scholarly journals Drusen in patient-derived hiPSC-RPE models of macular dystrophies

2017 ◽  
Vol 114 (39) ◽  
pp. E8214-E8223 ◽  
Author(s):  
Chad A. Galloway ◽  
Sonal Dalvi ◽  
Sandy S. C. Hung ◽  
Leslie A. MacDonald ◽  
Lisa R. Latchney ◽  
...  
Keyword(s):  
Vision Loss ◽  
Retinal Pigment ◽  
Retinal Dystrophy ◽  
Age Related Macular Degeneration ◽  
In Vivo Studies ◽  
Specific Cell ◽  
Rpe Cells ◽  
Age Related ◽  
Pathway Gene ◽  
Human Pathology

Age-related macular degeneration (AMD) and related macular dystrophies (MDs) are a major cause of vision loss. However, the mechanisms underlying their progression remain ill-defined. This is partly due to the lack of disease models recapitulating the human pathology. Furthermore, in vivo studies have yielded limited understanding of the role of specific cell types in the eye vs. systemic influences (e.g., serum) on the disease pathology. Here, we use human induced pluripotent stem cell-retinal pigment epithelium (hiPSC-RPE) derived from patients with three dominant MDs, Sorsby’s fundus dystrophy (SFD), Doyne honeycomb retinal dystrophy/malattia Leventinese (DHRD), and autosomal dominant radial drusen (ADRD), and demonstrate that dysfunction of RPE cells alone is sufficient for the initiation of sub-RPE lipoproteinaceous deposit (drusen) formation and extracellular matrix (ECM) alteration in these diseases. Consistent with clinical studies, sub-RPE basal deposits were present beneath both control (unaffected) and patient hiPSC-RPE cells. Importantly basal deposits in patient hiPSC-RPE cultures were more abundant and displayed a lipid- and protein-rich “drusen-like” composition. Furthermore, increased accumulation of COL4 was observed in ECM isolated from control vs. patient hiPSC-RPE cultures. Interestingly, RPE-specific up-regulation in the expression of several complement genes was also seen in patient hiPSC-RPE cultures of all three MDs (SFD, DHRD, and ADRD). Finally, although serum exposure was not necessary for drusen formation, COL4 accumulation in ECM, and complement pathway gene alteration, it impacted the composition of drusen-like deposits in patient hiPSC-RPE cultures. Together, the drusen model(s) of MDs described here provide fundamental insights into the unique biology of maculopathies affecting the RPE–ECM interface.

scholarly journals PGC-1α Protects RPE Cells of the Aging Retina against Oxidative Stress-Induced Degeneration through the Regulation of Senescence and Mitochondrial Quality Control. The Significance for AMD Pathogenesis

2018 ◽  
Vol 19 (8) ◽  
pp. 2317 ◽  
Author(s):  
Kai Kaarniranta ◽  
Jakub Kajdanek ◽  
Jan Morawiec ◽  
Elzbieta Pawlowska ◽  
Janusz Blasiak
Keyword(s):  
Oxidative Stress ◽  
Cellular Senescence ◽  
Mitochondrial Biogenesis ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Sirtuin 1 ◽  
Age Related Macular Degeneration ◽  
Rpe Cells ◽  
Age Related

PGC-1α (peroxisome proliferator-activated receptor gamma coactivator 1-alpha) is a transcriptional coactivator of many genes involved in energy management and mitochondrial biogenesis. PGC-1α expression is associated with cellular senescence, organismal aging, and many age-related diseases, including AMD (age-related macular degeneration), an important global issue concerning vision loss. We and others have developed a model of AMD pathogenesis, in which stress-induced senescence of retinal pigment epithelium (RPE) cells leads to AMD-related pathological changes. PGC-1α can decrease oxidative stress, a key factor of AMD pathogenesis related to senescence, through upregulation of antioxidant enzymes and DNA damage response. PGC-1α is an important regulator of VEGF (vascular endothelial growth factor), which is targeted in the therapy of wet AMD, the most devastating form of AMD. Dysfunction of mitochondria induces cellular senescence associated with AMD pathogenesis. PGC-1α can improve mitochondrial biogenesis and negatively regulate senescence, although this function of PGC-1α in AMD needs further studies. Post-translational modifications of PGC-1α by AMPK (AMP kinase) and SIRT1 (sirtuin 1) are crucial for its activation and important in AMD pathogenesis.

scholarly journals Release of Iron-Loaded Ferritin in Sodium Iodate-Induced Model of Age Related Macular Degeneration: An In-Vitro and In-Vivo Study

Antioxidants ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1253
Author(s):  
Ajay Ashok ◽  
Suman Chaudhary ◽  
Aaron S. Wise ◽  
Neil A. Rana ◽  
Dallas McDonald ◽  
...  
Keyword(s):  
Macular Degeneration ◽  
Retinal Pigment ◽  
Age Related Macular Degeneration ◽  
In Vivo Studies ◽  
Epithelial Cell Line ◽  
Sodium Iodate ◽  
Rpe Cells ◽  
Age Related

To evaluate the role of iron in sodium iodate (NaIO3)-induced model of age-related macular degeneration (AMD) in ARPE-19 cells in-vitro and in mouse models in-vivo. ARPE-19 cells, a human retinal pigment epithelial cell line, was exposed to 10 mM NaIO3 for 24 h, and the expression and localization of major iron modulating proteins was evaluated by Western blotting (WB) and immunostaining. Synthesis and maturation of cathepsin-D (cat-D), a lysosomal enzyme, was evaluated by quantitative reverse-transcriptase polymerase chain reaction (RT-qPCR) and WB, respectively. For in-vivo studies, C57BL/6 mice were injected with 40 mg/kg mouse body weight of NaIO3 intraperitoneally, and their retina was evaluated after 3 weeks as above. NaIO3 induced a 10-fold increase in ferritin in ARPE-19 cells, which co-localized with LC3II, an autophagosomal marker, and LAMP-1, a lysosomal marker. A similar increase in ferritin was noted in retinal lysates and retinal sections of NaIO3-injected mice by WB and immunostaining. Impaired synthesis and maturation of cat-D was also noted. Accumulated ferritin was loaded with iron, and released from retinal pigmented epithelial (RPE) cells in Perls’ and LAMP-1 positive vesicles. NaIO3 impairs lysosomal degradation of ferritin by decreasing the transcription and maturation of cat-D in RPE cells. Iron-loaded ferritin accumulates in lysosomes and is released in lysosomal membrane-enclosed vesicles to the extracellular milieu. Accumulation of ferritin in RPE cells and fusion of ferritin-containing vesicles with adjacent photoreceptor cells is likely to create an iron overload, compromising their viability. Moreover, reduced activity of cat-D is likely to promote accumulation of other cellular debris in lysosomal vesicles, contributing to AMD-like pathology.

scholarly journals Implication of N-methyl-D-aspartate receptor In Homocysteine Induced Age related Macular Degeneration

2021 ◽  
Author(s):  
Yara A. Samra ◽  
Dina Kira ◽  
Pragya Rajpurohit ◽  
Riyaz Mohamed ◽  
Leah Owen ◽  
...  
Keyword(s):  
Macular Degeneration ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Age Related Macular Degeneration ◽  
Mice Model ◽  
Rpe Cells ◽  
Aspartate Receptor ◽  
Age Related ◽  
Retinal Structure

Abstract Background: Age related macular degeneration (AMD) is a leading cause of vision loss in old people. Elevated homocysteine (Hcy), known as Hyperhomocysteinemia (HHcy) was reported in association with AMD. We previously reported that HHcy induces AMD like features. The current study suggests activation of N-Methyl-D-aspartate receptor (NMDAR) in retinal pigment epithelium (RPE) cells as a mechanism for HHcy-induced AMD. Serum Hcy and cystathione-β-synthase enzyme (CBS) were assessed by ELISA in AMD patients. The involvement of NMDAR in Hcy’s induced AMD features were evaluated 1)-In-vitro using ARPE-19 cells, primary RPE isolated from mice model of HHcy (CBS) and mouse choroidal endothelial cells (MCEC). 2)-In-vivo using wild type mice and mice deficient in RPE cells NMDAR (NMDARR -/-) with/without intravitreal injection of Hcy. Expression of retinal isolectin-B4, Ki67, HIF-1α, VEGF, NMDAR1 and albumin were assessed by immunofluorescence (IF), Western blot (WB), Optical coherence tomography (OCT), and fluorescein angiography (FA) to evaluate retinal structure, fluorescein leakage and development of choroidal neovascularization (CNV) in living mice. Results: Serum of the neovascular AMD patients showed significant increase in Hcy and decrease in CBS levels. Moreover, Hcy significantly increased angiogenic markers; HIF-1α, VEGF and NMDAR in RPE cells and Ki67 in MCEC. Hcy-injected WT mice showed disrupted retinal morphology and development of CNV. Knocking down NMDAR in RPE improved retinal structure and CNV induction.Conclusion: Our findings underscore the potential role for NMDAR in RPE cells in mediating Hcy-induced features of AMD and CNV induction, thus NMDAR inhibition could provide a promising therapeutic target for AMD.

scholarly journals CFH Loss in Human RPE Cells Leads to Inflammation and Complement System Dysregulation via the NF-κB Pathway

2021 ◽  
Vol 22 (16) ◽  
pp. 8727
Author(s):  
Angela Armento ◽  
Tiziana L. Schmidt ◽  
Inga Sonntag ◽  
David A. Merle ◽  
Mohamed Ali Jarboui ◽  
...  
Keyword(s):  
Complement System ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Factor H ◽  
Age Related Macular Degeneration ◽  
Complement Factor ◽  
Major Pathway ◽  
Rpe Cells ◽  
Age Related

Age-related macular degeneration (AMD), the leading cause of vision loss in the elderly, is a degenerative disease of the macula, where retinal pigment epithelium (RPE) cells are damaged in the early stages of the disease, and chronic inflammatory processes may be involved. Besides aging and lifestyle factors as drivers of AMD, a strong genetic association to AMD is found in genes of the complement system, with a single polymorphism in the complement factor H gene (CFH), accounting for the majority of AMD risk. However, the exact mechanism of CFH dysregulation confers such a great risk for AMD and its role in RPE cell homeostasis is unclear. To explore the role of endogenous CFH locally in RPE cells, we silenced CFH in human hTERT-RPE1 cells. We demonstrate that endogenously expressed CFH in RPE cells modulates inflammatory cytokine production and complement regulation, independent of external complement sources, or stressors. We show that loss of the factor H protein (FH) results in increased levels of inflammatory mediators (e.g., IL-6, IL-8, GM-CSF) and altered levels of complement proteins (e.g., C3, CFB upregulation, and C5 downregulation) that are known to play a role in AMD. Moreover, our results identify the NF-κB pathway as the major pathway involved in regulating these inflammatory and complement factors. Our findings suggest that in RPE cells, FH and the NF-κB pathway work in synergy to maintain inflammatory and complement balance, and in case either one of them is dysregulated, the RPE microenvironment changes towards a proinflammatory AMD-like phenotype.

scholarly journals A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD

2021 ◽  
Vol 22 (21) ◽  
pp. 11979
Author(s):  
Peng Shang ◽  
Nadezda A. Stepicheva ◽  
Haitao Liu ◽  
Olivia Chowdhury ◽  
Jonathan Franks ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Explant Culture ◽  
Age Related Macular Degeneration ◽  
Rpe Cells ◽  
Age Related

Degeneration of retinal pigment epithelium (RPE) is one of the most critical phenotypic changes of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly. While cultured polarized RPE cells with original properties are valuable in in vitro models to study RPE biology and the consequences of genetic and/or pharmacological manipulations, the procedure to establish mouse primary PRE cell culture or pluripotent stem cell-derived RPE cells is time-consuming and yields a limited number of cells. Thus, establishing a mouse in situ RPE culture system is highly desirable. Here we describe a novel and efficient method for RPE explant culture that allows for obtaining biologically relevant RPE cells in situ. These RPE explants (herein referred to as RPE flatmounts) are viable in culture for at least 7 days, can be efficiently transduced with adenoviral constructs, and/or treated with a variety of drugs/chemicals followed by downstream analysis of the signaling pathways/biological processes of interest, such as assessment of the autophagy flux, inflammatory response, and receptor tyrosine kinases stimulation. This method of RPE explant culture is highly beneficial for pharmacological and mechanistic studies in the field of RPE biology and AMD research.

scholarly journals CFH loss in human RPE cells leads to inflammation and complement system dysregulation via the NF-ƙB pathway

2021 ◽  
Author(s):  
Angela Armento ◽  
Tiziana Luisa Schmidt ◽  
Inga Sonntag ◽  
David Merle ◽  
Mohamed-ali Jarboui ◽  
...  
Keyword(s):  
Complement System ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Factor H ◽  
Age Related Macular Degeneration ◽  
Complement Factor ◽  
Major Pathway ◽  
Rpe Cells ◽  
Age Related

Age-related macular degeneration (AMD), the leading cause of vision loss in the elderly, is a degenerative disease of the macula, where retinal pigment epithelium (RPE) cells are damaged in the early stages of the disease and chronic inflammatory processes may be involved. Besides ageing and lifestyle factors as drivers of AMD, a strong genetic association to AMD is found in genes of the complement system, with a single polymorphism in the complement factor H gene (CFH), accounting for the majority of AMD risk. However, the exact mechanism by which CFH dysregulation confers such a great risk for AMD and its role in RPE cells homeostasis is unclear. To explore the role of endogenous CFH locally in RPE cells, we silenced CFH in human hTERT-RPE1 cells. We demonstrate that endogenously expressed CFH in RPE cells modulates inflammatory cytokine production and complement regulation, independent of external complement sources or stressors. We show that loss of the factor H protein (FH) results in increased levels of inflammatory mediators (e.g. IL-6, IL-8, GM-CSF) and altered levels of complement proteins (e.g. C3, CFB upregulation and C5 downregulation) that are known to play a role in AMD. Moreover, we identified the NF-ƙB pathway as the major pathway involved in the regulation of these inflammatory and complement factors. Our findings suggest that in RPE cells, FH and the NF-ƙB pathway work in synergy to maintain inflammatory and complement balance and in case either one of them is dysregulated, the RPE microenvironment changes towards a pro-inflammatory AMD-like phenotype.

scholarly journals Scaffold-Free Retinal Pigment Epithelium Microtissues Exhibit Increased Release of PEDF

2021 ◽  
Vol 22 (21) ◽  
pp. 11317
Author(s):  
Abdullah Al-Ani ◽  
Derek Toms ◽  
Saud Sunba ◽  
Kayla Giles ◽  
Yacine Touahri ◽  
...  
Keyword(s):  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Critical Role ◽  
Age Related Macular Degeneration ◽  
Rpe Cells ◽  
Age Related ◽  
Wide Range ◽  
And Function

The retinal pigmented epithelium (RPE) plays a critical role in photoreceptor survival and function. RPE deficits are implicated in a wide range of diseases that result in vision loss, including age-related macular degeneration (AMD) and Stargardt disease, affecting millions worldwide. Subretinal delivery of RPE cells is considered a promising avenue for treatment, and encouraging results from animal trials have supported recent progression into the clinic. However, the limited survival and engraftment of transplanted RPE cells delivered as a suspension continues to be a major challenge. While RPE delivery as epithelial sheets exhibits improved outcomes, this comes at the price of increased complexity at both the production and transplant stages. In order to combine the benefits of both approaches, we have developed size-controlled, scaffold-free RPE microtissues (RPE-µTs) that are suitable for scalable production and delivery via injection. RPE-µTs retain key RPE molecular markers, and interestingly, in comparison to conventional monolayer cultures, they show significant increases in the transcription and secretion of pigment-epithelium-derived factor (PEDF), which is a key trophic factor known to enhance the survival and function of photoreceptors. Furthermore, these microtissues readily spread in vitro on a substrate analogous to Bruch’s membrane, suggesting that RPE-µTs may collapse into a sheet upon transplantation. We anticipate that this approach may provide an alternative cell delivery system to improve the survival and integration of RPE transplants, while also retaining the benefits of low complexity in production and delivery.

scholarly journals Chrysin Alleviates DNA Damage to Improve Disturbed Immuno-Homeostasis and Pro-Angiogenic Environment in Laser-Induced Choroidal Neovascularization

2021 ◽  
Author(s):  
Jing Wang ◽  
Qiyu Bo ◽  
Minwen Zhou ◽  
Hong Wang ◽  
Hong Zhu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Choroidal Neovascularization ◽  
Vision Loss ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Age Related Macular Degeneration ◽  
Cell Viability Assay ◽  
Bioinformatics Analyses ◽  
Rpe Cells ◽  
Age Related

Abstract Background: Choroidal neovascularization (CNV) is a devastating pathology of numerous ocular diseases, such as wet age-related macular degeneration (wAMD), which causes irreversible vision loss. Although anti-VEGF therapy has been widely used, poor response or no response exits in some patients, suggesting that some other important angiogenic components play roles. Therefore, the underlined mechanism need to be clarified and new target of anti-angiogenic therapy is urgently needed. Damaged retinal pigment epithelium (RPE) cells have been demonstrated to activate inflammasome, drive a degenerative tissue environment and an enhanced pro-angiogenic response, which emphasizes the dysfunction of RPE, may be the hallmark of the pathogenesis.Methods: C57BL/6J male mice aged between 6 and 8 weeks were subjected to laser-induced CNV models. Chrysin was administered intragastrically at 25 mg/kg daily for 3 days or one week after laser-treated. Then to observe the CNV areas and CNV thickness, immunofluorescence staining of choroidal flatmount, SD-OCT and fluorescein angiograghy were performed, respectively. To further confirm the effect of chrysin on stress-induced DNA damage in RPE cells, RPE cells were administered with A2E and western-blot, cell viability assay, immunofluorescence chromosome PNA-FISH and SA-β-gal staining were performed. To elucidate the underlying mechanism, we performed RNA-seq and bioinformatics analyses.Results: In this study, we demonstrated that chrysin could successfully alleviated choroidal neovascularization. We show that DNA damage of RPE cells is remarkable in laser-induced choroidal neovascularization, resulting in inflammation response, which can be ameliorated by chrysin through inactivation of STAT3. Also, we identify that chrysin can reduce DNA damage, especially telomere erosion, simultaneously compromise the dysfunction of RPE and the secretion of SASP factor in vitro. Mechanistically, KEGG pathway analyzes show that chrsyin improves inflammatory imbalance mainly through down-regulation of IL17 pathway in the laser- induced CNV development.Conclusions: Our results indicate the interplay between DNA damage, perturbed RPE homeostasis, inflammatory imbalance and angiogenesis in laser-induced choroidal neovascularization. Importantly, chrysin may be an effective therapeutic supplement for CNV.

scholarly journals Potential of Telomerase in Age-Related Macular Degeneration—Involvement of Senescence, DNA Damage Response and Autophagy and a Key Role of PGC-1α

2021 ◽  
Vol 22 (13) ◽  
pp. 7194
Author(s):  
Janusz Blasiak ◽  
Joanna Szczepanska ◽  
Michal Fila ◽  
Elzbieta Pawlowska ◽  
Kai Kaarniranta
Keyword(s):  
Dna Damage ◽  
Macular Degeneration ◽  
Dna Damage Response ◽  
Vision Loss ◽  
Retinal Pigment ◽  
Ectopic Expression ◽  
Age Related Macular Degeneration ◽  
Rpe Cells ◽  
Age Related ◽  
Damage Response

Age-related macular degeneration (AMD), the main cause of vision loss in the elderly, is associated with oxidation in the retina cells promoting telomere attrition. Activation of telomerase was reported to improve macular functions in AMD patients. The catalytic subunit of human telomerase (hTERT) may directly interact with proteins important for senescence, DNA damage response, and autophagy, which are impaired in AMD. hTERT interaction with mTORC1 (mTOR (mechanistic target of rapamycin) complex 1) and PINK1 (PTEN-induced kinase 1) activates macroautophagy and mitophagy, respectively, and removes cellular debris accumulated over AMD progression. Ectopic expression of telomerase in retinal pigment epithelium (RPE) cells lengthened telomeres, reduced senescence, and extended their lifespan. These effects provide evidence for the potential of telomerase in AMD therapy. Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) may be involved in AMD pathogenesis through decreasing oxidative stress and senescence, regulation of vascular endothelial growth factor (VEGF), and improving autophagy. PGC-1α and TERT form an inhibitory positive feedback loop. In conclusion, telomerase activation and its ectopic expression in RPE cells, as well as controlled clinical trials on the effects of telomerase activation in AMD patients, are justified and should be assisted by PGC-1α modulators to increase the therapeutic potential of telomerase in AMD.

scholarly journals Role of amyloid β-peptide in the pathogenesis of age-related macular degeneration

2021 ◽  
Vol 6 (1) ◽  
pp. e000774
Author(s):  
Minwei Wang ◽  
Shiqi Su ◽  
Shaoyun Jiang ◽  
Xinghuai Sun ◽  
Jiantao Wang
Keyword(s):  
Mitochondrial Dysfunction ◽  
Macular Degeneration ◽  
Vision Loss ◽  
Cell Structure ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Amyloid Β ◽  
Age Related Macular Degeneration ◽  
Amyloid Β Peptide ◽  
Age Related

Age-related macular degeneration (AMD) is the most common eye disease in elderly patients, which could lead to irreversible vision loss and blindness. Increasing evidence indicates that amyloid β-peptide (Aβ) might be associated with the pathogenesis of AMD. In this review, we would like to summarise the current findings in this field. The literature search was done from 1995 to Feb, 2021 with following keywords, ‘Amyloid β-peptide and age-related macular degeneration’, ‘Inflammation and age-related macular degeneration’, ‘Angiogenesis and age-related macular degeneration’, ‘Actin cytoskeleton and amyloid β-peptide’, ‘Mitochondrial dysfunction and amyloid β-peptide’, ‘Ribosomal dysregulation and amyloid β-peptide’ using search engines Pubmed, Google Scholar and Web of Science. Aβ congregates in subretinal drusen of patients with AMD and participates in the pathogenesis of AMD through enhancing inflammatory activity, inducing mitochondrial dysfunction, altering ribosomal function, regulating the lysosomal pathway, affecting RNA splicing, modulating angiogenesis and modifying cell structure in AMD. The methods targeting Aβ are shown to inhibit inflammatory signalling pathway and restore the function of retinal pigment epithelium cells and photoreceptor cells in the subretinal region. Targeting Aβ may provide a novel therapeutic strategy for AMD.

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