Vitreous Levels Research Articles

Estimation of postmortem interval using vitreous potassium levels in cases of fatal road traffic collision

To produce a formula that can accurately predict postmortem interval (PMI) based on vitreous potassium levels using road traffic collision fatalities. Vitreous humour samples were taken from 78 individuals who had died following road traffic collisions between 2010 and 2015. Samples were obtained from both eyes and were sent for on-site analysis. Measurement of potassium was by an indirect ion-specific electrode Siemens diagnostics ADVIA 2400 chemistry system. Exact time of death was known from police reports, the time of postmortem was recorded and the postmortem interval was calculated. Linear regression was then used to analyse the relationship between the two. The impact of age was also assessed. PMI was between 6 and 162 hours. As vitreous potassium increases, the PMI also increases; exhibiting a linear relationship. This is illustrated by a regression equation of PMI = 6.42[K+] - 40.94, R = 0.67 (p < 0.001). This produced a formula closely comparable with three other studies proposed in previous literature and produces estimates that may exceed one calendar day. When both age and medical intervention are accounted for there is an insignificant improvement in prediction. Validated methods have been used to produce a formula for prediction of PMI using vitreous potassium. Although this is specific to road traffic collisions, the methods are transferable and can be seen to be comparable with other recently published methods. Nonetheless, if greater levels of accuracy are required it is suggested that biomarkers delivering a higher level of precision should still be sought.

Changes in vitreous VEGF, bFGF and fibrosis in proliferative diabetic retinopathy after intravitreal bevacizumab.

To evaluate the relationship between intravitreal bevacizumab (IVB) treatment and the levels of vitreous vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and vitreous-retina surface fibrosis in patients with proliferative diabetic retinopathy (PDR). This study was a prospective, open-label, controlled, randomized clinical trial. Sixty-eight eyes of PDR patients (n=53) and macular hole patients (n=15) were enrolled in this study. Thirty-four eyes of the PDR patients received IVB before vitrectomy. Twenty-three of the 34 PDR patients received IVB treatment 5d before vitrectomy (subgroup a), and 11 of the 34 PDR patients received IVB treatment greater than 2wk prior to vitrectomy (subgroup b). Nineteen of the PDR patients did not receive IVB treatment at any time prior to vitrectomy. The levels of bFGF and VEGF in vitreous samples were measured using enzyme-linked immunosorbent assay (ELISA) and the degree of vitreoretinal fibrosis was characterized using clinical data and data obtained intra-operatively. In PDR patients, VEGF and bFGF levels were significantly increased compared to non-PDR (control) subject's eyes (P<0.01). In PDR patients, vitreous VEGF levels were significantly decreased following IVB treatment compared to PDR patients that did not receive IVB treatment (P<0.01). The degree of vitreoretinal fibrosis was significantly increased in subgroup b compared to subgroup a(P<0.05) and to patients that did not receive IVB (P<0.05). Vitreous bFGF levels were significantly greater in subgroup b than subgroup a (P<0.01) or in patients who did not receive IVB treatment (P<0.05). A Spearman's rank correlation test indicated that higher levels of vitreous bFGF, but not VEGF, correlated with the degree of vitreoretinal fibrosis. We found that bFGF levels increase in PDR patient's vitreous after IVB treatment longer than two weeks prior to vitrectomy and correlated with the degree of fibrosis after IVB treatment. These findings suggest vitreous fibrosis is increased in PDR patients after IVB treatment may be due to increased levels of bFGF.

ANGPTL-4 correlates with vascular endothelial growth factor in patients with proliferative diabetic retinopathy.

To investigate the relationship between angiopoietin-like protein 4 (ANGPTL-4) and vascular endothelial growth factor (VEGF) in the serum and vitreous of eyes in patients with proliferative diabetic retinopathy (PDR). Thirty-five eyes of 35 patients with PDR, 20 eyes of 20 patients with non-proliferative diabetic retinopathy, 20 eyes of 20 patients with diabetes but no diabetic retinopathy, and 14 eyes of 14 nondiabetic patients with an idiopathic macular hole (IMH) were recruited from Shanghai First People's Hospital. The ANGPTL-4 and VEGF concentrations were determined using enzyme-linked immunosorbent assays. Group means were compared using one-way analysis of variance with GraphPad Prism 4.0 and SPSS ver. 17.0. The research followed the tenets of the Declaration of Helsinki. The ANGPTL-4 and VEGF levels were significantly higher in the vitreous and serum of patients with PDR compared with patients with IMH. There were significant correlations between the ANGPTL-4 and VEGF levels in the vitreous and serum of patients with PDR. The vitreous and serum ANGPTL-4 levels were also significantly correlated in patients with PDR. The ANGPTL-4 in both the vitreous and serum correlated with the serum triglyceride and high-density lipoprotein cholesterol levels. The ANGPTL-4 levels were markedly elevated and the ANGPTL-4 expression was directly correlated with the VEGF expression in the vitreous and serum of patients with PDR. The vitreous and serum ANGPTL-4 levels were also significantly correlated with serum lipids in patients with PDR. Our results suggest that the ANGPTL-4 may be used as a new therapeutic target for the treatment of PDR.

Increased sCD200 Levels in Vitreous of Patients With Proliferative Diabetic Retinopathy and Its Correlation With VEGF and Proinflammatory Cytokines.

The purpose of this study was to determine the levels of sCD200 expression in the vitreous of proliferative diabetic retinopathy (PDR) patients and to clarify its correlation with different vitreoretinal conditions, VEGF and its receptors, and proinflammatory cytokines. The expression of sCD200, VEGF and its receptors, and other proinflammatory cytokines were examined by using ELISA. Clinical stratification was performed on patients with different vitreoretinal conditions for correlation analysis. The vitreous levels of sCD200 were significantly higher in the PDR group (182.2 ± 17.63 pg/mL) compared with those in the control group (56.86 ± 6.573 pg/mL; P < 0.0001). The venous blood levels of sCD200 were 26.71 ± 4.32 pg/mL in the PDR group and 19.94 ± 3.87 pg/mL in the control group (P = 0.2614). The vitreous levels of sCD200 were significantly elevated in PDR patients with diabetic macular edema (DME; 266.9 ± 28.82 pg/mL) or traction retinal detachment (TRD; 256.9 ± 34.50 pg/mL) compared with the PDR group without DME (136.9 ± 15.13 pg/mL; P < 0.0001) or TRD (146.9 ± 15.97 pg/mL; P = 0.0024). The vitreous levels of CCL2, CXCL4, CXCL9, CXCL10, VEGF, sVEGFR-1, sVEGFR-2, IL-6, IL-8, IL-10, and IL-18 were also elevated significantly in the PDR group. Statistical association was found between sCD200 levels and VEGF (r = 0.6566, P < 0.0001), sVEGFR-1 (r = 0.5574, P = 0.006), sVEGFR-2 (r = 0.3605, P = 0.0362), CCL-2 (r = 0.6001, P = 0.0002), IL-6 (r = 0.5704, P = 0.0004), IL-8 (r = 0.3712, P = 0.0307), and IL-10 (r = 0.3618, P = 0.0355). Expression of sCD200 may contribute to retinal angiogenesis by interacting with VEGF-mediated inflammatory response and represents a potential therapeutic target for the patients with PDR.

Role of High‐mobility group box‐1 in diabetic retinal vasculopathy and neuropathy

SummaryHigh‐mobility mobility group box‐1 (HMGB1) is a non‐histone DNA‐binding protein that plays an important role in chromatin organization and transcriptional regulation. Extracellular HMGB1 functions as a proinflammatory cytokine and triggers the inflammatory response through the activation of multiple receptors such as the receptor for advanced glycation end products (RAGE), toll‐like receptor‐2 (TLR2), TLR4 and TLR9. We demonstrated that HMGB1 and RAGE were expressed in fibrovascular epiretinal membranes from patients with proliferative diabetic retinopathy (PDR). In addition, we demonstrated increased levels of HMGB1 in the vitreous samples from patients with PDR and that there were significant positive correlations between the vitreous levels and HMGB1 and the levels of the biomarkers of inflammation and oxidative injury. Furthermore, we demonstrated that diabetes induced significant upregulation of the expression of HMGB1 and RAGE in the retinas of rats and mice and that HMGB1 mediates diabetes‐induced oxidative stress, activation of inflammatory signaling pathways, breakdown of the blood‐retinal barrier and neuropathy in the retina. Therefore, compounds inhibiting HMGB1 may be novel therapeutic agents for diabetic retinopathy.

Mutual enhancement between high‐mobility group box‐1 and NADPH oxidase‐derived reactive oxygen species mediates diabetes‐induced upregulation of retinal apoptotic markers

SummaryWe hypothesized that a novel mechanism exists where HMGB1and NADPH oxidase (NOX)‐derived ROS are mutually enhanced in the diabetic retina, which may be a novel mechanism for promoting upregulation of retinal apoptotic markers induced by diabetes. To test this hypothesis we analyzed the vitreous samples from PDR and nondiabetic patients, retinas from rats and human retinal microvascular endothelial cells. We found that HMGB1 and the oxidative stress marker protein carbonyl content levels in the vitreous fluid from PDR patients were significantly higher than in controls. There was a significant positive correlation between vitreous fluid levels of HMGB1 and the levels of protein carbonyl content. HMGB1 enhanced interleukin‐1β, ROS, NOX2, and PARP‐1 and cleaved caspase‐3 expression by HRMEC. Diabetes and intravitreal injection of HMGB1 in normal rats induced significant upregulation of ROS, NOX2, PARP‐1 and cleaved caspase‐3 in the retina. Constant glycyrrhizin and apocynin intake from onset of diabetes did not affect the metabolic status of the diabetic rats, but restored these increased mediators to control values. Our results suggest that there is a mutual enhancement between HMGB1 and NOX‐derived ROS in the diabetic retina.

Evaluation of T‐cell related cytokines in the vitreous of proliferative diabetic retinopathy

SummaryMacrophages are involved in low‐grade inflammation in diabetes, and play pathogenic roles in proliferative diabetic retinopathy (PDR) by producing proinflammatory cytokines. T cells as well as other cells are activated by proinflammatory cytokines, and the infiltration into the vitreous of patients with PDR has been shown. We have recently found that both positive rates and levels of IL4, IL‐6, IL‐10, IL‐17A, IL‐21, IL‐22, and TNFα in the vitreous of PDR patients were significantly higher than those in the serum, and that vitreous levels of these cytokines and IL‐31 were significantly higher in PDR patients than in epiretinal membrane or macular hole patients. In addition, although vitreous of IFN‐g and sCD40L in the vitreous of endogenous uveitis (EU) patients were significantly higher than those of PDR patients, vitreous levels of IL‐4, IL‐17A, IL‐22, IL‐31, and TNFa were conversely higher in the vitreous of PDR patients than those of EU patients. Although it is unclear whether these cytokines synergistically play facilitative roles or inhibitory roles for the progression of PDR, our study suggested that Th2‐ and Th17‐related immune responses are involved in the pathogenesis of PDR.

Elevated Levels of Cytokines Associated with Th2 and Th17 Cells in Vitreous Fluid of Proliferative Diabetic Retinopathy Patients

Macrophages are involved in low-grade inflammation in diabetes, and play pathogenic roles in proliferative diabetic retinopathy (PDR) by producing proinflammatory cytokines. T cells as well as other cells are also activated by proinflammatory cytokines, and infiltration into the vitreous of patients with PDR has been shown. In this study, we measured helper T (Th) cell-related cytokines in the vitreous of PDR patients to define the characteristics of Th-mediated immune responses associated with PDR. The study group consisted of 25 type 2 diabetic patients (25 eyes) with PDR. The control group consisted of 27 patients with epiretinal membrane (ERM), 26 patients with idiopathic macular hole (MH), and 26 patients with uveitis associated with sarcoidosis. Vitreous fluid was obtained at the beginning of vitrectomy, and centrifuging for cellular removals was not performed. Serum was also collected from PDR patients. IL-1β, IL-4, IL-6, IL-10, IL-17A, IL-17F, IL-21, IL-22, IL-23, IL-25, IL-31, IL-33, IFN-γ, soluble sCD40L, and TNFα in the vitreous and serum samples were measured. Both percent detectable and levels of IL-4, IL-6, IL-17A, IL-21, IL-22, and TNFα in the vitreous were significantly higher than those in the serum in PDR patients. Vitreous levels of these cytokines and IL-31 were significantly higher in PDR than in ERM or MH patients. Vitreous levels of IL-4, IL-17A, IL-22, IL-31, and TNFα in PDR patients were also significantly higher than those of sarcoidosis patients. In PDR patients, vitreous IL-17A level correlated significantly with vitreous levels of IL-22 and IL-31, and especially with IL-4 and TNFα. Although it is unclear whether these cytokines play facilitative roles or inhibitory roles for the progression of PDR, the present study indicated that Th2- and Th17-related immune responses are involved in the pathogenesis of PDR.

EFFECT OF PEGAPTANIB AND RANIBIZUMAB ON PLASMA AND VITREOUS HOMOCYSTEINE IN PATIENTS WITH EXUDATIVE AGE-RELATED MACULAR DEGENERATION.

To investigate homocysteine (Hcy) concentration in the blood plasma and the vitreous in patients with exudative age-related macular degeneration receiving intravitreal anti-vascular endothelial growth factor therapy. Plasma Hcy and vitreous Hcy levels were analyzed in 73 exudative age-related macular degeneration patients (50.7% received pegaptanib 0.3 mg and 50.3% received ranibizumab 0.5 mg) and compared with 80 controls and 40 patients with idiopathic epiretinal membranes, respectively. Homocysteine concentration was measured by immunonephelometric particle test, and it was determined before and after antiangiogenic therapy. The mean Hcy concentrations (± SD) of blood plasma and vitreous were 13.0 ± 4.2 μmol/L and 1.00 ± 0.3 μmol/L in patients treated with pegaptanib; in ranibizumab group, they were 12.8 ± 2.5 μmol/L and 1.4 ± 0.6 μmol/L, respectively. The results of plasma and vitreous Hcy indicated statistically significant differences between exudative age-related macular degeneration patients and control groups (P = 0.03 and P = 0.02). After 6 months with both intravitreal therapies, the plasma and vitreous Hcy concentrations did not change significantly (P = 0.1). Pegaptanib and ranibizumab did not increase the plasma or vitreous Hcy concentrations.

Elevated vitreous body glial fibrillary acidic protein in retinal diseases.

PurposeIncreased expression of glial fibrillary acidic protein (GFAP) is a characteristic of gliotic activation (Müller cells and astrocytes) in the retina. This study assessed vitreous body GFAP levels in various forms of retinal pathology.MethodsThis prospective study included 82 patients who underwent vitrectomy (46 retinal detachments (RDs), 13 macular hole (MHs), 15 epiretinal glioses (EGs), 8 organ donors). An established enzyme–linked immunosorbent assay (ELISA, SMI26) was used for quantification of GFAP.ResultsThe highest concentration of vitreous body GFAP in organ donors was 20 pg/mL and it was used as the cutoff. A significant proportion of patients suffering from RD (65 %) to EG (53 %) had vitreous body GFAP levels above this cutoff when compared to organ donors (0 %, p < 0.0001, p = 0.0194, respectively, Fisher’s exact test) and MH (8 %, p < 0.0001, p = 0.0157, respectively). In RD and EG, vitreous body GFAP levels were correlated with axial length (R = 0.69, R = 0.52, p < 0.05 for both).ConclusionsThe data suggest that human vitreous body GFAP is a protein biomarker for glial activation in response to retinal pathologies. Vitreous body GFAP levels may be of interest as a surrogate outcome for experimental treatment strategies in translational studies.

The impact of ranibizumab on the level ofintercellular adhesion molecule type 1 in the vitreousofeyes with proliferative diabetic retinopathy.

This study was to investigate the impact of ranibizumab on the level of intercellular adhesion molecule type 1 (ICAM-1) in the vitreous of eyes with PDR. This is an interventional case-control study. A total of 82 eyes from 82 patients who had undergone vitreous surgery for the treatment of retinal disorders were included. Twenty-two eyes with PDR received an intravitreal ranibizumab injection (IVR) 3-7days before vitrectomy and were grouped as 'PDR with recent IVR' or Group 1. Sixteen eyes with PDR received IVR more than 7days before vitrectomy and were grouped as 'PDR with remote IVR' or Group 2. Twenty-two matched PDR eyes did not receive IVR before vitrectomy and were grouped as 'PDR without IVR' or Group 3. Finally, 22 eyes from 22 patients with idiopathic macular pucker (IMP) served as the 'non-diabetic control' group, or Group 4. Vitreous samples were obtained at the time of vitrectomy from all eyes, and the levels of vascular endothelium growth factor (VEGF) and ICAM-1 were analysed using ELISA. PDR without IVR (Group 3) had the highest vitreous VEGF concentration; the difference was significant compared with those in the PDR with recent IVR (Group 1), PDR with remote IVR (Group 2) and the non-diabetic control group (Group 4) (p<0.001). Group 2 had a lower vitreous VEGF level than Group 1 (p=0.041). Group 1 had the highest vitreous ICAM-1 levels (p<0.001 versus. Groups 2, 3 and 4); Group 2 had a lower vitreous ICAM-1 level than Group 3 (p=0.028). The vitreous fluid level of ICAM-1 was significantly increased within 1week of IVR administration, but markedly decreased after a week of administration in eyes with PDR. This suggests that leucostasis, vascular leakage and endothelial dysfunction may be amplified in the early days after IVR, but that a therapeutic effect of IVR in these processes may appear after 1week of ranibizumab administration in eyes with PDR.

Oxidative stress impact on growth hormone secretion in the eye.

AimTo evaluate the influence of oxidative stress on extrapituitary growth hormone (GH) secretion in the eye and to analyze the interdependence between eye and serum GH levels under normal and hypoxic conditions.MethodsPars plana vitrectomy (PPV) was performed in 32 patients with developed proliferative diabetic retinopathy (PDR) and 49 non-diabetic controls, both of whom required this procedure as part of their regular treatment in the period from April 2013 to December 2014. During PPV, vitreous samples were taken and blood was simultaneously collected from the cubital vein. GH levels in serum and vitreous samples were measured by electrochemical luminescence assay. Oxidative stress was measured by enzyme-linked immunosorbent assay of advanced oxidation protein products (AOPP) and lipid hydroperoxide (LPO) in serum and vitreous.ResultsSerum AOPP levels were significantly higher than vitreous levels in both groups (P < 0.001 for each group) and LPO levels were significantly higher only in PDR group (P < 0.001). There was a significant positive correlation between serum and vitreous LPO levels in PDR group (r = 0.909; P < 0.001). Serum GH levels were significantly higher than vitreous levels in both groups (P < 0.001 for each group). Serum GH levels were significantly higher in PDR group than in controls (P = 0.012). Vitreous GH values were slightly higher in PDR group, but the difference was not significant.ConclusionOur study confirms that GH production in the eye is autonomous and independent of oxidative stress or pituitary GH influence.

A new model for the estimation of time of death from vitreous potassium levels corrected for age and temperature

Analysis of potassium concentration in the vitreous fluid of the eye is frequently used by forensic pathologists to estimate the postmortem interval (PMI), particularly when other methods commonly used in the early phase of an investigation can no longer be applied. The postmortem rise in vitreous potassium has been recognized for several decades and is readily explained by a diffusion of potassium from surrounding cells into the vitreous fluid. However, there is no consensus regarding the mathematical equation that best describes this increase. The existing models assume a linear increase, but different slopes and starting points have been proposed. In this study, vitreous potassium levels, and a number of factors that may influence these levels, were examined in 462 cases with known postmortem intervals that ranged from 2h to 17 days. We found that the postmortem rise in potassium followed a non-linear curve and that decedent age and ambient temperature influenced the variability by 16% and 5%, respectively. A long duration of agony and a high alcohol level at the time of death contributed less than 1% variability, and evaluation of additional possible factors revealed no detectable impact on the rise of vitreous potassium. Two equations were subsequently generated, one that represents the best fit of the potassium concentrations alone, and a second that represents potassium concentrations with correction for decedent age and/or ambient temperature. The former was associated with narrow confidence intervals in the early postmortem phase, but the intervals gradually increased with longer PMIs. For the latter equation, the confidence intervals were reduced at all PMIs. Therefore, the model that best describes the observed postmortem rise in vitreous potassium levels includes potassium concentration, decedent age, and ambient temperature. Furthermore, the precision of these equations, particularly for long PMIs, is expected to gradually improve by adjusting the constants as more reference data are added over time. A web application that facilitates this calculation process and allows for such future modifications has been developed.

The Expression of GDF-15 in the Human Vitreous in the Presence of Retinal Pathologies with an Inflammatory Component

Purpose: The presence of growth differentiation factor-15 (GDF-15), a protein implicated in the regulation of the inflammatory response, was investigated in the vitreous of patients with vitreoretinal disorders.Methods: Vitreous and plasma samples were collected from patients with idiopathic epiretinal membrane (IERM), macular hole (MH), rhegmatogenous retinal detachment (RRD), nucleus drop (ND), or proliferative diabetic retinopathy (PDR). GDF-15 concentrations were measured using ELISA.Results: The vitreous levels of GDF-15 were higher in ND (5) and PDR (14) patients (1494 ± 243 and 904 ± 138 pg/mL, respectively) than RRD (3), MH (3), and IERM (8) patients (302 ± 160, 288 ± 24, and 254 ± 91 pg/mL, respectively). The vitreous levels of GDF-15 were significantly higher in patients with inflammatory vitreoretinal disorders (p < 0.0001).Conclusions: This is the first report showing that GDF-15 appears to be expressed in the vitreous, and that its expression is significantly higher in the presence of a vitreoretinal disorder in which there is an inflammatory component.

Mutual enhancement between high-mobility group box-1 and NADPH oxidase-derived reactive oxygen species mediates diabetes-induced upregulation of retinal apoptotic markers.

The expression of the proinflammatory cytokine high-mobility group box-1 (HMGB1) is upregulated in epiretinal membranes and vitreous fluid from patients with proliferative diabetic retinopathy (PDR) and in the diabetic retina. We hypothesized that a novel mechanism exists where HMGB1 and NADPH oxidase (Nox)-derived reactive oxygen species (ROS) are mutually enhanced in the diabetic retina, which may be a novel mechanism for promoting upregulation of retinal apoptotic markers induced by diabetes. Vitreous samples from 48 PDR and 34 nondiabetic patients, retinas from 1-month diabetic rats and from normal rats intravitreally injected with HMGB1 and human retinal microvascular endothelial cells (HRMEC) stimulated with HMGB1 were studied by enzyme-linked immunosorbent and spectrophotometric assays, Western blot analysis, RT-PCR, and immunofluorescence. We also studied the effect of the HMGB1 inhibitor glycyrrhizin and apocynin on diabetes-induced biochemical changes in the retinas of rats (n = 5-7 in each groups). HMGB1 and the oxidative stress marker protein carbonyl content levels in the vitreous fluid from PDR patients were significantly higher than in controls (p = 0.021; p = 0.005, respectively). There was a significant positive correlation between vitreous fluid levels of HMGB1 and the levels of protein carbonyl content (r = 0.62, p = 0.001). HMGB1 enhanced interleukin-1β, ROS, Nox2, poly (ADP-ribose) polymerase (PARP)-1, and cleaved caspase-3 production by HRMEC. Diabetes and intravitreal injection of HMGB1 in normal rats induced significant upregulation of ROS, Nox2, PARP-1, and cleaved caspase-3 in the retina. Constant glycyrrhizin and apocynin intake from onset of diabetes did not affect the metabolic status of the diabetic rats, but restored these increased mediators to control values. The results of this study suggest that there is a mutual enhancement between HMGB1 and Nox-derived ROS in the diabetic retina, which may promote diabetes-induced upregulation of retinal apoptotic markers.

Increased levels of inflammatory immune mediators in vitreous from pseudophakic eyes.

To determine if pseudophakic eyes have an increased and sustained level of inflammatory immune mediators in the vitreous compared to phakic eyes. Vitreous fluid samples were obtained from 73 patients undergoing elective pars plana vitrectomy (PPV) as a result of a macular hole, epiretinal membrane, vitreous macular traction, or vitreous floaters. Forty eyes were pseudophakic and had previously undergone uncomplicated cataract surgery, ranging from a few months to several years prior to PPV. The vitreous samples were analyzed for 29 different inflammatory immune mediators using multiplex bead immunoassays. A total of 14 cytokines (eotaxin, interferon-γ-induced protein-10 [IP-10], monocyte chemotactic protein-1 [MCP-1], macrophage derived chemokine [MDC], macrophage inflammatory protein [MIP]-1α, MIP-1β, thymus activation regulated chemokine [TARC], IL-12p40, IL-15, IL-16, IL-7, VEGF, IL-6, and IL-8) were detected in the vitreous of both study groups. Using multiple linear regression analysis, pseudophakia was significantly correlated with higher levels of vitreous immune mediators compared to phakia. Elevated vitreous levels were estimated to decrease over time for IL-6, IL-8, IL-15, IL-16, and VEGF, though they remained elevated for many months and even years compared to the levels detected in phakic eyes. This is the first study to demonstrate that cataract surgery and pseudophakia can induce increased vitreous levels of a substantial range of inflammatory immune mediators. The elevated levels seem to be maintained for a long period of time. These increased levels of cytokines may be involved in inflammatory processes leading to several complications to cataract surgery, both early and late.

Increased expression of angiogenic and inflammatory proteins in the vitreous of patients with ischemic central retinal vein occlusion.

BackgroundCentral retinal vein occlusion (CRVO) is a common disease characterized by a disrupted retinal blood supply and a high risk of subsequent vision loss due to retinal edema and neovascular disease. This study was designed to assess the concentrations of selected signaling proteins in the vitreous and blood of patients with ischemic CRVO.MethodsVitreous and blood samples were collected from patients undergoing surgery for ischemic CRVO (radial optic neurotomy (RON), n = 13), epiretinal gliosis or macular hole (control group, n = 13). Concentrations of 40 different proteins were determined by an ELISA-type antibody microarray.ResultsExpression of proteins enriched in the vitreous (CCL2, IGFBP2, MMP10, HGF, TNFRSF11B (OPG)) was localized by immunohistochemistry in eyes of patients with severe ischemic CRVO followed by secondary glaucoma. Vitreal expression levels were higher in CRVO patients than in the control group (CRVO / control; p < 0.05) for ADIPOQ (13.6), ANGPT2 (20.5), CCL2 (MCP1) (3.2), HGF (4.7), IFNG (13.9), IGFBP1 (14.7), IGFBP2 (1.8), IGFBP3 (4.1), IGFBP4 (1.7), IL6 (10.8), LEP (3.4), MMP3 (4.3), MMP9 (3.6), MMP10 (5.4), PPBP (CXCL7 or NAP2) (11.8), TIMP4 (3.8), and VEGFA (85.3). In CRVO patients, vitreal levels of CCL2 (4.2), HGF (23.3), IGFBP2 (1.23), MMP10 (2.47), TNFRSF11B (2.96), and VEGFA (29.2) were higher than the blood levels (vitreous / blood, p < 0.05). Expression of CCL2, IGFBP2, MMP10, HGF, and TNFRSF11B was preferentially localized to the retina and the retinal pigment epithelium (RPE).ConclusionProteins related to hypoxia, angiogenesis, and inflammation were significantly elevated in the vitreous of CRVO patients. Moreover, some markers known to indicate atherosclerosis may be related to a basic vascular disease underlying RVO. This would imply that local therapeutic targeting might not be sufficient for a long term therapy in a systemic disease but hypothetically reduce local changes as an initial therapeutic approach.

Low blood and vitreal BDNF, LXA4 and altered Th1/Th2 cytokine balance are potential risk factors for diabetic retinopathy

ObjectiveThe study was conducted to observe the serum and vitreous levels of LXA4, BDNF and Th1/Th2 cytokines in type 2 diabetes mellitus (DM) and changes associated with diabetic retinopathy (DR). Further, the in vitro study was performed to analyze the exposure of BDNF and LXA4 on LPS-induced pro-inflammatory state in ARPE 19 cells. Materials and MethodsTotally 114 individuals were recruited in a prospective case control study. Of these, 27 were type 2 DM cases with no complications, 30 cases were type 2 DM with non proliferative diabetic retinopathy (NPDR), 30 were type 2 DM with proliferative diabetic retinopathy (PDR), and 27 were healthy control. ELISA was done to estimate the serum and vitreous levels of BDNF, VEGF and PEDF. FACS cytometric Bead Array system was used to analyze the serum cytokines. ResultsThe serum BDNF and LXA4 levels were significantly reduced in both NPDR and PDR cases compared to control (p=0.005, 0.01; p=0.033, 0.015). Serum IL-6 was significantly increased in the PDR group (p=0.04). BDNF showed a significant negative correlation with VEGF levels (r=−0.522, p<0.01) and positive correlation with IL-10 (r=0.67, p<0.05) in serum. A significant odds ratio for the serum BDNF (OR: 3.20, p=0.025) as well as serum IL-6 (OR: 1.244, p=0.042) indicated them as potential risk factors for progression of type 2 DM to DR. A significant decrease in both the LXA4 (p=0.013) and BDNF (p=0.0008) with increase in cytokines IL-6 and IL-10 levels were observed in the vitreous of PDR cases ((p=0.04, 0.01). In vitro studies showed that both LXA4 (10nmol/L) and BDNF (500pg) decreased the IL-6 levels (p=0.036, 0.0002), in LPS induced pro-inflammatory condition in ARPE 19 cells, thereby their anti-inflammatory effect. ConclusionsThis study reports that low serum BDNF and higher IL-6 levels are potential risk factors for DR in type 2 DM. This study supports the role of BDNF in modulating the pro- and anti-inflammatory cytokines, and low level of BDNF is associated with development of diabetic retinopathy.

Vitreous estrogen levels in patients with an idiopathic macular hole.

PurposeEstrogen, a female hormone, activates collagenase and might be associated with the pathogenesis of vitreoretinal collagen fiber disease. The purpose of the present study was to investigate the vitreous levels of estrone (E1) and estradiol (E2) in subjects with an idiopathic macular hole (IMH).MethodsVitreous samples were obtained from ten female patients with an IMH and from nine female patients with other retinal diseases (six with rhegmatogenous retinal detachment and three with age-related macular degeneration) as a control at the time of vitreous surgery. E1 and E2 levels in the vitreous samples were then determined using the Coat-A-Count® Estradiol Radioimmunoassay (RIA) Kit and the DSL-70 Estrone RIA Kit, respectively.ResultsThe mean vitreous levels of E1 and E2 in the subjects with IMH were 1.83±2.00 pg/mL and 7.03±2.97 pg/mL, respectively, whereas in the control subjects they were 2.42±1.25 pg/mL and 4.90±2.90 pg/mL, respectively. Thus, the vitreous E2 levels in the subjects with IMH were significantly higher than in the controls (P<0.05).ConclusionThe findings of this study suggest that E2 might be associated with the pathogenesis of IMH, but further investigation is needed to elucidate that association.

Measurement of serum and vitreous concentrations of anti-type II collagen antibody in diabetic retinopathy.

BackgroundAutoimmune mechanisms have been postulated as a cause of diabetic retinopathy (DR), as several autoantibodies have reportedly been detected in the serum of DR patients. In this present study, we measured serum and vitreous levels of anti-type II collagen (anti-II-C) antibodies in DR patients and investigated their association with the mechanism of development of DR.Patients and methodsBlood samples were obtained from patients with proliferative DR and from patients with diabetic macular edema who underwent vitrectomy at Osaka Medical College, Takatsuki City, Osaka, Japan. Diabetic patients without DR were also included. The control group consisted of age- and sex-matched patients with noninflammatory eye diseases who underwent eye surgery for retinal detachment or for cataracts. The levels of anti-II-C immunoglobulin (Ig)G antibody in the vitreous and serum were measured using a human/monkey anti-II-C IgG assay kit.ResultsThe serum levels of anti-II-C IgG antibody were significantly higher in the DR patients than in the patients with noninflammatory eye disease (56.8±33.8 units/mL versus 30.5±13.7 units/mL, respectively; P<0.05, Fisher’s exact test). These levels were also significantly higher in the diabetic patients without DR than in the patients with noninflammatory eye disease (76.3±49.7 units/mL versus 30.5±13.7 units/mL, respectively; P<0.01, Fisher’s exact test). However, anti-II-C IgG antibody levels were unable to be detected in all of the obtained vitreous fluid samples.ConclusionThe development and progression of DR may be related to a mechanism involving intraocular type II collagen, which normally has immunological tolerance as a sequestered antigen. In DR, the disruption of the blood–retinal barrier leads to contact between the intraocular type II collagen and immunocompetent cells, and to subsequent activation of the autoimmune mechanism.