Multimodal imaging of bilateral retinal pigment epithelial immunoglobulin light chain deposition in patients with systemic immunoglobulin light chain deposition.

OBJECTIVE: To describe multimodal imaging of peculiar bilateral globular subretinal deposits and acquired serous retinal detachment in patients with systemic immunoglobulin light chain deposition. DESIGN: A retrospective observational case series. PARTICIPANTS: We examined six eyes in three patients (one with multiple myeloma, one with membranous nephropathy, and one with immunoglobulin A nephropathy) at the Eye and ENT Hospital of Fudan University. The patients presented with peculiar globular subretinal deposits along the retinal pigment epithelium (RPE)‒Bruch's membrane complex and acquired serous retinal detachment. METHODS: Fundus appearance was documented with multimodal imaging, which included fundus photography, fundus autofluorescence, spectral domain optical coherence tomography (OCT), swept-source OCT (SS-OCT), en-face OCT, and SS-OCT angiography. Additional evaluations included serum protein electrophoreses, positron emission tomography computed tomography, and renal and bone biopsies to assess the primary diseases. MAIN OUTCOME MEASURES: Multimodal imaging, course, and prognosis of bilateral RPE immunoglobulin light chain deposition in patients with systemic immunoglobulin light chain deposition. RESULTS: Bilateral, multiple, speckled, or patchy RPE changes in the posterior fundus that corresponded to striking multifocal hyperautofluorescence on fundus autofluorescence and lumpy, globular hyperreflective deposits along the RPE‒Bruch's membrane complex were identified as characteristic features of bilateral RPE light chain deposition. These features may be accompanied by dense light chain deposits in the choriocapillaris and choroid vessels, diffuse choroidal thickening, and "angiographically silent" serous retinal detachment in patients with systemic immunoglobulin light chain deposition. CONCLUSIONS: We have documented the characteristic features, clinical course, and prognosis of bilateral RPE immunoglobulin light chain deposition in patients with systemic immunoglobulin light chain deposition. Appropriate evaluations, including serum protein electrophoresis and hematologic consultation, are recommended to manage patients with this fundus abnormality.

High score of LDH plus dNLR predicts poor survival in patients with HER2-positive advanced breast cancer treated with trastuzumab emtansine.

OBJECTIVE: To investigate the prognostic value of derived neutrophil to lymphocyte ratio (dNLR) and lactate dehydrogenase (LDH) in patients with advanced HER2 positive breast cancer treated with trastuzumab emtansine. METHODS: Fifty one patients with advanced HER2 positive breast cancer who received T-DM1 treatment in Harbin Medical University Cancer Hospital were selected. The clinical data and blood test indexes were collected, and the ROC curve determined the optimal cut-off value. Kaplan-Meier survival curve and Cox regression model was used to analyze the effect of different levels of dNLR,LDH,LNI (dNLR combined with LDH index) before and after T-DM1 treatment on the survival of patients. RESULTS: The median PFS and OS of the patients with advanced HER2 positive breast cancer who received T-DM1 treatment were 6.9 months and 22.2 months, respectively. The optimal cut-off value of LDH and dNLR before T-DM1 treatment was 244 U / L (P = 0.003) and 1.985 (P = 0.013), respectively. Higher LDH and dNLR were significantly correlated with shorter median PFS and OS (P < 0.05). The median PFS of patients with LNI (0), LNI (1) and LNI (2) were 8.1 months, 5.5 months and 2.3 months, respectively, P = 0.007. Univariate and multivariate analysis showed that LDH > 244 U / L, dNLR > 1.985, LNI > 0, ECOG ≥1 and HER-2 (IHC2 +, FISH+) before the T-DM1 treatment were the poor prognostic factors. LDH uptrend after the T-DM1 treatment also predicted poor prognosis. CONCLUSION: Serum LDH > 244 U / L and dNLR > 1.985 before the T-DM1 treatment were prognostic risk factors for patients with advanced HER2 positive breast cancer receiving T-DM1 treatment. The higher LNI score was significantly associated with shorter PFS and OS. LDH uptrend after T-DM1 treatment was also related to the poor prognosis.

Intravitreal injection of mitochondrial DNA induces cell damage and retinal dysfunction in rats

BACKGROUND: 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. METHODS: 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 path-way 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. RESULTS: 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-ß mRNAs and the protein levels of STING, phosph-TBK1, phospho-IRF3, and IFN-ß 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. CONCLUSIONS: Cytosolic mtDNA triggers an inflammatory response. It also promotes apoptosis and the dysfunction of the retina.

Changes of macular vessel density and thickness in gas and silicone oil tamponades after vitrectomy for macula-on rhegmatogenous retinal detachment.

PURPOSE: To investigate the macular vessel density and thickness in macular-on rhegmatogenous retinal detachment (RRD) after vitrectomy with gas and silicone oil (SO) tamponade. METHODS: Patients with macular-on RRD eyes, treated with a single successful vitrectomy with gas or SO tamponade and a minimum 30 months follow-up, were reviewed. Best-corrected visual acuity (BCVA), macular vessel density and retinal thickness by using optical coherence tomography angiography, were compared to the contralateral eyes. RESULTS: Sixteen eyes with gas tamponade and 17 eyes with SO tamponade were included in the study. LogMAR best-corrected visual acuity (BCVA) slightly improved from 0.25 ± 0.18 (Snellen 20/36) to 0.17 ± 0.23 (Snellen 20/30) in eyes with gas tamponade, and decreased from 0.30 ± 0.22 (Snellen 20/40) to 0.49 ± 0.28 (Snellen 20/62) in eyes with SO tamponade. The parafoveal vessel densities in superficial vascular complex (SVC) and the corresponding inner retinal thickness (IRT) were similar between the affected eyes and the contralateral eyes in gas tamponade group (P = 0.578, P = 0.943), while significantly reduced in the affected eyes, compared to the contralateral eyes in SO tamponade group (P < 0.001, P < 0.001). CONCLUSION: Eyes in SO tamponade group had worse BCVA, lower SVC vessel densities and thinner corresponding IRT after vitrectomy for macular-on RRD, than those in gas tamponade group.

MicroRNA-29b-3p inhibits cell proliferation and angiogenesis by targeting VEGFA and PDGFB in retinal microvascular endothelial cells.

Purpose: Excessive angiogenesis, also known as neovascularization, has considerable pathophysiologic roles in several retinal diseases, including retinopathy of prematurity, diabetic retinopathy, and exudative age-related macular degeneration. Accumulated evidence has revealed that miRNAs play important roles in endothelial cell dysfunction and angiogenesis. However, the role of microRNA-29b-3p (miR-29b-3p) in retinal angiogenesis is still unclear. Therefore, we investigated whether and how miR-29b-3p affects the function of retinal microvascular endothelial cells (RMECs). Methods: The overexpression and inhibition of miR-29b-3p were achieved by transfecting rat RMECs with an miR-29b-3p mimic and inhibitor, respectively. The proliferation, migration, and angiogenesis of RMECs were evaluated using a Cell Counting Kit-8 assay, Ki67 staining, western blotting (of proliferating cell nuclear antigen, cyclin A2, cyclin D1, and cyclin E1), wound healing test, and tube formation assay. The expression levels of vascular endothelial growth factor A (VEGFA) and platelet-derived growth factor B (PDGFB) were examined with quantitative real-time PCR and western blotting, respectively. Results: Overexpression of miR-29b-3p statistically significantly inhibited the function of RMECs in cell proliferation and angiogenesis, while inhibition of miR-29b-3p increased the proliferative and angiogenic activities of RMECs. Moreover, VEGFA and PDGFB, as the targets of miR-29b-3p, were statistically significantly downregulated by the miR-29b mimic, whereas the miR-29b-3p inhibitor had the opposite effects. Conclusions: miR-29b-3p negatively regulates RMEC proliferation and angiogenesis, at least partly by targeting VEGFA and PDGFB. These data may provide a potential therapeutic strategy for treating ocular neovascular diseases.

RETINAL TELANGIECTASIA IN PATIENTS WITH PATHOLOGIC MYOPIA: A CASE SERIES.

PURPOSE: To describe the characteristics of retinal telangiectasia in eyes with pathologic myopia. METHODS: The study included 10 patients (18 eyes) who were diagnosed with pathologic myopia combined with retinal telangiectasia. The patients visited our retinal clinic every 3 months. Nine eyes underwent vitrectomy for vision-threatening complications after diagnosis. All eyes underwent comprehensive ophthalmologic examinations including multimodal retinal imaging at presentation and at each follow-up. RESULTS: Retinal telangiectasia in pathologic myopia was characterized by saccular aneurysmal dilatation of the capillary bed without hard exudates in color fundus photographs and hyporeflective saccular structure in infrared reflectance fundus photographs, and it was filled in the early retinal arteriovenous phase with minimal dye leakage in the late phase of fundus fluorescein angiography. Spectral domain optical coherence tomography and optical coherence tomographic angiography showed that retinal telangiectasia was primarily located in the superficial retina, together with myopic traction maculopathy. In the 9 eyes that underwent vitrectomy, the retinal telangiectasia regressed within 3 months of surgery. Retinal telangiectasia remained stable in the other nine eyes, but these eyes were at risk of spontaneous bleeding. CONCLUSION: Retinal telangiectasia is a relatively quiescent and uncommon disorder in patients with pathologic myopia that might be closely related to myopic traction maculopathy.

Synthesized glucocorticoid-induced leucine zipper peptide inhibits photoreceptor apoptosis and protects retinal function in light-induced retinal degeneration model.

BACKGROUND: This study aimed to investigate the neuroprotective function of a synthesized glucocorticoid-induced leucine zipper peptide (GILZ-p) in a light-induced retinal degeneration model. METHODS: The GILZ98-134 peptide was synthesized and injected intravitreally into Sprague Dawley rats. Retinal injury was then induced in the rats by exposing their eyes to constant white light (5000 lux) for 24 h. The activation of retinal caspases-9/3 and the release of cytochrome c from the mitochondria to the cytosol were measured at 1, 3, 5 and 7 d after light injury. Photoreceptor apoptosis was evaluated with terminal-deoxynucleotidyl-transferase-mediated deoxyuridine triphosphate-biotin nick end labelling (TUNEL) staining at 3 d after injury. Haematoxylin and eosin staining and electroretinography were used to observe the changes in the retinal morphology and function, respectively, at 7 and 14 d after light injury. RESULTS: The intravitreally injected synthesized GILZ-p successfully penetrated to the retina and significantly inhibited the activation of retinal caspase-3 and caspase-9 at 1, 3, 5 and 7 d after light injury, and reduced the number of TUNEL-positive photoreceptors at 3 d after light injury. GILZ-p pre-treatment also alleviated cytochrome c release and rescued mitochondria-mediated apoptosis after injury. Simultaneously, GILZ-p pre-treatment also mitigated the light-induced thinning of the outer nuclear layer and the loss of retinal function at 7 and 14 d after light injury, respectively. CONCLUSIONS: The synthesized GILZ-p prevented light-induced photoreceptor apoptosis and protected retinal function from degeneration, and is therefore a potential therapeutic option for degenerative retinal diseases.

Light-Induced Lipocalin 2 Facilitates Cellular Apoptosis by Positively Regulating Reactive Oxygen Species/Bim Signaling in Retinal Degeneration.

Purpose: Lipocalin 2 (LCN2) is reported to be one of the key regulators of cell survival and death; however, its effect on retinal degeneration is unclear. Therefore, we aimed to investigate the role of LCN2 and its underlying mechanisms in light-induced retinal degeneration. Methods: A recombinant lentivirus expressing a short hairpin RNA targeting LCN2 mRNA and a recombinant lentivirus overexpressing LCN2 were used to downregulate and upregulate retinal LCN2, respectively. Seven days after intravitreal injection of the lentiviruses, rats were exposed to blue light (2500 lux) for 24 hours. Retinal function and morphology were evaluated with ERG and hematoxylin-eosin staining, respectively. TUNEL staining was used to detect apoptotic cells. The levels of reactive oxygen species (ROS) were evaluated with dihydroethidium labeling. Western blotting and real-time PCR were used to examine protein and mRNA expression levels, respectively. Results: Retinal LCN2 expression was significantly upregulated after light exposure. Light exposure reduced the amplitudes of a- and b-waves on the ERG and the thickness of the outer nuclear layer and promoted photoreceptor apoptosis. These phenomena were clearly attenuated by LCN2 knockdown, whereas LCN2 overexpression had the opposite effects. The overexpression of LCN2 facilitated photoreceptor apoptosis by increasing ROS generation and Bim expression. On the opposite, LCN2 knockdown mitigated the generation of light-exposure-induced ROS and the activation of the Bim-mediated mitochondrial apoptotic pathway. Conclusions: Light-induced LCN2 is a proapoptotic factor in the retina, and LCN2 knockdown protects photoreceptors from apoptosis by inhibiting ROS production and Bim expression. LCN2 is a potential therapeutic target for light-induced retinal degeneration.

A Synthesized Glucocorticoid- Induced Leucine Zipper Peptide Inhibits Retinal Müller Cell Gliosis.

Purpose: The anti-inflammatory activities of protein glucocorticoid-induced leucine zipper (GILZ) have been demonstrated in vivo and in vitro. Here, we examined the potential effect of a synthetic peptide derived from the leucine zipper motif and proline-rich region of GILZ on suppressing inflammatory responses in primary cultured rat Müller cells. Methods: Peptides were selected from amino acids 98-134 of the GILZ protein (GILZ-p). Solid-phase peptide synthesis was used to generate the cell-penetrating peptide TAT, which was bound to the amino terminus of GILZ-p. Primary cultured retinal Müller cells were stimulated with lipopolysaccharide (LPS) alone or in combination with different concentrations of GILZ-p, and the interaction of GILZ-p with nuclear factor (NF)-κB p65 in Müller cells was investigated by western blotting, immunoprecipitation, and immunofluorescence. The expression of the Müller cell gliosis marker glial fibrillary acidic protein (GFAP), functional protein aquaporin (AQP)-4, and the inflammatory cytokines interleukin (IL)-1ß, tumor necrosis factor (TNF) α, intercellular adhesion molecule (ICAM)-1, and monocyte chemoattractant protein (MCP)-1 was measured by Western Blotting. The concentration of those cytokines in culture medium was measured by using Enzyme-Linked Immunosorbent Assay. Results: The synthesized GILZ-p, which was water-soluble, entered cells and bound with NF-κB p65, inhibiting p65 nuclear translocation. GILZ-p inhibited the LPS-induced expression of GFAP, IL-1ß, TNFα, ICAM-1, and MCP-1 in Müller cells and prevented the LPS-induced downregulation of AQP4. Conclusions: These results indicate that GILZ-p interacted with NF-κB p65 and suppressed p65 nuclear translocation, thereby inhibiting inflammatory cytokine release and Müller cell gliosis.

Lipocalin 2 Suppresses Ocular Inflammation by Inhibiting the Activation of NF-κß Pathway in Endotoxin-Induced Uveitis.

BACKGROUND/AIMS: Lipocalin 2 (LCN2), an important mediator of a variety of cellular processes, is involved in regulating the inflammatory response, but its roles in different inflammatory diseases are controversial. Because the role of LCN2 in ocular inflammation has been unclear until now, we explored the function of LCN2 in lipopolysaccharide (LPS)-induced ocular inflammation in vivo and in vitro. METHODS: Endotoxin-induced uveitis (EIU) was induced in male Sprague Dawley rats by the intravitreal injection of LPS. The expression and location of LCN2 in the retina were detected with western blotting and immunohistochemistry, respectively. We determined the clinical scores for anterior inflammation, quantified the infiltrated inflammatory cells, and measured the pro-inflammatory factors to determine the anti-inflammatory effects of LCN2 in EIU eyes. Cultured primary rat Müller cells were stimulated with LPS and the expression and secretion of LCN2 were measured with real-time PCR, western blotting, and an ELISA. After Müller cells were cotreated with LPS and LCN2 or PBS, the expression and secretion of TNF-α, IL-6, and MCP-1 were examined with realtime PCR, western blotting, and ELISAs. Western blotting and immunofluorescence were used to detect the phosphorylation and cellular distribution of nuclear factor kappaB (NF-κB) subunit p65. RESULTS: In EIU, the expression of LCN2 was significantly upregulated in the retina, especially in the outer nuclear layer (mainly composed of Müller cells). LPS stimulation of cultured Müller cells also markedly elevated LCN2 expression. Intravitreal injection of LCN2 significantly reduced the clinical scores, inflammatory infiltration, and protein leakage in EIU, which correlated with the reduced levels of proinflammatory factors in the aqueous humor and retina. LCN2 treatment also reduced the expression and secretion of TNF-α, IL-6, and MCP-1 in LPS-stimulated Müller cells. LCN2 inhibited the inflammatory response by inhibiting the phosphorylation and translocation of NF-κB p65. CONCLUSIONS: LCN2 protects against ocular inflammation, at least in part, by negatively regulating the activation of the NF-κB signaling pathway. LCN2 may be a promising anti-inflammatory therapy for ocular diseases, such as uveitis.

Glucocorticoid-Induced Leucine Zipper Protects the Retina From Light-Induced Retinal Degeneration by Inducing Bcl-xL in Rats.

Purpose: The aim of the present study was to investigate the neuroprotective effects of glucocorticoid-induced leucine zipper (GILZ) in a light-induced retinal degeneration model and to explore the underlying mechanisms. Methods: Intravitreal injection of recombinant GILZ-overexpressing lentivirus (OE-GILZ-rLV) and short hairpin RNA targeting GILZ recombinant lentivirus (shRNA-GILZ-rLV) was performed to up- and downregulate retinal GILZ, respectively. Three days after stable transduction, rats were exposed to continuous bright light (5000 lux) for 2 days. Retinal function was assessed by full-field electroretinography (ERG), and the retinal structure was examined for photoreceptor survival and death in rats kept under a 12-hour light:2-hour dark cycle following light exposure. The expression levels of retinal Bcl-xL, caspase-9, and caspase-3 were examined by Western blotting or real-time PCR at 1, 3, 5, and 7 days after light exposure. Results: Exposure to bright light downregulated retinal GILZ in parallel with the downregulation of Bcl-xL and the upregulation of active caspase-3. Overexpression of retinal GILZ attenuated the decrease of Bcl-xL and the activation of caspase-9 and caspase-3 at 1, 3, 5, and 7 days after bright light exposure, respectively. GILZ silencing aggravated the downregulation of Bcl-xL induced by bright light exposure. Bright light exposure reduced the amplitude of ERG, increased the number of apoptotic photoreceptor cells, and decreased retinal thickness; and GILZ overexpression could attenuate all these effects. Conclusions: Overexpression of GILZ by OE-GILZ-rLV transduction protected the retina from light-induced cellular damage by activating antiapoptotic pathways.

Glucocorticoid-Induced Leucine Zipper Suppresses ICAM-1 and MCP-1 Expression by Dephosphorylation of NF-κB p65 in Retinal Endothelial Cells.

Purpose: Glucocorticoid-induced leucine zipper (GILZ) is involved in anti-inflammatory activities in several animal models and in various cell types. In this study, we explored the role of GILZ in rat retinal vascular endothelial cells. Methods: Glucocorticoid-induced leucine zipper overexpression or silencing was established using GILZ overexpressing recombinant lentivirus (OE-GILZ-rLV) or short-hairpin RNA targeting GILZ recombinant lentivirus (shRNA-GILZ-rLV), respectively, in rat primary retinal microvascular endothelial cells (RMECs) and intact retina. Seventy-two hours after transfection, RMECs were stimulated with 1000 ng/mL lipopolysaccharide (LPS), 20 µM isoliensinine (an alkaloid derived from the embryos of Nelumbo nucifera, could enhance the dephosphorylation of p65 at Ser536), or PBS for another 24 hours. Western blotting and immunofluorescence were performed to measure protein expression. The concentrations of intercellular adhesion molecule (ICAM)-1 and monocyte chemoattractant protein (MCP)-1 in the RMEC culture media were measured by ELISA. Results: Lipopolysaccharide downregulated GILZ expression in RMECs in a time- and dose-dependent manner, and the decrease in GILZ expression was accompanied by increased ICAM-1 and MCP-1 expression. Glucocorticoid-induced leucine zipper overexpression decreased LPS-induced ICAM-1 and MCP-1 expression, whereas GILZ silencing significantly attenuated the production of both cytokines. Glucocorticoid-induced leucine zipper overexpression also inhibited LPS-induced nuclear factor-κB p65 nuclear translocation in RMECs that was mediated by enhanced p65 dephosphorylation. The dephosphorylation of NF-κB p65 further downregulated ICAM-1 and MCP-1 expression in RMECs. Conclusions: Glucocorticoid-induced leucine zipper overexpression inhibited NF-κB p65 nuclear translocation by enhancing p65 dephosphorylation. Exogenous GILZ regulated ICAM-1 and MCP-1 expression, which was probably mediated by enhanced p65 dephosphorylation.