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Müller glial cells inhibit proliferation of retinal endothelial cells via TGF-β2 and Smad signaling.Glia Sep 2014Neovascularization is a sight-threatening complication of ischemic proliferative retinopathies. Transforming growth factor (TGF)-β, a cytokine with multiple functions...
Neovascularization is a sight-threatening complication of ischemic proliferative retinopathies. Transforming growth factor (TGF)-β, a cytokine with multiple functions in the retina, participates in the control of pathological angiogenesis and neovascularization. Retinal glial (Müller) cells produce TGF-β2 under physiological and post-ischemic conditions. To characterize glial cell-derived mediators of angiogenesis regulation in glial-endothelial interactions in the retina, we co-cultured primary Müller cells and bovine microvascular retinal endothelial cells (BRECs). Müller cell-derived TGF-β2 was bound by the BRECs, which were found to express serine/threonine kinase TGF-β receptors, and stimulated TGF-β-dependent anti-proliferative signaling pathways. The proliferation of BRECs was attenuated by exogenous TGF-β2 as well as by the presence of Müller cell culture media. The following intracellular signaling mechanisms were found to be involved in the anti-angiogenic action of Müller cell-derived TGF-β2: (i) binding of TGF-β2 to BRECs is mediated by the type-II TGF-β receptor, leading to (ii) activation and phosphorylation of receptor-activated Smads; (iii) Müller cell-derived TGF-β2 activates Smad2 and Smad3 to (iv) attenuate the phosphorylation state of the MAP kinases, extracellular signal-regulated kinase (ERK)-1/-2. Neutralizing TGF-β or TGF-β type-II receptor or blocking the activation of Smads partially abrogated the effect of Müller cell-conditioned media on BRECs. Together, our data suggest that Müller cells release TGF-β2, inhibiting the proliferation of retinal endothelial cells via activation of Smad2/Smad3 and attenuation of ERK signaling. Given the context-dependent action of TGF-β2 on angiogenesis, our results may have implications for understanding the pathogenesis of retinal angiopathies, such as diabetic retinopathy, and the anti-angiogenic role of TGF-β therein.
Topics: Animals; Cattle; Cell Hypoxia; Cell Proliferation; Cells, Cultured; Coculture Techniques; Endothelial Cells; Ependymoglial Cells; Guinea Pigs; Humans; MAP Kinase Signaling System; Phosphorylation; Rats, Long-Evans; Recombinant Proteins; Retina; Smad Proteins; Transforming Growth Factor beta2; Vascular Endothelial Growth Factor A
PubMed: 24838936
DOI: 10.1002/glia.22694 -
Scientific Reports May 2021Administration of the mitochondrial complex I inhibitor rotenone provides an excellent model to study the pathomechanism of oxidative stress-related neural degeneration...
Administration of the mitochondrial complex I inhibitor rotenone provides an excellent model to study the pathomechanism of oxidative stress-related neural degeneration diseases. In this study, we examined the glial roles in retinal cell survival and degeneration under the rotenone-induced oxidative stress condition. Mouse-derived Müller, microglial (BV-2), and dissociated retinal cells were used for in vitro experiments. Gene expression levels and cell viability were determined using quantitative reverse transcription-polymerase chain reaction and the alamarBlue assay, respectively. Conditioned media were prepared by stimulating glial cells with rotenone. Retinal ganglion cells (RGCs) and inner nuclear layer (INL) were visualized on rat retinal sections by immunohistochemistry and eosin/hematoxylin, respectively. Rotenone dose-dependently induced glial cell death. Treatment with rotenone or rotenone-stimulated glial cell-conditioned media altered gene expression of growth factors and inflammatory cytokines in glial cells. The viability of dissociated retinal cells significantly increased upon culturing in media conditioned with rotenone-stimulated or Müller cell-conditioned media-stimulated BV-2 cells. Furthermore, intravitreal neurotrophin-5 administration prevented the rotenone-induced reduction of RGC number and INL thickness in rats. Thus, glial cells exerted both positive and negative effects on retinal cell survival in rotenone-induced neural degeneration via altered expression of growth factors, especially upregulation of microglia-derived Ntf5, and proinflammatory cytokines.
Topics: Animals; Cell Survival; Culture Media, Conditioned; Cytokines; Ependymoglial Cells; Intercellular Signaling Peptides and Proteins; Microglia; Nerve Degeneration; Oxidative Stress; Rats; Retinal Ganglion Cells; Rotenone
PubMed: 34045544
DOI: 10.1038/s41598-021-90604-w -
Visual Neuroscience Jul 2021The electroretinogram (ERG) has been employed for years to collect information about retinal function and pathology. The usefulness of this noninvasive test depends on...
The electroretinogram (ERG) has been employed for years to collect information about retinal function and pathology. The usefulness of this noninvasive test depends on our understanding of the cell sources that generate the ERG. Important contributors to the ERG are glial Müller cells (MCs), which are capable of generating substantial transretinal potentials in response to light-induced changes in extracellular K+ concentration ([K+]o). For instance, the MCs generate the slow PIII (sPIII) component of the ERG as a reaction to a photoreceptor-induced [K+]o decrease in the subretinal space. Similarly, an increase of [K+]o related to activity of postreceptor retinal neurons also produces transretinal glial currents, which can potentially influence the amplitude and shape of the b-wave, one of the most frequently analyzed ERG components. Although it is well documented that the majority of the b-wave originates from On-bipolar cells, some contribution from MCs was suggested many years ago and has never been experimentally rejected. In this work, detailed information about light-evoked [K+]o changes in the isolated mouse retina was collected and then analyzed with a relatively simple linear electrical model of MCs. The results demonstrate that the cornea-positive potential generated by MCs is too small to contribute noticeably to the b-wave. The analysis also explains why MCs produce the large cornea-negative sPIII subcomponent of the ERG, but no substantial cornea-positive potential.
Topics: Animals; Electroretinography; Ependymoglial Cells; Mice; Microelectrodes; Photic Stimulation; Potassium; Retina
PubMed: 34294176
DOI: 10.1017/S0952523821000092 -
PloS One 2021Diabetic retinopathy (DR), the most common complication of diabetes mellitus, is associated with oxidative stress, nuclear factor-κB (NFκB) activation, and excess...
Diabetic retinopathy (DR), the most common complication of diabetes mellitus, is associated with oxidative stress, nuclear factor-κB (NFκB) activation, and excess production of vascular endothelial growth factor (VEGF) and intracellular adhesion molecule-1 (ICAM-1). Muller glial cells, spanning the entirety of the retina, are involved in DR inflammation. Mitigation of DR pathology currently occurs via invasive, frequently ineffective therapies which can cause adverse effects. The application of far-red to near-infrared (NIR) light (630-1000nm) reduces oxidative stress and inflammation in vitro and in vivo. Thus, we hypothesize that 670nm light treatment will diminish oxidative stress preventing downstream inflammatory mechanisms associated with DR initiated by Muller cells. In this study, we used an in vitro model system of rat Müller glial cells grown under normal (5 mM) or high (25 mM) glucose conditions and treated with a 670 nm light emitting diode array (LED) (4.5 J/cm2) or no light (sham) daily. We report that a single 670 nm light treatment diminished reactive oxygen species (ROS) production and preserved mitochondrial integrity in this in vitro model of early DR. Furthermore, treatment for 3 days in culture reduced NFκB activity to levels observed in normal glucose and prevented the subsequent increase in ICAM-1. The ability of 670nm light treatment to prevent early molecular changes in this in vitro high glucose model system suggests light treatment could mitigate early deleterious effects modulating inflammatory signaling and diminishing oxidative stress.
Topics: Animals; Cells, Cultured; Energy Metabolism; Ependymoglial Cells; Glucose; Infrared Rays; Mitochondria; Oxidative Stress; Rats; Reactive Oxygen Species; Sweetening Agents
PubMed: 34860856
DOI: 10.1371/journal.pone.0260968 -
Cell and Tissue Research Dec 2022T helper 22 (Th22) cells have been implicated in diabetic retinopathy (DR), but it remains unclear whether Th22 cells involve in the pathogenesis of DR. To investigate...
T helper 22 (Th22) cells have been implicated in diabetic retinopathy (DR), but it remains unclear whether Th22 cells involve in the pathogenesis of DR. To investigate the role of Th22 cells in DR mice, the animal models were established by intraperitoneal injection of STZ and confirmed by fundus fluorescein angiography and retinal haematoxylin-eosin staining. IL-22BP was administered by intravitreal injection. IL-22 level was measured by ELISA in vivo and in vitro. The expression of IL-22Rα1 in the retina was assessed by immunofluorescence. We assessed GFAP, VEGF, ICAM-1, inflammatory-associated factors and the integrity of blood-retinal barrier in control, DR, IL-22BP, and sham group. Müller cells were co-cultured with Th22 cells, and the expression of the above proteins was measured by immunoblotting. Plasmid transfection technique was used to silence Act1 gene in Müller cells. Results in vivo and in vitro indicated that Th22 cells infiltrated into the DR retinal and IL-22Rα1 expressed in Müller cells. Th22 cells promoted Müller cells activation and inflammatory factor secretion by secreting IL-22 compared with high-glucose stimulation alone. In addition, IL-22BP ameliorated the pathological alterations of the retina in DR. Inhibition of the inflammatory signalling cascade through Act1 knockdown alleviated DR-like pathology. All in all, the results suggested that Th22 cells infiltrated into the retina and secreted IL-22 in DR, and then IL-22 binding with IL-22Rα1 activated the Act1/TRAF6 signal pathway, and promoted the inflammatory of Müller cells and involved the pathogenesis of DR.
Topics: Mice; Animals; Diabetic Retinopathy; Ependymoglial Cells; TNF Receptor-Associated Factor 6; Diabetes Mellitus, Experimental; Vascular Endothelial Growth Factor A; Retina
PubMed: 36201050
DOI: 10.1007/s00441-022-03689-8 -
Viruses Jun 2023Retinopathy is a recently recognized complication of dengue, affecting up to 10% of hospitalized patients. Research on the pathogenesis has focused largely on effects of...
Retinopathy is a recently recognized complication of dengue, affecting up to 10% of hospitalized patients. Research on the pathogenesis has focused largely on effects of dengue virus (DENV) at the blood-retinal barrier. Involvement of retinal Müller glial cells has received little attention, although this cell population contributes to the pathology of other intraocular infections. The goal of our work was to establish the susceptibility of Müller cells to infection with DENV and to identify characteristics of the cellular antiviral, inflammatory, and immunomodulatory responses to DENV infection in vitro. Primary human Müller cell isolates and the MIO-M1 human Müller cell line were infected with the laboratory-adapted Mon601 strain and DENV serotype 1 and 2 field isolates, and cell-DENV interactions were investigated by immunolabelling and quantitative real-time polymerase chain reaction. Müller cells were susceptible to DENV infection, but experiments involving primary cell isolates indicated inter-individual variation. Viral infection induced an inflammatory response (including tumour necrosis factor-α, interleukin [IL]-1β, and IL-6) and an immunomodulatory response (including programmed death-ligand [PD-L]1 and PD-L2). The type I interferon response was muted in the Müller cell line compared to primary cell isolates. The highest infectivity and cell responses were observed in the laboratory-adapted strain, and overall, infectivity and cell responses were stronger in DENV2 strains. This work demonstrates that Müller cells mount an antiviral and immune response to DENV infection, and that this response varies across cell isolates and DENV strain. The research provides a direction for future efforts to understand the role of human retinal Müller glial cells in dengue retinopathy.
Topics: Humans; Dengue; Dengue Virus; Ependymoglial Cells; Cell Line; Antiviral Agents
PubMed: 37515098
DOI: 10.3390/v15071410 -
Pharmaceutical Biology Dec 2021Oxidative injury in a high-glucose (HG) environment may be a mechanism of diabetic retinopathy (DR) and edaravone can protect retinal ganglion cells by scavenging ROS.
CONTEXT
Oxidative injury in a high-glucose (HG) environment may be a mechanism of diabetic retinopathy (DR) and edaravone can protect retinal ganglion cells by scavenging ROS.
OBJECTIVE
To explore the effect of edaravone on HG-induced injury.
MATERIALS AND METHODS
First, Müller cells were cultured by different concentrations of glucose for different durations to obtain a suitable culture concentrations and duration. Müller cells were then divided into Control, HG + Vehicle, HG + Eda-5 μM, HG + Eda-10 μM, HG + Eda-20 μM, and HG + Eda-40 μM groups. Cells were cultured by 20 mM glucose and different concentrations of edaravone for 72 h.
RESULTS
The IC of glucose at 12-72 h is 489.3, 103.5, 27.92 and 20.71 mM, respectively. When Müller cells were cultured in 20 mM glucose for 72 h, the cell viability was 52.3%. Edaravone significantly increased cell viability compared to Vehicle (68.4% vs 53.3%; 78.6% vs 53.3%). The EC of edaravone is 34.38 μM. HG induced high apoptosis rate (25.5%), while edaravone (20 and 40 μM) reduced it to 12.5% and 6.89%. HG increased the DCF fluorescence signal (189% of Control) and decreased the mitochondrial membrane potential by 57%. Edaravone significantly decreased the DCF fluorescence signal (144% and 132% of Control) and recovered the mitochondrial membrane potential to 68% and 89% of Control. Furthermore, HG decreased the expression of TRX1, PGC-1α, NRF1 and TFAM, which were restored by edaravone.
DISCUSSION AND CONCLUSION
These findings provide a new potential approach for the treatment of DR and indicated new molecular targets in the prevention of DR.
Topics: Animals; Apoptosis; Cell Survival; Cells, Cultured; Diabetic Retinopathy; Edaravone; Ependymoglial Cells; Free Radical Scavengers; Glucose; Inhibitory Concentration 50; Membrane Potential, Mitochondrial; Nuclear Respiratory Factor 1; Oxidative Stress; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Rats; Rats, Sprague-Dawley; Thioredoxins; Transcription Factors
PubMed: 34506218
DOI: 10.1080/13880209.2021.1972123 -
Ophthalmic & Physiological Optics : the... Nov 2020To investigate presumed activated retinal astrocytes and Müller cells (ARAM) detected by scanning laser ophthalmoscopy (SLO) and spectral domain optical coherence...
PURPOSE
To investigate presumed activated retinal astrocytes and Müller cells (ARAM) detected by scanning laser ophthalmoscopy (SLO) and spectral domain optical coherence tomography, and to investigate its presence in healthy controls as well as its relationship to posterior vitreal detachment (PVD) and glaucoma.
METHODS
This retrospective study involved 1337 eyes of 805 controls between ages 8 and 90, and 250 eyes of 146 patients with glaucoma between the ages of 28 and 95. Subjects were counted as possessing ARAM only if they met the following criteria: (1) a patchy, discrete, glittering appearance on SLO, (2) a distinct, flat, hyper-reflective layer at the internal limiting membrane on at least one B-scan crossing the glittering area and (3) absence of any surface wrinkling retinopathy. The diagnosis of PVD was based on both the patient's clinical examination and imaging data. Frequency tables were used to describe categorical variables and differences were compared by means of χ . Analyses were separated based on right and left eye, first on controls and then between glaucomatous eyes and age-similar sex-matched controls.
RESULTS
ARAM was found in both healthy controls and patients with glaucoma at similar frequencies. There was no association between having glaucoma and the presence of ARAM. ARAM was not different between the sexes but was associated with age and having a PVD.
CONCLUSIONS
This large retrospective study found that ARAM can be seen in healthy controls, is associated with PVD and possibly independently with age, and occurred at similar frequency in glaucomatous eyes.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Astrocytes; Child; Ependymoglial Cells; Female; Glaucoma, Open-Angle; Humans; Male; Middle Aged; Ophthalmoscopy; Retina; Retrospective Studies; Tomography, Optical Coherence; Young Adult
PubMed: 32885879
DOI: 10.1111/opo.12731 -
International Journal of Molecular... Jun 2023The disorganization of retinal inner layers (DRIL) is an optical coherence tomography (OCT) biomarker strictly associated with visual outcomes in patients with diabetic... (Observational Study)
Observational Study
The disorganization of retinal inner layers (DRIL) is an optical coherence tomography (OCT) biomarker strictly associated with visual outcomes in patients with diabetic macular edema (DME) whose pathophysiology is still unclear. The aim of this study was to characterize in vivo, using retinal imaging and liquid biopsy, DRIL in eyes with DME. This was an observational cross-sectional study. Patients affected by center-involved DME were enrolled. All patients underwent spectral domain optical coherence tomography (SD-OCT) and proteomic analysis of aqueous humor (AH). The presence of DRIL at OCT was analyzed by two masked retinal experts. Fifty-seven biochemical biomarkers were analyzed from AH samples. Nineteen eyes of nineteen DME patients were enrolled. DRIL was present in 10 patients (52.63%). No statistically significant difference was found between DME eyes with and without DRIL, considering the AH concentration of all the analyzed biomarkers except for glial fibrillary acidic protein (GFAP), a biomarker of Müller cells dysfunction ( = 0.02). In conclusion, DRIL, in DME eyes, seems to strictly depend on a major dysfunction of Müller cells, explaining its role not only as imaging biomarker, but also as visual function Müller cells-related parameter.
Topics: Humans; Macular Edema; Diabetic Retinopathy; Cross-Sectional Studies; Ependymoglial Cells; Proteomics; Retrospective Studies; Visual Acuity; Fluorescein Angiography; Retina; Tomography, Optical Coherence; Biomarkers; Diabetes Mellitus
PubMed: 37298558
DOI: 10.3390/ijms24119607 -
Current Eye Research Aug 2023This study aimed to investigate the role of salidroside (SAL) in the cellular communication between Müller cells and retinal ganglion cells in diabetic mice.
PURPOSE
This study aimed to investigate the role of salidroside (SAL) in the cellular communication between Müller cells and retinal ganglion cells in diabetic mice.
METHODS
The diabetes mellitus (DM) animal models were established by the intraperitoneal injection of streptozotocin and treatment with SAL gavage or by the injection of IL-22BP into the vitreous cavity. Immunohistochemistry was used to measure the expression of the glial fibrillary acidic protein in Müller cells. The expression of IL-22 and IL-22Rα1 in retinal tissues was assessed by immunofluorescence. Western blotting was used to measure the expression of inflammatory and apoptosis-related proteins. Hematoxylin-eosin staining, TUNEL staining, and flow cytometry were used to analyze the apoptosis of retinal ganglion cells. The effect of cellular interactions was explored by Transwell assays.
RESULTS
Western blotting showed that glial fibrillary acidic protein, IL-22 protein expression was significantly upregulated in the DM animal models compared with the control mice. Immunofluorescence showed that IL-22 was highly expressed in Müller cells and IL-22Rα1 was expressed in ganglion cells in the retina of DM mice. Hematoxylin-eosin and TUNEL staining results showed an increase in the number of ganglion cells apoptotic in DM. However, SAL reversed these phenomena. Meanwhile, after coculture with Müller cells, Western blotting suggested that ganglion cells secreted p-STAT3, and c-caspase3 protein expression was increased. More interestingly, the treatment of IL-22BP and SAL inhibited the expression of the p-STAT3 and c-caspase3 proteins. Flow cytometry indicates that compared with the control group, the apoptosis rate of ganglion cells was increased in the high glucose group, while the apoptosis rate of cells in the recombinant IL-22 protein group was significantly increased, while the SAL inhibited ganglion cells apoptosis.
CONCLUSION
SAL inhibits the apoptosis of retinal ganglion cells the IL-22/STAT3 pathway in Müller cells.
Topics: Mice; Animals; Diabetic Retinopathy; Ependymoglial Cells; Glial Fibrillary Acidic Protein; Diabetes Mellitus, Experimental; Eosine Yellowish-(YS); Hematoxylin; Apoptosis
PubMed: 37287123
DOI: 10.1080/02713683.2023.2204208