in glycolysis, TCA, and fatty acid metabolism, with a concurrent decline in oxidative phosphorylation, suggesting a reprogramming of energy utilization. Additionally, we detected upregulation of proteins involved in the stress response and protein turnover. Predicted upstream regulators also showed divergent results, with inhibition of inflammation and immune response with aging and activation of these processes with AMD. Our results support the idea that AMD is not simply advanced aging but rather the culmination of perturbed protein homeostasis, defective bioenergetics, and increased oxidative stress within the aging RPE, exacerbated by environmental factors and the genetic background of an individual.
| S-EPMC9683958 | biostudies-literature
Project description:Age-related macular degeneration (AMD) is the leading cause of vision loss in individuals over the age of 65. Histopathological changes become evident in the retinal pigment epithelium (RPE), a monolayer that provides metabolic support for the overlying photoreceptors, even at the earliest stages of AMD that precede vision loss. In a previous global RPE proteome analysis, changes were identified in the content of several mitochondrial proteins associated with AMD. In this study, the subproteome of mitochondria isolated from human donor RPE graded with the Minnesota Grading System (MGS) was analyzed.Human donor eye bank eyes were categorized into one of four progressive stages (MGS 1-4) based on the clinical features of AMD. After dissection of the RPE, mitochondrial proteins were isolated and separated by two-dimensional gel electrophoresis based on their charge and mass. Protein spot densities were compared between the four MGS stages. Peptides from spots that changed significantly with MGS stage were extracted and analyzed by using mass spectrometry to identify the protein.Western blot analyses verified that mitochondria were consistently enriched between MGS stages. The densities of eight spots increased or decreased significantly as a function of MGS stage. These spots were identified as the alpha-, beta-, and delta-ATP synthase subunits, subunit VIb of the cytochrome c oxidase complex, mitofilin, mtHsp70, and the mitochondrial translation factor Tu.The results are consistent with the hypothesis that mitochondrial dysfunction is associated with AMD and further suggest specific pathophysiological mechanisms involving altered mitochondrial translation, import of nuclear-encoded proteins, and ATP synthase activity.
| S-EPMC4397576 | biostudies-literature