Identification of Key lncRNAs in Gout Under Copper Death and Iron Death Mechanisms: A Study Based on ceRNA Network Analysis and Random Forest Algorithm.

This study focused on identifying potential key lncRNAs associated with gout under the mechanisms of copper death and iron death through ceRNA network analysis and Random Forest (RF) algorithm, which aimed to provide new insights into the molecular mechanisms of gout, and potential molecular targets for future therapeutic strategies of gout. Initially, we conducted an in-depth bioinformatics analysis of gout microarray chips to screen the key cuproptosis-related genes (CRGs) and key ferroptosis-related genes (FRGs). Using these data, we constructed a key ceRNA network for gout. Finally, key lncRNAs associated with gout were identified through the RF algorithm combined with ROC curves, and validated using the Comparative Toxicogenomics Database (CTD). We successfully identified NLRP3, LIPT1, and DBT as key CRGs associated with gout, and G6PD, PRKAA1, LIG3, PHF21A, KLF2, PGRMC1, JUN, PANX2, and AR as key FRGs associated with gout. The key ceRNA network identified four downregulated key lncRNAs (SEPSECS-AS1, LINC01054, REV3L-IT1, and ZNF883) along with three downregulated mRNAs (DBT, AR, and PRKAA1) based on the ceRNA theory. According to CTD validation inference scores and biological functions of target mRNAs, we identified a potential gout-associated lncRNA ZNF883/hsa-miR-539-5p/PRKAA1 regulatory axis. This study identified the key lncRNA ZNF883 in the context of copper death and iron death mechanisms related to gout for the first time through the application of ceRNA network analysis and the RF algorithm, thereby filling a research gap in this field and providing new insights into the molecular mechanisms of gout. We further found that lncRNA ZNF883 might function in gout patients by regulating PRKAA1, the mechanism of which was potentially related to uric acid reabsorption in the proximal renal tubules and inflammation regulation. The proposed lncRNA ZNF883/hsa-miR-539-5p/PRKAA1 regulatory axis might represent a potential RNA regulatory pathway for controlling the progression of gout disease. This discovery offered new molecular targets for the treatment of gout, and had significant implications for future therapeutic strategies in managing the gout.

Mapping Regional Homogeneity and Functional Connectivity of the Visual Cortex in Resting-State fMRI.

A combined regional homogeneity (ReHo) and functional connectivity (FC) method, a type of noninvasive functional magnetic resonance imaging (fMRI) method, has been used to evaluate synchronous neuronal activity changes in retinitis pigmentosa (RP). The purpose of this study is to describe our method for analysis of intra- and interregional synchronizations of changes in neuronal activity in RP patients. The advantages of the combined ReHo and FC method are that it is both noninvasive and sufficiently sensitive to investigate changes in cerebral synchronous neuronal activity changes in vivo. Here, 16 RP patients and 14 healthy controls closely matched in age, sex, and education underwent resting-state fMRI scans. Two sample t-tests were conducted to compare ReHo and FC across groups. Our results showed that visual network disconnection and reorganization of the retino-thalamocortical pathway and dorsal visual stream occurred in the RP patients. Here, we describe the details of this method, its use, and the impact of its key parameters in a step-by-step manner.

Automated identification of retinopathy of prematurity by image-based deep learning.

BACKGROUND: Retinopathy of prematurity (ROP) is a leading cause of childhood blindness worldwide but can be a treatable retinal disease with appropriate and timely diagnosis. This study was performed to develop a robust intelligent system based on deep learning to automatically classify the severity of ROP from fundus images and detect the stage of ROP and presence of plus disease to enable automated diagnosis and further treatment. METHODS: A total of 36,231 fundus images were labeled by 13 licensed retinal experts. A 101-layer convolutional neural network (ResNet) and a faster region-based convolutional neural network (Faster-RCNN) were trained for image classification and identification. We applied a 10-fold cross-validation method to train and optimize our algorithms. The accuracy, sensitivity, and specificity were assessed in a four-degree classification task to evaluate the performance of the intelligent system. The performance of the system was compared with results obtained by two retinal experts. Moreover, the system was designed to detect the stage of ROP and presence of plus disease as well as to highlight lesion regions based on an object detection network using Faster-RCNN. RESULTS: The system achieved an accuracy of 0.903 for the ROP severity classification. Specifically, the accuracies in discriminating normal, mild, semi-urgent, and urgent were 0.883, 0.900, 0.957, and 0.870, respectively; the corresponding accuracies of the two experts were 0.902 and 0.898. Furthermore, our model achieved an accuracy of 0.957 for detecting the stage of ROP and 0.896 for detecting plus disease; the accuracies in discriminating stage I to stage V were 0.876, 0.942, 0.968, 0.998 and 0.999, respectively. CONCLUSIONS: Our system was able to detect ROP and differentiate four-level classification fundus images with high accuracy and specificity. The performance of the system was comparable to or better than that of human experts, demonstrating that this system could be used to support clinical decisions.

Large-Scale Neuronal Network Dysfunction in Diabetic Retinopathy.

Diabetic retinopathy (DR) patients are at an increased risk of cognitive decline and dementia. There is accumulating evidence that specific functional and structural architecture changes in the brain are related to cognitive impairment in DR patients. However, little is known regarding whether the functional architecture of resting-state networks (RSNs) changes in DR patients. The purpose of this study was to investigate the intranetwork functional connectivity (FC) and functional network connectivity (FNC) of RSN changes in DR patients using independent component analysis (ICA). Thirty-four DR patients (18 men and 16 women; mean age, 53.53 ± 8.67 years) and 38 nondiabetic healthy controls (HCs) (15 men and 23 women; mean age, 48.63 ± 11.83 years), closely matched for age, sex, and education, underwent resting-state magnetic resonance imaging scans. ICA was applied to extract the nine RSNs. Then, two-sample t-tests were conducted to investigate different intranetwork FCs within nine RSNs between the two groups. The FNC toolbox was used to assess interactions among RSNs. Pearson correlation analysis was conducted to explore the relationship between intranetwork FCs and clinical variables in the DR group. A receiver operating characteristic (ROC) curve was conducted to assess the ability of the intranetwork FCs of RSNs in discriminating between the two groups. Compared to the HC group, DR patients showed significant decreased intranetwork FCs within the basal ganglia network (BGN), visual network (VN), ventral default mode network (vDMN), right executive control network (rECN), salience network (SN), left executive control network (lECN), auditory network (AN), and dorsal default mode network (dDMN). In addition, FNC analysis showed increased VN-BGN, VN-vDMN, VN-dDMN, vDMN-lECN, SN-BGN, lECN-dDMN, and AN-BGN FNCs in the DR group, relative to the HC group. Furthermore, altered intranetwork FCs of RSNs were significantly correlated with the glycosylated hemoglobin (HbA1c) level in DR patients. A ROC curve showed that these specific intranetwork FCs of RSNs discriminated between the two groups with a high degree of sensitivity and specificity. Our study highlighted that DR patients had widespread deficits in both low-level perceptual and higher-order cognitive networks. Our results offer important insights into the neural mechanisms of visual loss and cognitive decline in DR patients.

Assessment of cerebral low-frequency oscillations in patients with retinal vein occlusion: a preliminary functional MRI study.

BACKGROUND: There is increasing evidence that patients with retinal vein occlusion exhibit cerebral vascular changes and are at an increased risk of stroke. However, it remains unknown whether patients with retinal vein occlusion exhibit changes in intrinsic brain activity. PURPOSE: This study investigated intrinsic brain activity changes in patients with retinal vein occlusion by assessing the amplitude of low-frequency fluctuations. MATERIAL AND METHODS: Forty-five patients with retinal vein occlusion (22 men, 23 women, mean age 56.55 ± 6.97 years) and 43 healthy controls (13 men, 30 women; mean age 53.53 ± 8.19 years) closely matched in age, sex, and education level underwent resting-state MRI scans. The amplitude of low-frequency fluctuation method was used to compare intrinsic brain activity between the two groups. RESULTS: Compared with healthy controls, patients with retinal vein occlusion exhibited significantly lower amplitude of low-frequency fluctuation values in the left middle occipital gyrus, right middle occipital gyrus, and right calcarine. However, patients with retinal vein occlusion showed significantly higher amplitude of low-frequency fluctuations in the bilateral cerebellum 6, right hippocampus, left insula, and left fusiform (voxel-level P < 0.01, Gaussian random field correction, cluster-level P < 0.05). CONCLUSION: Our results demonstrated that patients with retinal vein occlusion showed abnormal spontaneous neural activities in the visual cortices, cerebellum, and Papez circuit, which might indicate impaired vision, cognition, and emotional function in patients with retinal vein occlusion. These findings offer important insights into the neural mechanism of retinal vein occlusion.

Abnormal intrinsic functional network hubs and connectivity following peripheral visual loss because of inherited retinal degeneration.

Previous neuroimaging studies have shown that the long-term effects of peripheral vision loss lead to functional and morphological reorganization in visual cortices. However, it has not been determined whether whole-brain functional network centrality changes occur during peripheral vision loss. This study aimed to investigate functional network centrality and connectivity changes in individuals with peripheral vision loss because of retinitis pigmentosa (RP) by using voxel-wise degree centrality (DC) and seed-based resting-state functional connectivity (rsFC) methods. In total, 30 RP patients (18 men and 12 women, mean age: 38.77±14.44 years) and 30 healthy controls (HCs) (18 men and 12 women, mean age: 34.57±10.70 years) matched for age, sex, cognition, education, and visual expertise underwent resting-state magnetic resonance imaging scans. Graph theory-based network analysis was carried out to investigate DC between the two groups. A seed-based rsFC analysis was then carried out to further reveal the abnormal functional connectivity of the altered DC brain region. Pearson's correlation was used to analyze the relationships of DC and rsFC index with the clinical variables in RP patients: visual function (best-corrected visual acuity and visual field, VF) and optical coherence tomography testing (mean retinal nerve fiber layer). Compared with HCs, RP patients had significantly lower DC values in the bilateral cuneus/calcarine/precuneus (CUN/CAL/PreCUN) [Brodmann's area (BA) 17/18/19/30/31]. In addition, RP patients showed decreased rsFC index, relative to that of HCs, from bilateral CUN/CAL/PreCUN to bilateral lingual/cuneus/calcarine (LIG/CUN/CAL) (BA 18/19/30) and the bilateral postcentral gyrus/superior parietal lobule (BA 3/5/7/40). In contrast, RP patients showed increased rsFC index, relative to that of HCs, from bilateral CUN/CAL/PreCUN to bilateral thalamus/caudate (voxel-level P<0.01; Gaussian random-field correction, cluster-level P<0.05). Moreover, the course of RP showed a negative correlation with the mean DC values of the bilateral CUN/CAL/PreCUN (r=-0.480; P=0.007) and the mean FC values of the bilateral LIG/CUN/CAL (r=-0.484; P=0.007); the mean DC values of the bilateral CUN/CAL/PreCUN in RP showed a negative correlation with the right eye VF (r=-0.411; P=0.024) and left eye VF (r=-0.426; P=0.019). Our results showed that RP patients showed abnormal function network hubs in various brain regions related to visual, thalamocortical, and sensorimotor networks; these might reflect impaired top-down modulations, visual imagery, and visuomotor coordination in RP patients. Moreover, the DC index can be used as a biomarker to indicate the severity of visual loss in RP patients.

Inhibitory effects of low decibel infrasound on the cardiac fibroblasts and the involved mechanism.

INTRODUCTION: Infrasound is a mechanical vibration wave with frequency between 0.0001 and 20 Hz. It has been established that infrasound of 120 dB or stronger is dangerous to humans. However, the biological effects of low decibel infrasound are largely unknown. The purpose of this study was to investigate the effects of low decibel infrasound on the cardiac fibroblasts. MATERIALS AND METHODS: The cardiac fibroblasts were isolated and cultured from Sprague-Dawley rats. The cultured cells were assigned into the following four groups: control group, angiotensin II (Ang II) group, infrasound group, and Ang II+infrasound group. The cell proliferation and collagen synthesis rates were evaluated by means of [3H]-thymidine and [3H]-proline incorporation, respectively. The levels of TGF-ß were determined by enzyme-linked immunosorbent assay. Moreover, RNAi approaches were used for the analysis of the biological functions of miR-29a, and the phosphorylation status of Smad3 was detected using western blotting analysis. RESULTS: The results showed that low decibel infrasound significantly alleviated Ang II-induced enhancement of cell proliferation and collagen synthesis. DISCUSSION: Compared with the control, Ang II markedly decreased the expression of miR-29a levels and increased the secretion of TGF-ß and phosphorylation of Smad3, which was partly reversed by the treatment with low decibel infrasound. Importantly, knockdown of miR-29a diminished the effects of infrasound on the cardiac fibroblasts. In conclusion, low decibel infrasound inhibits Ang II-stimulated cardiac fibroblasts via miR-29a targeting TGF-ß/Smad3 signaling.

Activation of the sigma receptor 1 modulates AMPA receptor-mediated light-evoked excitatory postsynaptic currents in rat retinal ganglion cells.

Sigma receptor (σR), a unique receptor family, is classified into three subtypes: σR1, σR2 and σR3. It was previously shown that σR1 activation induced by 1µM SKF10047 (SKF) suppressed N-methyl-d-aspartate (NMDA) receptor-mediated responses of rat retinal ganglion cells (GCs) and the suppression was mediated by a distinct Ca(2+)-dependent phospholipase C (PLC)-protein kinase C (PKC) pathway. In the present work, using whole-cell patch-clamp techniques in rat retinal slice preparations, we further demonstrate that SKF of higher dosage (50µM) significantly suppressed AMPA receptor (AMPAR)-mediated light-evoked excitatory postsynaptic currents (L-EPSCs) of retinal ON-type GCs (ON GCs), and the effect was reversed by the σR1 antagonist BD1047, suggesting the involvement of σR1. The SKF (50µM) effect was unlikely due to a change in glutamate release from bipolar cells, as suggested by the unaltered paired-pulse ratio (PPR) of AMPAR-mediated EPSCs of ON GCs. SKF (50µM) did not change L-EPSCs of ON GCs when the G protein inhibitor GDP-ß-S or the protein kinase G (PKG) inhibitor KT5823 was intracellularly infused. Calcium imaging further revealed that SKF (50µM) did not change intracellular calcium concentration in GCs and persisted to suppress L-EPSCs when intracellular calcium was chelated by BAPTA. The SKF (50µM) effect was intact when protein kinase A (PKA) and phosphatidylinostiol (PI)-PLC signaling pathways were both blocked. We conclude that the SKF (50µM) effect is Ca(2+)-independent, PKG-dependent, but not involving PKA, PI-PLC pathways.

Signalling mechanism for somatostatin receptor 5-mediated suppression of AMPA responses in rat retinal ganglion cells.

Somatostatin (SRIF) is involved in a variety of physiological functions via the activation of five subtypes of specific receptors (sst1-5). Here, we investigated the effects of SRIF on AMPA receptor (AMPAR)-mediated currents (AMPA currents) in isolated rat retinal ganglion cells (GCs) using patch-clamp techniques. Immunofluorescence double labelling demonstrated the expression of sst5 in rat GCs. Consistent to this, whole cell AMPA currents of GCs were dose-dependently suppressed by SRIF, and the effect was reversed by the sst5 antagonist BIM-23056. Intracellular dialysis of GDP-ß-S or pre-incubation with the Gi/o inhibitor pertussis toxin (PTX) abolished the SRIF effect. The SRIF effect was mimicked by the administration of either 8-Br-cAMP or forskolin, but was eliminated by the protein kinase A (PKA) antagonists H-89/KT5720/Rp-cAMP. Moreover, SRIF increased intracellular Ca(2+) levels and did not suppress the AMPA currents when GCs were infused with an intracellular Ca(2+)-free solution or in the presence of ryanodine receptor modulators caffeine/ryanodine. Furthermore, the SRIF effect was eliminated when the activity of calmodulin (CaM), calcineurin and protein phosphatase 1 (PP1) was blocked with W-7, FK-506 and okadaic acid, respectively. SRIF persisted to suppress the AMPA currents when cGMP-protein kinase G (PKG) and phosphatidylinositol (PI)-/phosphatidylcholine (PC)-phospholipase C (PLC) signalling pathways were blocked. In rat flat-mount retinas, SRIF suppressed AMPAR-mediated light-evoked excitatory postsynaptic currents (L-EPSCs) in GCs. We conclude that a distinct Gi/o/cAMP-PKA/ryanodine/Ca(2+)/CaM/calcineurin/PP1 signalling pathway comes into play due to the activation of sst5 to mediate the SRIF effect on GCs.

Orexin-A differentially modulates AMPA-preferring responses of ganglion cells and amacrine cells in rat retina.

By activating their receptors (OX1R and OX2R) orexin-A/B regulate wake/sleeping states, feeding behaviors, but the function of these peptides in the retina remains unknown. Using patch-clamp recordings and calcium imaging in rat isolated retinal cells, we demonstrated that orexin-A suppressed α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA)-preferring receptor-mediated currents (AMPA-preferring currents) in ganglion cells (GCs) through OX1R, but potentiated those in amacrine cells (ACs) through OX2R. Consistently, in rat retinal slices orexin-A suppressed light-evoked AMPA-preferring receptor-mediated excitatory postsynaptic currents in GCs, but potentiated those in ACs. Intracellular dialysis of GDP-ß-S or preincubation with the Gi/o inhibitor pertussis toxin (PTX) abolished both the effects. Either cAMP/the protein kinase A (PKA) inhibitor Rp-cAMP or cGMP/the PKG blocker KT5823 failed to alter the orexin-A effects. Whilst both of them involved activation of protein kinase C (PKC), the effects on GCs and ACs were respectively eliminated by the phosphatidylinositol (PI)-phospholipase C (PLC) inhibitor and phosphatidylcholine (PC)-PLC inhibitor. Moreover, in GCs orexin-A increased [Ca(2+)]i and the orexin-A effect was blocked by intracellular Ca(2+)-free solution and by inositol 1,4,5-trisphosphate (IP3) receptor antagonists. In contrast, orexin-A did not change [Ca(2+)]i in ACs and the orexin-A effect remained in intracellular or extracellular Ca(2+)-free solution. We conclude that a distinct Gi/o/PI-PLC/IP3/Ca(2+)-dependent PKC signaling pathway, following the activation of OX1R, is likely responsible for the orexin-A effect on GCs, whereas a Gi/o/PC-PLC/Ca(2+)-independent PKC signaling pathway, following the activation of OX2R, mediates the orexin-A effect on ACs. These two actions of orexin-A, while working in concert, provide a characteristic way for modulating information processing in the inner retina.

Upregulation of glutamate-aspartate transporter by glial cell line-derived neurotrophic factor ameliorates cell apoptosis in neural retina in streptozotocin-induced diabetic rats.

AIMS: Dysfunction of glutamate uptake, largely mediated by the glutamate-aspartate transporter (GLAST), may lead to retinal cell apoptosis in diabetic retinopathy. The aim of this study is to examine how cell apoptosis and the expression level of GLAST in neural retina of a diabetic rat model are changed and whether the neuroretinal apoptosis could be ameliorated by the administration of glial cell line-derived neurotrophic factor (GDNF). METHODS: Diabetes was induced by intraperitoneal injection of streptozotocin (STZ) in Sprague-Dawley rats. GLAST protein expression levels were determined by Western blotting, whereas apoptosis of retinal neurons was evaluated by TUNEL staining. To assess the role of GDNF in ameliorating the STZ-induced retinal changes, GDNF/GDNF with siRNA directed against GLAST was injected into the vitreous after STZ injection. RESULTS: In rat retinas 4 weeks after the onset of STZ-induced diabetes, TUNEL-positive cells were significantly increased, whereas GLAST levels were significantly reduced. Intraocular administration of GDNF at the early stage of diabetes remarkably increased the GLAST levels and decreased TUNEL-positive signals in the retinas. These effects of GDNF were largely abolished by coadministration of GLAST siRNA. CONCLUSIONS: GDNF, administrated at the early stage of diabetes, could rescue retinal cells from neurodegeneration by upregulating the expression of GLAST.

Distribution of somatostatin receptor 5 in mouse and bullfrog retinas.

Somatostatin (SRIF), as a neuroactive peptide in the CNS, may act as a neuromodulator through activation of five specific receptor subtypes (sst(1)-sst(5)). In this work we conducted a comparative study of the expression of sst(5) in mouse and bullfrog retinas by immunofluorescence double labeling. Basically, the expression profiles of sst(5) in the retinas of the two species were similar. That is, in the inner retina sst(5) was localized to dopaminergic and cholinergic amacrine cells, stained by tyrosine hydroxylase (TH) and choline acetyltransferase (ChAT) respectively, and cells in the ganglion cell layer, whereas in the outer retina immunostaining for sst(5) was observed in horizontal cells. However, a more widespread, abundant distribution of labeling for sst(5), as compared to mouse retina, was seen in bullfrog retina: strong labeling for sst(5) was diffusely distributed in both outer and inner plexiform layers (OPL and IPL) in the bullfrog retina, but the labeling was only observed in the IPL of the mouse retina. In addition, bullfrog photoreceptors, both rods and cones, but not mouse ones, were labeled by sst(5). In combination with the experiments showing that SRIF-immunoreactivity was mainly found in the inner retina, our results suggest that SRIF, released from SRIF-containing cells in the inner retina, may play a neuromodulatory role in both outer and inner retina mediated by volume transmission via sst(5) in bullfrog retina, while the SRIF action may be largely restricted to the mouse inner retina.