Antioxidant Actions of Selenium in Orbital Fibroblasts: A Basis for the Effects of Selenium in Graves’ Orbitopathy

Thyroid ◽  
2017 ◽  
Vol 27 (2) ◽  
pp. 271-278 ◽  
Author(s):  
Giovanna Rotondo Dottore ◽  
Marenza Leo ◽  
Giamberto Casini ◽  
Francesco Latrofa ◽  
Luca Cestari ◽  
...  
Keyword(s):  
Graves Orbitopathy ◽  
Orbital Fibroblasts

Selenium rescues orbital fibroblasts from cell death induced by hydrogen peroxide: another molecular basis for the effects of selenium in graves’ orbitopathy

Endocrine ◽  
2017 ◽  
Vol 58 (2) ◽  
pp. 386-389 ◽  
Author(s):  
Giovanna Rotondo Dottore ◽  
Riccardo Chiarini ◽  
Maria De Gregorio ◽  
Marenza Leo ◽  
Giamberto Casini ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Death ◽  
Molecular Basis ◽  
Graves Orbitopathy ◽  
Orbital Fibroblasts

Erratum to: Selenium rescues orbital fibroblasts from cell death induced by hydrogen peroxide: another molecular basis for the effects of selenium in graves’ orbitopathy

Endocrine ◽  
2017 ◽  
Vol 58 (2) ◽  
pp. 390-390 ◽  
Author(s):  
Giovanna Rotondo Dottore ◽  
Riccardo Chiarini ◽  
Maria De Gregorio ◽  
Marenza Leo ◽  
Giamberto Casini ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Death ◽  
Molecular Basis ◽  
Graves Orbitopathy ◽  
Orbital Fibroblasts

PERK mediates oxidative stress and adipogenesis in Graves’ orbitopathy pathogenesis

2021 ◽  
Author(s):  
JaeSang Ko ◽  
Ji-Young Kim ◽  
Min Kyung Chae ◽  
Eun Jig Lee ◽  
Jin Sook Yoon
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Endoplasmic Reticulum ◽  
Transcription Factor ◽  
Er Stress ◽  
Adipogenic Differentiation ◽  
Ros Generation ◽  
Mrna Levels ◽  
Graves Orbitopathy ◽  
Orbital Fibroblasts

We examined endoplasmic reticulum (ER) stress-related gene expression in orbital tissues from patients with Graves’ orbitopathy (GO) and the effects of silencing protein kinase RNA-like endoplasmic reticulum kinase (PERK) in primary orbital fibroblast cultures to demonstrate the therapeutic potential of PERK-modulating agents in GO management. The expression of ER stress related genes in orbital tissue harvested from individuals with or without GO was studied using real-time polymerase chain reaction. The role of PERK in GO pathogenesis was examined through small-interfering RNA (siRNA)-mediated silencing in cultured primary orbital fibroblasts. Intracellular reactive oxygen species (ROS) levels induced in response to cigarette smoke extract (CSE) or hydrogen peroxide were measured using 5-(and 6)-carboxy-20,70-dichlorodihydrofluorescein diacetate staining and flow cytometry. Cells were stained with Oil Red O, and adipogenesis-related transcription factor expression was evaluated through western blotting after adipogenic differentiation. PERK, activating transcription factor 4 (ATF4), and CCAAT-enhancer-binding protein (C/EBP)-homologous protein(CHOP)mRNA levels were significantly higher in GO orbital tissues than in non-GO orbital tissues. PERK silencing inhibited CSE- or hydrogen peroxide-induced ROS generation. After adipogenic differentiation, GO orbital fibroblasts revealed decreased lipid droplets and downregulation of C/EBPα, C/EBPβ, and peroxisome proliferator-activator gamma (PPARγ) in PERK siRNA-transfected cells. The orbital tissues of patients with GO were exposed to chronic ER stress and subsequently exhibited enhanced unfolded protein response (especially through the PERK pathway). PERK silencing reduced oxidative stress and adipogenesis in GO orbital fibroblasts in vitro. Our results imply that PERK-modulating agents can potentially be used to manage GO.

Wnt signalling inhibits adipogenesis in orbital fibroblasts from patients with Graves’ orbitopathy

2021 ◽  
pp. bjophthalmol-2020-316898
Author(s):  
Sang Joon Jung ◽  
Yeon Jeong Choi ◽  
Tae Kwann Park ◽  
Sang Earn Woo ◽  
Bo-Yeon Kim ◽  
...  
Keyword(s):  
Cyclin D1 ◽  
Adipogenic Differentiation ◽  
Wnt Signalling ◽  
Dependent Manner ◽  
Expression Levels ◽  
Graves Orbitopathy ◽  
Orbital Fibroblast ◽  
Vitro Model ◽  
Orbital Fibroblasts

Background/AimsTo investigate the role of Wnt signalling in adipogenesis using an in vitro model of Graves’ orbitopathy (GO).MethodsOrbital fat was obtained from patients with GO and non-GO participants for primary orbital fibroblast (OF) culture. Expression levels of Wnt5a, Wnt10b, β-catenin, phospho-β-catenin and cyclin D1 were compared between GO and non-GO OFs. These expression levels were also determined during adipogenesis of GO and non-GO OFs. The effects of a stimulator and inhibitor of Wnt signalling on adipogenesis of GO and non-GO OFs were investigated.ResultsWestern blotting analysis showed significant reductions in β-catenin and cyclin D1 and significant enhancement of phospho-β-catenin in OFs from patients with GO, compared with OFs from non-GO participants (p<0.05). Expression of Wnt5a, Wnt10b, β-catenin and cyclin D1 in OFs was highest on day 0, and then gradually declined after induction of adipogenic differentiation. The expression levels of PPARγ, C/EBPα and C/EBPβ were reduced in Wnt stimulator-treated OFs in a dose-dependent manner. Oil red O staining confirmed that a stimulator of Wnt inhibited adipogenesis in GO OFs.ConclusionThese results indicate that Wnt signalling inhibits adipogenesis in OFs from patients with GO and non-GO participants. Further studies are required to examine the potential of Wnt signalling as a target for therapeutic strategies.

scholarly journals Fucoxanthin improves functional recovery of orbitopathy in Graves’ disease by downregulating IL-17 mRNA expression in a mouse model

2020 ◽  
Vol 19 (5) ◽  
pp. 933-941
Author(s):  
Lichao Chai ◽  
Jing Wang ◽  
Yan Wei
Keyword(s):  
Mrna Expression ◽  
Graves Disease ◽  
Oxidative Potential ◽  
Stress Markers ◽  
Tnf Α ◽  
Anti Oxidant ◽  
Graves Orbitopathy ◽  
Orbital Fibroblast ◽  
Complete Cell ◽  
Orbital Fibroblasts

Purpose: To explore the efficacy of fucoxanthin (FX), a carotenoid, against inflammation via inhibition of IL-17 mRNA expression, and its anti-oxidant activity in Graves’ orbitopathy (GO)-induced mice model.Methods: The effects of FX on IL-6, IL-8, IL-17, MCP-1, and TNF-α, in orbital fibroblast tissues extracted from GO-induced BALB/c mice was  investigated. Anti-oxidative stress markers, 8-hydroxy-2’- deoxyguanosine (8-OHdG) and malondialdehyde (MDA) levels were quantified in tear samples collected from GO-induced FX treated mice.Results: FX administration in cultured human orbital fibroblast cells revealed almost complete cell viability and no cell apoptosis. FX resulted in IL-1β induced Beclin-1 and Atg-5 silencing, in cultured human orbital fibroblasts. BALB/c mice immunized with Ad-TSHR289 indicated elevated levels ofthyroid peroxidase and thyroglobulin antibodies in the serum sample. FX predominantly downregulated the mRNA expression of IL-17, and also reduced increased 8-OHdG and MDA in the tear secretion of GO-induced mice.Conclusion: FX may be an effective and useful molecule for the treatment of GO, through its antiinflammatory and anti-oxidative potential, but it requires further investigation to ascertain its therapeutic effectiveness. Keywords: Anti-inflammatory, Anti-oxidant, Fucoxanthin, Graves’ disease, Graves’ orbitopathy, IL-17

Action of three bioavailable antioxidants in orbital fibroblasts from patients with Graves’ orbitopathy (GO): a new frontier for GO treatment?

2017 ◽  
Vol 41 (2) ◽  
pp. 193-201 ◽  
Author(s):  
G. Rotondo Dottore ◽  
I. Ionni ◽  
F. Menconi ◽  
G. Casini ◽  
S. Sellari-Franceschini ◽  
...  
Keyword(s):  
New Frontier ◽  
Graves Orbitopathy ◽  
Orbital Fibroblasts

scholarly journals Novel Roles of Chloroquine and Hydroxychloroquine in Graves’ Orbitopathy Therapy by Targeting Orbital Fibroblasts

2020 ◽  
Vol 105 (6) ◽  
pp. 1906-1917 ◽  
Author(s):  
Yan Guo ◽  
Hai Li ◽  
Xueying Chen ◽  
Huasheng Yang ◽  
Hongyu Guan ◽  
...  
Keyword(s):  
Western Blot ◽  
Real Time ◽  
Fluorescent Protein ◽  
Inhibitory Effect ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Graves Orbitopathy ◽  
Orbital Fibroblasts

Abstract Context Graves’ orbitopathy (GO) causes infiltrative exophthalmos by inducing excessive proliferation, adipogenesis, and glycosaminoglycan production in orbital fibroblasts (OFs). Interference with OF autophagy is a potential therapy for proptosis. Objectives Here, we aimed to evaluate the effects of chloroquine (CQ) and hydroxychloroquine (HCQ), the autophagy inhibitors commonly used in clinical practice, on OFs. Design/Setting/Participants OFs isolated from patients with GO (GO-OFs) or control individuals (non-GO-OFs) were cultured in proliferation medium (PM) or subjected to differentiation medium. OFs were treated with CQ or HCQ (0, 0.5, 2, and 10 μM), and subsequently examined in vitro. Main Outcome Measures CCK-8, EdU incorporation, and flow cytometry assays were used to assess cellular viability. Adipogenesis was assessed with Western blot analysis, real-time polymerase chain reaction (PCR) , and Oil Red O staining. Hyaluronan production was determined by real-time PCR and enzyme-linked immunosorbent assay. Autophagy flux was detected through red fluorescent protein (RFP)-green fluorescent protein (GFP)-LC3 fluorescence staining and Western blot analyses. Results CQ/HCQ halted proliferation and adipogenesis in GO-OFs in a concentration-dependent manner through blockage of autophagy, phenotypes that were not detected in non-GO-OFs. The inhibitory effect of CQ/HCQ on hyaluronan secretion of GO-OFs was also concentration dependent, mediated by downregulation of hyaluronan synthase 2 rather than hyaluronidases. Moreover, CQ (10 μM) induced GO-OF apoptosis without aggravating oxidative stress. Conclusions The antimalarials CQ/HCQ affect proliferation, adipogenesis, and hyaluronan generation in GO-OFs by inhibiting autophagy, providing evidence that they can be used to treat GO as autophagy inhibitors.

Genetic Profiling of Orbital Fibroblasts from Patients with Graves’ Orbitopathy

2021 ◽  
Author(s):  
Giovanna Rotondo Dottore ◽  
Ilaria Bucci ◽  
Giulia Lanzolla ◽  
Iacopo Dallan ◽  
Angela Sframeli ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Cell Proliferation ◽  
Differential Gene Expression ◽  
Global Dna Methylation ◽  
Whole Genome ◽  
Graves Orbitopathy ◽  
Differential Gene ◽  
And Control ◽  
Orbital Fibroblasts

Abstract Context Graves’ orbitopathy (GO) is an autoimmune disease that persists when immunosuppression is achieved. Orbital fibroblasts from GO patients display peculiar phenotypes even if not exposed to autoimmunity, possibly reflecting genetic or epigenetic mechanisms, which we investigated here. Objective We aimed to explore potential genetic or epigenetic differences using primary cultures of orbital fibroblasts from GO and control patients. Methods Cell proliferation, hyaluronic acid (HA) secretion, and HA synthases (HAS) were measured. Next-generation sequencing and gene expression analysis of the whole genome were performed, as well as real-time-PCR of selected genes and global DNA methylation assay on orbital fibroblasts from 6 patients with GO and 6 control patients from a referral center. Results Cell proliferation was higher in GO than in control fibroblasts. Likewise, HA in the cell medium was higher in GO fibroblasts. HAS-1 and HAS-2 did not differ between GO and control fibroblasts, whereas HAS-3 was more expressed in GO fibroblasts. No relevant gene variants were detected by whole-genome sequencing. However, 58 genes were found to be differentially expressed in GO compared with control fibroblasts, and RT-PCR confirmed the findings in 10 selected genes. We postulated that the differential gene expression was related to an epigenetic mechanism, reflecting diverse DNA methylation, which we therefore measured. In support of our hypothesis, global DNA methylation was significantly higher in GO fibroblasts. Conclusions We propose that, following an autoimmune insult, DNA methylation elicits differential gene expression and sustains the maintenance of GO.

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