Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The Silk-protein Sericin Induces Rapid Melanization of Cultured Primary Human Retinal Pigment Epithelial Cells by Activating the NF-κB Pathway.

ABSTRACT: Restoration of the retinal pigment epithelial (RPE) cells to prevent further loss of vision in patients with age-related macular degeneration represents a promising novel treatment modality. Development of RPE transplants, however, requires up to 3 months of cell differentiation. We explored whether the silk protein sericin can induce maturation of primary human retinal pigment epithelial (hRPE) cells. Microarray analysis demonstrated that sericin up-regulated RPE-associated transcripts (RPE65 and CRALBP). Upstream analysis identified the NF-κB pathway as one of the top sericin-induced regulators. ELISA confirmed that sericin stimulates the main NF-κB pathway. Increased levels of RPE-associated proteins (RPE65 and the pigment melanin) in the sericin-supplemented cultures were confirmed by western blot, spectrophotometry and transmission electron microscopy. Sericin also increased cell density and reduced cell death following serum starvation in culture. Inclusion of NF-κB agonists and antagonists in the culture medium showed that activation of the NF-κB pathway appears to be necessary, but not sufficient, for sericin-induced RPE pigmentation. We conclude that sericin promotes pigmentation of cultured primary hRPE cells by activating the main NF-κB pathway. Sericin's potential role in culture protocols for rapid differentiation of hRPE cells derived from embryonic or induced pluripotent stem cells should be investigated.

SUBMITTER: Eidet JR

PROVIDER: S-EPMC4778122 | biostudies-other | 2016 Mar

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

TRAF2 is an NF-κB-activating oncogene in epithelial cancers.

Project description:Aberrant nuclear factor (NF)-κB activation is frequently observed in human cancers. Genome characterization efforts have identified genetic alterations in multiple components of the NF-κB pathway, some of which have been shown to be essential for cancer initiation and tumor maintenance. Here, using patient tumors and cancer cell lines, we identify the NF-κB regulator, TRAF2 (tumor necrosis factor (TNF) receptor-associated factor 2), as an oncogene that is recurrently amplified and rearranged in 15% of human epithelial cancers. Suppression of TRAF2 in cancer cells harboring TRAF2 copy number gain inhibits proliferation, NF-κB activation, anchorage-independent growth and tumorigenesis. Cancer cells that are dependent on TRAF2 also require NF-κB for survival. The phosphorylation of TRAF2 at serine 11 is essential for the survival of cancer cells harboring TRAF2 amplification. Together, these observations identify TRAF2 as a frequently amplified oncogene.

| S-EPMC4067463 | biostudies-literature

Ubiquitin Specific Protease USP48 Destabilizes NF-κB/p65 in Retinal Pigment Epithelium Cells.

Project description:Activation of NF-κB transcription factor is strictly regulated to accurately direct cellular processes including inflammation, immunity, and cell survival. In the retina, the modulation of the NF-κB pathway is essential to prevent excessive inflammatory responses, which plays a pivotal role in many retinal neurodegenerative diseases, such as age-related macular degeneration (AMD), diabetic retinopathy (DR), and inherited retinal dystrophies (IRDs). A critical cytokine mediating inflammatory responses in retinal cells is tumor necrosis factor-alpha (TNFα), leading to the activation of several transductional pathways, including NF-κB. However, the multiple factors orchestrating the appropriate regulation of NF-κB in retinal cells still remain unclear. The present study explores how the ubiquitin-specific protease 48 (USP48) downregulation impacts the stability and transcriptional activity of NF-κB/p65 in retinal pigment epithelium (RPE), at both basal conditions and following TNFα stimulation. We described that USP48 downregulation stabilizes p65. Notably, the accumulation of p65 is mainly detectable in the nuclear compartment and it is accompanied by an increased NF-κB transcriptional activity. These results delineate a novel role of USP48 in negatively regulating NF-κB in retinal cells, providing new opportunities for therapeutic intervention in retinal pathologies.

| S-EPMC9456453 | biostudies-literature

Neurogenin1 effectively reprograms cultured chick retinal pigment epithelial cells to differentiate toward photoreceptors.

Project description:Photoreceptors are highly specialized sensory neurons in the retina, and their degeneration results in blindness. Replacement with developing photoreceptor cells promises to be an effective therapy, but it requires a supply of new photoreceptors, because the neural retina in human eyes lacks regeneration capability. We report efficient generation of differentiating, photoreceptor-like neurons from chick retinal pigment epithelial (RPE) cells propagated in culture through reprogramming with neurogenin1 (ngn1). In reprogrammed culture, a large number of the cells (85.0% +/- 5.9%) began to differentiate toward photoreceptors. Reprogrammed cells expressed transcription factors that set in motion photoreceptor differentiation, including Crx, Nr2E3, NeuroD, and RXRgamma, and phototransduction pathway components, including transducin, cGMP-gated channel, and red opsin of cone photoreceptors (equivalent to rhodopsin of rod photoreceptors). They developed inner segments rich in mitochondria. Furthermore, they responded to light by decreasing their cellular free calcium (Ca(2+)) levels and responded to 9-cis-retinal by increasing their Ca(2+) levels after photobleaching, hallmarks of photoreceptor physiology. The high efficiency and the advanced photoreceptor differentiation indicate ngn1 as a gene of choice to reprogram RPE progeny cells to differentiate into photoreceptor neurons in future cell replacement studies.

| S-EPMC2927132 | biostudies-literature

RIP140-Mediated NF-κB Inflammatory Pathway Promotes Metabolic Dysregulation in Retinal Pigment Epithelium Cells.

Project description:Metabolic dysregulation of the retinal pigment epithelium (RPE) has been implicated in age-related macular degeneration (AMD). However, the molecular regulation of RPE metabolism remains unclear. RIP140 is known to affect oxidative metabolism and mitochondrial biogenesis by negatively controlling mitochondrial pathways regulated by PPAR-γ co-activator-1 α(PGC-1α). This study aims to disclose the effect of RIP140 on the RPE metabolic program in vitro and in vivo. RIP140 protein levels were assayed by Western blotting. Gene expression was tested using quantitative real-time PCR (qRT-PCR), ATP production, and glycogen concentration assays, and the release of inflammatory factors was analyzed by commercial kits. Mice photoreceptor function was measured by electroretinography (ERG). In ARPE-19 cells, RIP140 overexpression changed the expression of the key metabolic genes and lipid processing genes, inhibited mitochondrial ATP production, and enhanced glycogenesis. Moreover, RIP140 overexpression promoted the translocation of NF-κB and increased the expression and production of IL-1β, IL-6, and TNF-α in ARPE-19 cells. Importantly, we also observed the overexpression of RIP140 through adenovirus delivery in rat retinal cells, which significantly decreased the amplitude of the a-wave and b-wave measured by ERG assay. Therapeutic strategies that modulate the activity of RIP140 could have clinical utility for the treatment of AMD in terms of preventing RPE degeneration.

| S-EPMC9689634 | biostudies-literature

Oxidative stress induces ferroptotic cell death in retinal pigment epithelial cells.

Project description:The dysfunction and cell death of retinal pigment epithelial (RPE) cells are hallmarks of late-stage dry (atrophic) age-related macular degeneration (AMD), for which no effective therapy has yet been developed. Previous studies have indicated that iron accumulation is a source of excess free radical production in RPE, and age-dependent iron accumulation in RPE is accelerated in patients with dry AMD. Although the pathogenic role of oxidative stress in RPE in the development of dry AMD is widely accepted, the mechanisms of oxidative stress-induced RPE cell death remain elusive. Here, we show that ferroptotic cell death, a mode of regulated necrosis mediated by iron and lipid peroxidation, is implicated in oxidative stress-induced RPE cell death in vitro. In ARPE-19 cells we observed that the ferroptosis inhibitors ferrostatin-1 and deferoxamine (DFO) rescued tert-butyl hydroperoxide (tBH)-induced RPE cell death more effectively than inhibitors of apoptosis or necroptosis. tBH-induced RPE cell death was accompanied by the three characteristics of ferroptotic cell death: lipid peroxidation, glutathione depletion, and ferrous iron accumulation, which were all significantly attenuated by ferrostatin-1 and DFO. Exogenous iron overload enhanced tBH-induced RPE cell death, but this effect was also attenuated by ferrostatin-1 and DFO. Furthermore, mRNA levels of numerous genes known to regulate iron metabolism were observed to be influenced by oxidative stress. Taken together, our observations suggest that multiple modes of cell death are involved in oxidative stress-induced RPE cell death, with ferroptosis playing a particularly important role.

| S-EPMC7418497 | biostudies-literature

Improvement of Storage Medium for Cultured Human Retinal Pigment Epithelial Cells Using Factorial Design.

Project description:Storage of human retinal pigment epithelium (hRPE) can contribute to the advancement of cell-based RPE replacement therapies. The present study aimed to improve the quality of stored hRPE cultures by identifying storage medium additives that, alone or in combination, contribute to enhancing cell viability while preserving morphology and phenotype. hRPE cells were cultured in the presence of the silk protein sericin until pigmentation. Cells were then stored for 10 days in storage medium plus sericin and either one of 46 different additives. Individual effects of each additive on cell viability were assessed using epifluorescence microscopy. Factorial design identified promising additive combinations by extrapolating their individual effects. Supplementing the storage medium with sericin combined with adenosine, L-ascorbic acid and allopurinol resulted in the highest cell viability (98.6 ± 0.5%) after storage for three days, as measured by epifluorescence microscopy. Flow cytometry validated the findings. Proteomics identified 61 upregulated and 65 downregulated proteins in this storage group compared to the unstored control. Transmission electron microscopy demonstrated the presence of melanosomes after storage in the optimized medium. We conclude that the combination of adenosine, L-ascorbic acid, allopurinol and sericin in minimal essential medium preserves RPE pigmentation while maintaining cell viability during storage.

| S-EPMC5890280 | biostudies-literature

Expression of R345W-Fibulin-3 Induces Epithelial-Mesenchymal Transition in Retinal Pigment Epithelial Cells.

Project description:Purpose:To investigate the role of protein misfolding in retinal pigment epithelial (RPE) cell dysfunction, the effects of R345W-Fibulin-3 expression on RPE cell phenotype were studied. Methods:Primary RPE cells were cultured to confluence on Transwells and infected with lentivirus constructs to express wild-type (WT)- or R345W-Fibulin-3. Barrier function was assessed by evaluating zonula occludens-1 (ZO-1) distribution and trans-epithelial electrical resistance (TER). Polarized secretion of vascular endothelial growth factor (VEGF), was measured by Enzyme-linked immunosorbent assay (ELISA). Differentiation status was assessed by qPCR of genes known to be preferentially expressed in terminally differentiated RPE cells, and conversion to an epithelial-mesenchymal transition (EMT) phenotype was assessed by a migration assay. Results:Compared to RPE cells expressing WT-Fibulin-3, ZO-1 distribution was disrupted and TER values were significantly lower in RPE cells expressing R345W-Fibulin-3. In cells expressing mutant Fibulin-3, VEGF secretion was attenuated basally but not in the apical direction, whereas Fibulin-3 secretion was reduced in both the apical and basal directions. Retinal pigment epithelial signature genes were downregulated and multiple genes associated with EMT were upregulated in the mutant group. Migration assays revealed a faster recovery rate in ARPE-19 cells overexpressing R345W-Fibulin-3 compared to WT. Conclusions:The results suggest that expression of R345W-Fibulin-3 promotes EMT in RPE cells.

| S-EPMC7317295 | biostudies-literature

Human Norovirus Induces Aquaporin 1 Production by Activating NF-κB Signaling Pathway.

Project description:Human norovirus (HuNoV) is one of the major pathogens of acute nonbacterial gastroenteritis. Due to the lack of a robust and reproducible in vitro culture system and an appropriate animal model, the mechanism underlying HuNoV-caused diarrhea remains unknown. In the current study, we found that HuNoV transfection induced the expression of aquaporin 1 (AQP1), which was further confirmed in the context of virus infection, whereas the enterovirus EV71 (enterovirus 71) did not have such an effect. We further revealed that VP1, the major capsid protein of HuNoV, was crucial in promoting AQP1 expression. Mechanistically, HuNoV induces AQP1 production through the NF-κB signaling pathway via inducing the expression, phosphorylation and nuclear translocation of p65. By using a model of human intestinal epithelial barrier (IEB), we demonstrated that HuNoV and VP1-mediated enhancement of small molecule permeability is associated with the AQP1 channel. Collectively, we revealed that HuNoV induced the production of AQP1 by activating the NF-κB signaling pathway. The findings in this study provide a basis for further understanding the significance of HuNoV-induced AQP1 expression and the potential mechanism underlying HuNoV-caused diarrhea.

| S-EPMC9028284 | biostudies-literature

Spermidine Attenuates High Glucose-Induced Oxidative Damage in Retinal Pigment Epithelial Cells by Inhibiting Production of ROS and NF-κB/NLRP3 Inflammasome Pathway.

Project description: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.

| S-EPMC10341649 | biostudies-literature

The expression of native and cultured human retinal pigment epithelial cells grown in different culture conditions.

Project description:AimTo determine the transcriptional proximity of retinal pigment epithelium (RPE) cells grown under different culture conditions and native RPE.MethodsARPE-19 cells were grown under five conditions in 10% CO(2): "subconfluent" in DMEM/F12+10% FBS, "confluent" in serum and serum withdrawn, and "differentiated" for 2.5 months in serum and serum withdrawn medium. Native RPE was laser microdissected. Total RNA was extracted, reverse transcribed, and radiolabelled probes were hybridised to an array containing 5,353 genes. Arrays were evaluated by hierarchical cluster analysis and significance analysis of microarrays.Results78% of genes were expressed by native RPE while 45.3--47.7% were expressed by ARPE-19 cells, depending on culture condition. While the most abundant genes were expressed by native and cultured cells, significant differences in low abundance genes were seen. Hierarchical cluster analysis showed that confluent and differentiated, serum withdrawn cultures clustered closest to native RPE, and that serum segregated cultured cells from native RPE. The number of differentially expressed genes and their function, and profile of expressed and unexpressed genes, demonstrate differences between native and cultured cells.ConclusionsWhile ARPE-19 cells have significant value for studying RPE behaviour, investigators must be aware of how culture conditions can influence the mRNA phenotype of the cell.

| S-EPMC1772910 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data