Müller cells, as the predominant glial element in the sensory retina, play a crucial role in healthy and diseased retina. Overactivation of Müller cells in response to damage is detrimental to the ...retina tissue. Current research shows that inhibiting glial fibrillary acidic protein (GFAP), a sensitive indicator of Müller cell activation, attenuated glial reactions and promoted neuroprotection. Recent evidence suggests that the transcript factor SOX9 (sex-determining region Y box 9), part of the SOX family, regulates GFAP expression of astrocytes in the central nervous system. However, in retina Müller cells, it is still unknown whether GFAP can be downregulated by reduced SOX9 function. The present results show that clustered regularly interspaced short palindromic repeats/Cas9-mediated SOX9 knockout not only inhibited GFAP expression in rat Müller cells but also attenuated cell migration ability. These results suggest that inhibition of SOX9 activity may be a novel therapeutic strategy for reduction of glial cell activity.
Attention is increasingly being given to microglia-related inflammation in neovascular diseases, such as diabetic retinopathy and age-related macular disease. Evidence shows that activated microglia ...contribute to disruption of the blood-retinal barrier, however, the mechanism is unclear. In this study, we aimed to clarify whether and how microglia affect the function of retinal microvascular endothelial cells (RMECs).
We activated microglia by Lipopolysaccharides (LPS) stimulation. After co-culturing static or activated microglia with RMECs using the Transwell system, we evaluated the function of RMECs. Vascular endothelial growth factor-A (VEGF-A) and platelet-derived growth factor-BB (PDGF-BB) levels in the supernatant from the lower chamber were evaluated by ELISA. Angiogenesis, migration, and proliferation of RMECs were assessed by tube formation, wound healing, and WST-1 assays. The expression levels of tight junction proteins (ZO-1 and occludin) and endothelial markers (CD31 and CD34) were examined by Western blot analysis.
We successfully established an LPS-activated microglia model and co-culture system of static or activated microglia with RMECs. In the co-culture system, we showed that microglia, especially activated microglia stimulated VEGF-A and PDGF-BB expression, enhanced angiogenesis, migration, proliferation, and permeability, and altered the phenotype of co-cultured RMECs.
Microglia, especially activated microglia, play important roles in angiogenesis and maintenance of vascular function hemostasis in the retinal microvasculature. The mechanism needs further investigation and clarification.
•We efficiently induced WJ-MSCs into neuron-like cells using a modified method.•After neuronal induction, most of WJ-MSCs expressed mature neuronal markers.•Microarray analysis revealed distinct ...miRNA profiles in WJ-MSCs derived neurons.•The expression of six miRNAs were further validated by stem-loop RT-PCR.
Mesenchymal stem cells (MSCs) have the capacity to generate multiple tissues of mesodermal origin, and also have the potential to trans-differentiate into neurons. We isolated MSCs from the Wharton’s jelly of the human umbilical cord (WJ-MSCs), and efficiently induced WJ-MSCs into neuron-like cells using a modified method. After neuronal induction for 12 days, most of WJ-MSCs expressed mature neuronal marker MAP2 (83±7%), and meanwhile some adopted neuronal morphology. WJ-MSCs also expressed Nestin (34±6%), NSE (30±5%), and GFAP (12±3%). Moreover, we used miRNA microarray to analyze the differentially expressed miRNAs in neuronal differentiation of WJ-MSCs. Microarray analysis revealed discrepant miRNA profiles in the uninduced WJ-MSCs and WJ-MSCs derived neurons. Six miRNAs were chosen for further qRT-PCR validation. Among these 6 miRNAs, four miRNAs (miR-1290, miR-26b, miR-194, and miR-124a) were up-regulated and 2 miRNAs (miR-4521 and miR-543) were down-regulated in the WJ-MSCs derived neurons. In conclusion, WJ-MSCs could be efficiently induced into neuron-like cells. More importantly, our findings suggested that miRNAs might play important roles in the neuronal differentiation of WJ-MSCs.
Retinal neurodegeneration is induced by a variety of environmental insults and stresses, but the exact mechanisms are unclear. In the present study, we explored the involvement of cytosolic ...mitochondrial DNA (mtDNA), resulting in the cGAS-STING dependent inflammatory response and apoptosis in retinal damage in vivo.
Retinal injury was induced with white light or intravitreal injection of lipopolysaccharide (LPS). After light- or LPS-induced injury, the amount of cytosolic mtDNA in the retina was detected by PCR. The mtDNA was isolated and used to transfect retinas in vivo. WB and real-time PCR were used to evaluate the activation of cGAS-STING pathway and the levels of apoptosis-associated protein at different times after mtDNA injection. Retinal cell apoptosis rate was detected by TUNEL staining. Full-field electroretinography (ERG) was used to assess the retinal function.
Light injury and the intravitreal injection of LPS both caused the leakage of mtDNA into the cytoplasm in retinal tissue. After the transfection of mtDNA in vivo, the levels of cGAS, STING, and IFN-β mRNAs and the protein levels of STING, phosph-TBK1, phospho-IRF3, and IFN-β were upregulated. mtDNA injection also induced the activation of caspase 3 and caspase 9. BAX and BAK were increased at both the mRNA and protein levels. The release of cytochrome c from the mitochondria to the cytosol was increased after mtDNA injection. The wave amplitudes on ERG decreased and retinal cell apoptosis was detected after mtDNA injection.
Cytosolic mtDNA triggers an inflammatory response. It also promotes apoptosis and the dysfunction of the retina.
The purpose of this study was to replicate genetic factors associated with the susceptibility to Behçet's disease (BD). We conducted a two-stage candidate genes association and functional study, ...involving 477 BD patients and 1,334 normal controls of Chinese Han descent.
The genotyping of five candidate genes/loci, including LOC100129342, KIAA1529, CPVL, UBASH3B and UBAC2, were performed using TaqMan single nucleotide polymorphism (SNP) assays. Real-time PCR and luciferase reporter assay were performed to test the function of the identified promoter polymorphism. The main outcome measures were genotype frequencies and expression levels in BD patients.
The first-stage study results showed that UBAC2 (rs9513584, Pc = 0.018, OR = 1.4), but not LOC100129342, KIAA1529, CPVL, UBASH3B was associated with the susceptibility to BD in Chinese Han. The fine-mapping association study of UBAC2 identified six risk SNPs for BD in the Chinese cohort; three of them were verified in validation study (rs3825427, first-stage Pc = 2.2 × 10-3, second-stage Pc = 9.3 × 10-3, combined Pc = 6.9 × 10-6; rs9517668, first-stage Pc = 1.7 × 10-3, second-stage Pc = 0.03, combined Pc = 3.3 × 10-4; rs9517701, first-stage Pc = 5.1 × 10-3, second-stage Pc = 9.0 × 10-3, combined Pc = 2.9 × 10-5; respectively). Functional analysis showed that the risk T allele of the promoter polymorphism rs3825427 had a significantly lower promoter activity than the non-risk G allele (P = 0.002) and a decreased expression of UBAC2 transcript variant 1 in peripheral blood mononuclear cells (PBMCs) and skin of normal controls carrying the risk T allele than that in individuals with the G allele (P = 0.045, P = 0.025; respectively). The mRNA expression of UBAC2 transcript variant 1 was significantly decreased in PBMCs and skin of BD patients as compared with controls (P = 0.025; P = 0.047, respectively). The mRNA expression of UBAC2 transcript variant 2 was significantly increased in skin of BD patients as compared with controls (P = 0.004).
This study replicates a predisposition gene to BD, UBAC2, and suggests that UBAC2 may be involved in the development of BD through its transcriptional modulation.
Retinal ischemia/reperfusion (I/R) injury is implicated in the etiology of various ocular disorders. Prior research has demonstrated that bone marrow tyrosine kinase on chromosome X (BMX) contributes ...to the advancement of ischemic disease and inflammatory reactions. Consequently, the current investigation aims to evaluate BMX's impact on retinal I/R injury and clarify its implied mechanism of action.
This study utilized male and female systemic BMX knockout (BMX−/−) mice to conduct experiments. The utilization of Western blot assay and immunofluorescence labeling techniques was employed to investigate variations in the expression of protein and tissue localization. Histomorphological changes were observed through H&E staining and SD-OCT examination. Visual function changes were assessed through electrophysiological experiments. Furthermore, apoptosis in the retina was identified using the TUNEL assay, as well as the ELISA technique, which has been utilized to determine the inflammatory factors level.
Our investigation results revealed that the knockdown of BMX did not yield a significant effect on mouse retina. In mice, BMX knockdown mitigated the negative impact of I/R injury on retinal tissue structure and visual function. BMX knockdown effectively reduced apoptosis, suppressed inflammatory responses, and decreased inflammatory factors subsequent to I/R injury. The outcomes of the current investigation revealed that BMX knockdown partially protected the retina through downregulating phosphorylation of AKT/ERK/STAT3 pathway.
Our investigation showed that BMX−/− reduces AKT, ERK, and STAT3 phosphorylation, reducing apoptosis and inflammation. Thus, this strategy protected the retina from structural and functional damage after I/R injury.
•BMX−/− mitigated the detrimental influence of I/R injury on retinal tissue structure and visual function.•BMX−/− effectively reduced cell death and inflammation.•BMX−/− protects the retina by blocking phosphorylation in the AKT/ERK/STAT3 pathway.
Nuclear factor of activated T cells, cytoplasmic 4 (NFATc4) is one of the four members of the NFAT family, which were described first as essential components of T cells activation and lately as ...important regulators for the initiation and coordination of the immune response, including B cells and natural killer cells. Accumulating evidence has demonstrated that NFATc4 exerted a pro-apoptotic effect in the pathogenesis of various experimental central nervous system diseases by upregulating Fas ligand (FasL) levels. However, the function of NFATc4 in the retina is still with limited acquaintance. To investigate whether NFATc4 is involved in retinal neuron apoptosis, we performed a light-induced retinal damage model in adult rats. A significant upregulation of NFATc4 was detected in the retina after light-induced damage by using Western blotting and reverse transcriptase PCR (RT-PCR). Besides this, NFATc4 was observed to be localized mainly in the retinal ganglion cells (RGCs). In addition, the expression patterns of active caspase-3, active caspase-8, and FasL were parallel with that of NFATc4. We also found the co-localization of NFATc4 with active caspase-3 and FasL in RGCs after light exposure. Collectively, we hypothesized that NFATc4 might participate in RGCs apoptosis by upregulating FasL levels.
Glucocorticoid-induced leucine zipper (GILZ) mediates several effects of glucocorticoids and has important anti-inflammatory properties. Here, we explored the role of GILZ in inhibiting retinal ...inflammation. Endotoxin-induced uveitis (EIU) was established in rats by intravitreal injection of lipopolysaccharide (LPS). GILZ levels decreased in the EIU retina after LPS injection. Retinal GILZ was downregulated by recombinant lentivirus-delivered short-hairpin RNA targeting GILZ (shRNA-GILZ-rLV) and upregulated by recombinant lentivirus-mediated GILZ overexpression (Oe-GILZ-rLV). GILZ silencing attenuated the anti-inflammatory effects of intravitreal injection of triamcinolone acetonide (TA) in the EIU retina, as demonstrated by increased retinal interleukin (IL)-1β, monocyte chemoattractant protein (MCP)-1and intercellular cell adhesion molecule-1 expression at 18 h after TA injection. A Bio-Plex cytokine assay and western blotting demonstrated that GILZ overexpression inhibited the effects of LPS, downregulating retinal IL-1β, MCP-1, MIP-1α, and IL-17 and inhibiting LPS-induced activation of the retinal toll-like receptor 4–myeloid differentiation factor 88 signaling pathway. At 48 and 72 h after LPS injection, the clinical score of inflammation was significantly lower in Oe-GILZ-rLV–transfected eyes than in blank-rLV–transfected eyes. Histological examination showed a 67.85% reduction of infiltrating inflammatory cells in the anterior chamber and a 58.97% reduction in vitreous cavity of Oe-GILZ-rLV transfected eyes at 48 h after LPS injection. Taken together, our results suggest that GILZ is a novel therapeutic target for the treatment of retinal inflammatory diseases.
•Retinal GILZ was downregulated or upregulated by specific recombinant lentiviruses.•Retinal GILZ overexpression attenuated expression of proinflammatory biomarkers.•GILZ overexpression reduced retinal IL-1β, MCP-1, MIP-1α, and IL-17 expression.•GILZ inhibited LPS-induced activation of the retinal TLR4–MyD88 signaling pathway.
We explore the choroid vasculature changes of acute Vogt-Koyanagi-Harada (VKH) disease using widefield optical coherence tomography angiography (OCTA).
In this retrospective, observational, ...longitudinal study, 16 patients with acute VKH disease (32 eyes, mean age: 42.19 ±13.66 years) were measured using widefield OCTA.
In this study, we first described the multiple dark foci in choriocapillaris and Sattler's layer in the panoramic montage of the five 12x12mm images in 30 eyes (93.8%) of acute VKH disease. OCTA follow-up in these 30 eyes demonstrated the diminished size and number of these dark foci in choriocapillaris and Sattler's layer after the initiation of treatment.
Widefield OCTA enables noninvasive identification of characteristics of flow void at the level of superficial choroidal vessels in the acute phase and may be a novel valuable tool for diagnosis and monitoring of disease progression in VKH disease in the future.
Introduction: The aim of the study was to describe the characteristics of open globe injury (OGI) and the relationship between the complications and visual outcomes in children with this type of ...injury. Methods: This was a retrospective chart review of 1,664 children, under the age of 16 years, who were hospitalized for OGI between January 1, 2007, and December 31, 2015. Each patient’s age, sex, cause and agent of injury, complications, visual acuity, and classification of ocular trauma were collected for review and analysis. Results: The mean age was 5.6 ± 3.4 years. Right eyes were particularly vulnerable to injury (right eye:left eye ratio = 1.2:1). Traumatic cataract was the most common complication. The average initial and final best corrected visual acuity were logarithm of the minimum angle of resolution (logMAR) 2.04 ± 0.78 and logMAR 1.74 ± 0.88, respectively. Logistic regression analysis showed that hyphema (odds ratio OR = 1.850), iris prolapse (OR = 1.702), vitreous hemorrhage (OR = 9.703), retinal detachment (OR = 11.938), endophthalmia (OR = 5.377), intraocular foreign body (OR = 3.346), and initial visual acuity <0.05 (OR = 9.017) were risk factors for visual acuity <0.05 at hospital discharge. Conclusion: OGI was most frequent in preschool children and boys. Right eyes were more vulnerable than left eyes. Poor visual outcomes were associated with hyphema, iris prolapse, vitreous hemorrhage, retinal detachment, endophthalmia, intraocular foreign body, and an initial visual acuity <0.05.
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