The aqueous humour dynamics in primary angle closure disease: a computational study.

PURPOSE: To create a computational fluid dynamics (CFD) model of ocular anterior segment for primary angle closure diseases (PACD) and assess the aqueous humour (AH) dynamics in different angle closure ranges (ACRs). METHODS: The ocular anterior segment geometry was obtained from an optical coherence tomography image by SOLIDWORKS. Three different angle opening distance at 750 µm from the scleral spur (AOD750) values were established to mimic three widths of anterior chamber angle. The AH dynamics were modelled using the Navier-Stokes equation. The 3D CFD model of the ocular anterior segment was created in COMSOL Multiphysics. The major outcome was the maximum flow velocity (MFV) and pressure in the ocular anterior segment. An in vitro simulation model was used to validate the computational results of the pressure and ACRs. RESULTS: The MFV and pressure both showed a non-linear association with ACR in the CFD models of PACD. The MFV and pressure started to elevate when ACR was larger than 180°, and increased dramatically when the ACR was larger than 270°. The in vitro experiment of the pressure changes was consistent with the CFD model. No significant differences of the MFV and pressure among the three AOD750 models. CONCLUSIONS: The association among the ACR, MFV and pressure is an ascending curve in PACD, and ACR of 180° and 270° are two critical turning points. Our results are consistent with clinical phenomenon and may be used to provide better guidances for the clinical management of PACD in different stages.

Role of N6-methyladenosine-related lncRnas in pseudoexfoliation glaucoma.

To explore the role of lncRNA m6A methylation modification in aqueous humour (AH) of patients with pseudoexfoliation glaucoma (PXG). Patients with open-angle PXG under surgery from June 2021 to December 2021 were selected. Age- and gender-matched patients with age-related cataract (ARC) were chosen as control. Patients underwent detailed ophthalmic examinations. 0.05-0.1 ml AH were extracted during surgery for MeRIP-Seq and RNA-Seq. Joint analysis was used to screen lncRNAs with differential m6A methylation modification and expression. Online software tools were used to draw lncRNA-miRNA-mRNA network (ceRNA). Expression of lncRNAs and mRNAs was confirmed using quantitative real-time PCR. A total of 4151 lncRNAs and 4386 associated m6A methylation modified peaks were identified in the PXG group. Similarly, 2490 lncRNAs and 2595 associated m6A methylation modified peaks were detected in the control. Compared to the ARC group, the PXG group had 234 hypermethylated and 402 hypomethylated m6A peaks, with statistically significant differences (| Fold Change (FC) |≥2, p < 0.05). Bioinformatic analysis revealed that these differentially methylated lncRNA enriched in extracellular matrix formation, tight adhesion, TGF- ß signalling pathway, AMPK signalling pathway, and MAPK signalling pathway. Joint analysis identified 10 lncRNAs with differential m6A methylation and expression simultaneously. Among them, the expression of ENST000000485383 and ROCK1 were confirmed downregulated in the PXG group by RT-qPCR. m6A methylation modification may affect the expression of lncRNA and participate in the pathogenesis of PXG through the ceRNA network. ENST000000485383-hsa miR592-ROCK1 May be a potential target pathway for further investigation in PXG m6A methylation.

Aqueous humour protein dysregulation in Asian eyes with primary open angle glaucoma.

The pathophysiology of Primary Open Angle Glaucoma (POAG) remains poorly understood. Through proteomic analysis of aqueous humour (AH) from POAG patients, we aim to identify changes in protein composition of these samples compared to control samples. High resolution mass spectrometry-based TMT6plex quantitative proteomics analysis is performed on AH samples collected from POAG patients, and compared against a control group of patients with cataracts. Data are available via ProteomeXchange with identifier PXD033153. 1589 proteins were quantified from the aqueous samples using Proteome Discoverer version 2.2 software. Among these proteins, 210 were identified as unique master proteins. The proteins which were up or down-regulated by ±3 fold-change were considered significant. Human neuroblastoma full-length cDNA clone CS0DD006YL02 was significantly upregulated in patients with severe POAG on >2 medications, while actin, cytoplasmic 1, V2-7 protein (fragment), immunoglobulin-like polypeptide 1 and phosphatidylethanolamine-binding protein 4 were only present in these patients with severe POAG on >2 medications. Beta-crystallin B1 and B2, Gamma-crystallin C, D and S were significantly downregulated in the severe POAG ≤2 glaucoma medications group. Beta-crystallin B2, Gamma-crystallin D and GCT-A9 light chain variable region (fragment) were significantly downregulated in the non-severe POAG group. Actin, cytoplasmic 1 was significantly upregulated in subjects with severe POAG who required more than 2 glaucoma medications. Crystallins (Beta-crystallin B1 and B2, Gamma-crystallin C, D and S) were significantly downregulated in subjects with severe POAG who required less than 2 glaucoma medications.

Genetic Diagnosis of Retinoblastoma Using Aqueous Humour-Findings from an Extended Cohort.

The identification of somatic RB1 variation is crucial to confirm the heritability of retinoblastoma. We and others have previously shown that, when tumour DNA is unavailable, cell-free DNA (cfDNA) derived from aqueous humour (AH) can be used to identify somatic RB1 pathogenic variation. Here we report RB1 pathogenic variant detection, as well as cfDNA concentration in an extended cohort of 75 AH samples from 68 patients. We show cfDNA concentration is highly variable and significantly correlated with the collection point of the AH. Cell-free DNA concentrations above 5 pg/µL enabled the detection of 93% of known or expected RB1 pathogenic variants. In AH samples collected during intravitreal chemotherapy treatment (Tx), the yield of cfDNA above 5 pg/µL and subsequent variant detection was low (≤46%). However, AH collected by an anterior chamber tap after one to three cycles of primary chemotherapy (Dx1+) enabled the detection of 75% of expected pathogenic variants. Further limiting our analysis to Dx1+ samples taken after ≤2 cycles (Dx ≤ 2) provided measurable levels of cfDNA in all cases, and a subsequent variant detection rate of 95%. Early AH sampling is therefore likely to be important in maximising cfDNA concentration and the subsequent detection of somatic RB1 pathogenic variants in retinoblastoma patients undergoing conservative treatment.

Advancing Personalized Medicine by Analytical Means: Selection of Three Metabolites That Allows Discrimination between Glaucoma, Diabetes, and Controls.

This paper aimed at devising an intelligence-based method to select compounds that can distinguish between open-angle glaucoma patients, type 2 diabetes patients, and healthy controls. Taking the concentration of 188 compounds measured in the aqueous humour (AH) of patients and controls, linear discriminant analysis (LDA) was used to identify the right combination of compounds that could lead to accurate diagnosis. All possibilities, using the leave-one-out approach, were considered through ad hoc programming and in silico massive data production and statistical analysis. Our proof of concept led to the selection of four molecules: acetyl-ornithine (Ac-Orn), C3 acyl-carnitine (C3), diacyl C42:6 phosphatidylcholine (PC aa C42:6), and C3-DC (C4-OH) acyl-carnitine (C3-DC (C4-OH)) that, taken in combination, would lead to a 95% discriminative success. 100% success was obtained with a non-linear combination of the concentration of three of these four compounds. By discarding younger controls to adjust by age, results were similar although one control was misclassified as a diabetes patient. Methods based on the consideration of individual clinical chemical parameters have limitations in the ability to make a reliable diagnosis, stratify patients, and assess disease progression. Leveraging human AH metabolomic data, we developed a procedure that selects a minimal number of metabolites (3-5) and designs algorithms that maximize the overall accuracy evaluating both positive predictive (PPV) and negative predictive (NPV) values. Our approach of simultaneously considering the levels of a few metabolites can be extended to any other body fluid and has potential to advance precision medicine. Artificial intelligence is expected to use algorithms that use the concentration of three to five molecules to correctly diagnose diseases, also allowing stratification of patients and evaluation of disease progression. In addition, this significant advance shifts focus from a single-molecule biomarker approach to that of an appropriate combination of metabolites.

Low expression of TGF-ß2 and matrilin2 in human aqueous humour with acute primary angle closure.

Primary angle-closure glaucoma (PACG) is the leading cause of irreversible blindness in the world. Angle closure induced by pupil block and secondary iris synechia is the fundamental pathology of the PACG. The molecular mechanisms of angle closure have not yet been clearly illustrated. This study was designed to investigate the protein difference in the aqueous humour and explore new biomarker of the PACG. Aqueous humour (AH) was collected from patients with acute primary angle closure (APAC) and cataract (n = 10 in APAC group) and patients with cataract only (n = 10 in control group). Samples were pooled and measured using label-free proteome technology. Then, the differentially expressed proteins (DEPs) were verified by ELISA using independent AH samples (n = 20 each group). More than 400 proteins were revealed in both groups through proteomics. Comparing the two groups, there were 91DEPs. These proteins participate in biological activities such as inflammation, fibrosis, nerve growth and degeneration and metabolism. We found that the expression of transforming growth factor-ß2 and matrilin2 was downregulated in the APAC group. The two proteins are related to inflammation and extracellular matrix formation, which might be involved in angle closure. This study characterized DEPs in AH of the APAC and found a downregulated protein matrilin2.

Correlating somatic copy number alteration in aqueous humour cfDNA with chemotherapy history, eye salvage and pathological features in retinoblastoma.

BackgroundThis study determined to probe the potential association between somatic copy number alteration (SCNA) in retinoblastoma (RB) aqueous humour (AH) and pathological high-risk factors, clinical features and previous chemotherapy history. METHODS: Single-centre retrospective cohort study from including 58 AH samples collected from 58 patients diagnosed. Among them, 41 samples were collected after enucleation and 17 samples were collected before intravitreal chemotherapy. SCNAs were accessed by conducting shallow whole-genome sequencing in cell-free (cf) DNA of AH. HRs and ORs were applied to measure risk factors. RESULTS: Canonical RB SCNAs including 1q gain (87%), 2p gain (50%), 6p gain (76%), 16q loss (69%) were frequently detected. Non-classical RB SCNAs in AH including 17q gain (53%), 19q loss (43%), 7q gain (35%) were also commonly observed. 19q loss was significantly more common in patients with cT3c or worse stage than others (p=0.034). 2p gain(p=0.001) and 7q gain(p=0.001) were both more common in patients with primary enucleation than those with previous chemotherapy. Interestingly, both 2p gain (HR=1.933, p=0.027) and 7q gain (HR=2.394, p=0.005) might predict enucleation. Correlation analysis with pathological features among enucleated eyes showed that 19q loss can predict a higher risk for both massive choroid invasion (OR=4.909, p=0.038) and postlaminar optic nerve invasion (OR=4.250, p=0.043). DISCUSSION: Sequencing of AH cfDNA in RB can provide sufficient in vivo information. 19q loss was a potential signature of advanced cases clinically and pathologically.Repeated sampling from eyes receiving sequential chemotherapy should be conducted to evaluate fluctuation of SCNA in future study.

Intraocular pressure variation from ocular compression in low and high myopia.

CLINICAL RELEVANCE: Change in intraocular pressure during acute ocular compression is related to aqueous humour dynamics. Monitoring intraocular pressure (IOP) change throughout ocular compression has potential to evaluate aqueous outflow facilities. BACKGROUND: Recent studies have monitored lamina cribrosa deformation using optical coherence tomography during ocular compression. IOP was measured only once immediately after ocular compression. This study aimed to evaluate IOP changes during and after ocular compression and compare the differences between low and high myopia. METHODS: Two groups of young, healthy adults were age-matched and underwent ocular compression. IOP was measured at baseline and monitored during a 2-min ocular compression followed by a 10-min recovery phase. Rebound tonometry was used and applied at 30-s intervals. RESULTS: Thirty low and 30 high myopes (60 right eyes) were included in the study. They had similar baseline IOP at 14.9 mmHg. IOP was elevated to 21.7 ± 3.8 mmHg and 22.3 ± 4.2 mmHg for the low and high myopic group, respectively (p = 0.877). Low myopes had faster IOP decay during ocular compression at -3.24 mmHg/min than high myopes at -2.58 mmHg/min (p = 0.0528). The IOP dropped below the baseline level after the release of the compressive force. Low myopes had IOP that returned to baseline levels faster (at 360 s) than high myopes (at 510 s). CONCLUSION: Measuring IOP once immediately after ocular compression could under-estimate the effect of IOP elevation during ocular compression. Further studies are required regarding IOP changes from ocular compression in aqueous humour dynamics.

Anterior segment involvement in vitreoretinal lymphoma: clinical manifestations, molecular findings and in vivo confocal microscopy.

BACKGROUND: Intermediate and posterior manifestations of vitreoretinal lymphoma (VRL) are well characterised. However, there is limited information on anterior segment involvement in VRL. This study aimed to describe the anterior manifestations of VRL, and their association with molecular testing. METHODS: Retrospective analysis of patients with biopsy-proven VRL. Study variables included anterior segment manifestations, findings from slit-lamp photos and in vivo confocal microscopy (IVCM) when available. MYD88 L265P mutation and cytology in the aqueous humour, retinal and systemic findings were also analysed. RESULTS: The analysis included 108 eyes of 55 VRL patients. Anterior segment involvement was present in at least one visit in 55 eyes (51%) of 33 patients (60%); it included keratic precipitates (dendritiform with branching and irregular margins in 33 eyes, dust-like in 16 eyes and large granulomatous in 12 eyes), cells in the anterior chamber (51 eyes) and posterior synechiae (2 eyes). IVCM was available for 41 eyes and showed different morphologies of keratic precipitates, including floral, spikes and mulberry patterns (66%, 56% and 20%, respectively). MYD88 L265P mutation in the aqueous humour was detected in 10/21 (48%) eyes with no anterior segment involvement and 24/37 (65%) eyes with anterior segment involvement. CONCLUSIONS: Anterior segment manifestations are often present in VRL and include dendritiform and dust-like keratic precipitates. IVCM in VRL can identify different patterns associated with keratic precipitates. MYD88 L265P mutation in the aqueous humour of VRL patients can also be found in eyes without significant anterior segment involvement.

Biotissue stent for supraciliary outflow in open-angle glaucoma patients: surgical procedure and first clinical results of an aqueous drainage biostent.

BACKGROUND/AIMS: To report a first-in-human trial in open-angle glaucoma (OAG) subjects treated with a new microinterventional biostent-reinforced cyclodialysis technique to enhance supraciliary aqueous drainage. METHODS: Subjects (N=10; 74.1±7.9 years old) with OAG and cataracts underwent combined phacoemulsification cataract surgery with implantation of a permanent endoscleral supraciliary biostent to reinforce a controlled cyclodialysis cleft. The biostent comprised decellularised scleral allograft tissue microtrephined into a polymer tubular implant intraoperative/postoperative safety, intraocular pressure (IOP) and glaucoma medications were tracked through 12 months postimplantation. RESULTS: Baseline medicated IOP averaged 24.2±6.9 mm Hg with subjects using 1.3±0.8 IOP-lowering medications. Successful biostent implantation was achieved in all individuals without significant complications. Immediate IOP lowering was sustained through 1 year. Twelve-month mean IOP was reduced 40% from baseline to 14.6±3.2 mm Hg (p=0.004; paired two-tailed t-test), and 80% of patients achieved >20% IOP reduction. Biostenting reduced glaucoma medication use 62%, from a baseline mean of 1.3 required medications to 0.5 medications (p=0.037) at postoperative 12 months. The biotissue implant was well tolerated and demonstrated good endothelial safety with only 11% endothelial cell loss at 12 months after combined phaco-biostenting surgery, similar to that expected after phacoemulsification alone. Mean BCVA increased from baseline 20/130 Snellen to 20/36 at postoperative 12 months (p=0.001). CONCLUSION: Supraciliary biostenting in OAG patients is well tolerated, has a good safety profile and produces long-term IOP-lowering while reducing glaucoma medication requirements.

Evaluating the long-term biological stability of cytokine biomarkers in ocular fluid samples.

PURPOSE: The quality of biological fluid samples is vital for optimal preanalytical procedures and a requirement for effective translational biomarker research. This study aims to determine the effects of storage duration and freeze-thawing on the levels of various cytokines in the human aqueous humour and vitreous samples. METHODS AND ANALYSIS: Human ocular aqueous humour and vitreous samples were obtained from 25 eyes and stored at -80°C for analysis. All samples were assayed for 27 cytokine biomarker concentrations (pg/mL) using a multiplex assay. Four sample storage durations following sample collection were evaluated (1 week, 3 months, 9 months and 15 months). Additionally, samples underwent up to three freeze-thaw cycles within the study period. RESULTS: Among the 27 cytokine biomarkers, concentrations of four cytokines (Interleukin (IL)-2, IL-10, IL-12 and platelet-derived growth factor-BB) were significantly decreased by storage duration at all time points, as early as 3 months following sample collection (range of 9%-37% decline between 1 week and 15 months, p<0.001). Freeze-thawing of up to three cycles did not significantly impact the cytokine biomarker concentrations in aqueous humour or vitreous. Separability of patient-specific cytokine biomarker profiles in the principal component analysis remained relatively the same over the 15 months of storage duration. CONCLUSION: The findings from this study suggest that several intraocular cytokine biomarkers in human aqueous humour and vitreous samples may be susceptible to degradation with long-term storage, as early as 3 months after collection. The overall patient-specific cytokine biomarker profiles are more stable than concentrations of individual cytokines. Future studies should focus on developing guidelines for optimal and standardised sample handling methods to ensure correct research findings about intraocular biomarkers are translated into clinical practice.

Advancement in Understanding Glaucoma: A Comprehensive Review.

Glaucoma, a silent thief of sight, remains a significant cause of irreversible blindness due to a substantial number of undiagnosed and untreated cases. To combat this insidious disease effectively, a multifaceted approach is imperative. Early detection is paramount in the battle against glaucoma. Patient history, including family history, plays a pivotal role in identifying those at risk. A comprehensive understanding of a patient's genetic predisposition can significantly enhance the accuracy of diagnosis and detection of suspicious cases. Treatment options include prescription eye drops, oral medicines, laser treatment, surgery, or a combination of approaches. Trabeculectomy involves the surgical creation of an aqueous humor drainage channel, while laser trabeculoplasty enhances aqueous outflow by modifying the trabecular meshwork. However, these procedures pose certain risks and complications. Exploration of alternative treatments with lower risks is underway. These innovative approaches hold promise in reducing the burdens associated with conventional treatments such as trabeculectomy. However, the effectiveness of these alternatives in the long term remains a subject of ongoing research. Neuroprotective drugs have also been in development to halt the progression of glaucoma. However, their success remains uncertain due to challenges, such as a lack of understanding of the underlying mechanisms, scarcity of suitable drugs, and regulatory hurdles in gaining approval. In essence, the overarching goal of glaucoma therapy is to reduce intraocular pressure through various means - medications, laser procedures, or innovative methods. The aim is to slow down the disease's progression, thereby preserving vision and improving the patient's quality of life. In conclusion, addressing the challenge of glaucoma requires a comprehensive approach encompassing early detection, innovative treatments, and ongoing research into potential cures. Only through concerted efforts can we hope to reduce the impact of this sight-stealing disease on individuals and society as a whole.

Vascular Cell Adhesion Molecule-1 (VCAM-1) contributes to macular fibrosis in neovascular age-related macular degeneration through modulating macrophage functions.

BACKGROUND: Neovascular age-related macular degeneration (nAMD) is a major cause of blindness in the elderly. The disease is due to the growth of abnormal blood vessels into the macula, leading to the loss of central vision. Intravitreal injection of vascular endothelial growth factor (VEGF) inhibitors (e.g., anti-VEGF) is the standard of care for nAMD. However, nearly 50% of patients do not respond or respond poorly to the therapy. More importantly, up to 70% of nAMD patients develop macular fibrosis after 10 years of anti-VEGF therapy. The underlying mechanism of nAMD-mediated macular fibrosis is unknown although inflammation is known to play an important role in the development of abnormal macular blood vessels and its progression to fibro-vascular membrane. In this study, we measured the intraocular levels of adhesion molecule VCAM-1, ICAM-1, CD44, CD62L, and CD62P in nAMD patients with and without macular fibrosis and investigated the link between the levels of adhesion molecule and clinical features (e.g., visual improvement, retinal thickness, etc.). We further investigated the effect of VCAM-1 in macrophage function in vitro and the development of subretinal fibrosis in vivo using a two-stage laser-induced protocol. RESULTS: The aqueous levels of ICAM-1, VCAM-1, CD44, and CD62L were significantly higher in nAMD patients compared to cataract controls. The aqueous level of VCAM-1 (but not other adhesion molecules) was significantly higher in patients with macular fibrosis than those without and the level correlated positively with the retinal thickness. VCAM-1 was highly expressed at the lesion site in the mouse model of subretinal fibrosis. Blocking VCAM-1 or its receptor VLA-4 significantly prevented macrophage infiltration and reduced subretinal fibrosis in vivo. VCAM-1 induced macrophage migration and upregulated the expression of Arg-1, Mmp12 and Il6 but down-regulated the expression of iNOS and Il1b in macrophages. CONCLUSIONS: VCAM-1 may contribute to the development of macular fibrosis in nAMD patients by modulating macrophage functions, including migration and profibrotic polarization.

Study of aqueous humour inflammatory mediators' levels in a cohort of Egyptian patients with diabetic macular oedema.

BACKGROUND: The aim was to study aqueous humour inflammatory mediators' levels in a cohort of Egyptian patients with diabetic macular oedema (DMO). METHODS: This was a case-control prospective study conducted on 2 groups: 25 eyes of 22 (11 females) patients seeking treatment for DMO as patients group, and 10 eyes of 10 (4 females) cataract patients as a control group. Aqueous humour was aspirated before intravitreal injection (patients' group) or cataract surgery (control group). Inflammatory mediators in aqueous humour were measured using a multiplex bead immunoassay kit of 27 pre-mixed cytokines. RESULTS: Eotaxin, interferon gamma-induced protein 10 (IP-10), monocyte chemoattractant protein-1 (MCP-1/CCL2) and interleukin-8 (IL-8/CXCL8) were found significantly higher in patients' group compared to control group (p = 0.043, 0.037, 0.001, 0.015 respectively). On the contrary, interferon-gamma (IFN-gamma) and granulocyte colony-stimulating factor (G-CSF) were found significantly higher in control group than patients' group (p = 0.003, 0.019 respectively). Basic fibroblast growth factor (Basic-FGF/FGF-2) and interleukin-1 receptor antagonist (IL-1ra) were found higher (but not statistically significant) in controls (p = 0.100 and 0.070 respectively). Additionally, a negative and significant correlation was found between Eotaxin level in aqueous humour and central macular thickness. CONCLUSIONS: Some mediators might be implicated in the pathogenesis of DMO either augmenting or suppressing role. Eotaxin, IP-10, MCP-1 and IL-8 might have a role in cases not responding to standard anti-vascular endothelial growth factor (VEGF) therapy. IL-1ra might have a protective role; therefore, the effectiveness of intravitreal injection of IL-1ra homologue needs to be studied in future clinical trials.

False negative result of polymerase chain reaction in very early stages of acute retinal necrosis.

BACKGROUND: Viral nucleic acid testing of intraocular fluid using polymerase chain reaction (PCR) is a major laboratory examination in the diagnosis of acute retinal necrosis (ARN). Importantly, false negative PCR results may occur in several special situations. We reported a case of ARN with a negative PCR result in the aqueous humour in the very early stages of disease. CASE PRESENTATION: A female patient presented to the ophthalmologist with complaints of blurred vision and redness in her left eye. Her medical history included ARN in her right eye 10 years prior. Although the result of the aqueous viral analysis by PCR in her left eye was negative the first time (one day after the appearance of ocular symptoms), ARN in her left eye was presumed based on the clinical signs. With timely antiviral and anti-inflammatory treatments, the retinal lesions diminished. The viral load of herpes simplex virus (HSV) turned positive (7.25 × 103 copies/mL) one week later, increased to 2.49 × 105 copies/mL after three weeks, and finally turned negative about five weeks after the onset of disease. The initial HSV-IgG level in the aqueous humour was 0.01 U/mL and increased to 222.64 U/mL in the final sampling. CONCLUSIONS: The results of PCR analysis can be negative in the very early stages of ARN. Diagnosis of ARN should be made based on the clinical features, and antiviral treatments should not be delayed. Repeated PCR analysis of the aqueous humour is necessary to confirm the diagnosis and monitor the disease process.

Proteomic study of aqueous humour in diabetic patients with cataracts by TMT combined with HPLC-MS/MS.

BACKGROUND: The purpose of this study is to identify the proteomic differences between the aqueous humour of diabetes patients with cataracts and that of non-diabetic sufferers of cataracts in a clinical setting. METHODS: Patients were divided into the diabetic experimental group and the non-diabetic control group. Aqueous humour specimens were obtained via cataract surgery. Sample proteins were treated with a TMT reagent, separated using a cation chromatography column, and analysed using a C18 desalting column. Proteins were identified using HPLC-MS/MS. The differential proteins were identified using both a p value of < 0.05 and a fold change of > 1.2. GO classification enrichment analysis, KEGG pathway enrichment analysis, protein interaction network analysis, and ingenuity pathway analysis were all carried out. The expression level of four differential proteins were verified by Western blot, and GC and TTR expressions were further examined using an expanded sample pool. RESULTS: The postprandial glucose levels between the experimental group (9.40 ± 1.35 mmol/L) and the control group (6.56 ± 0.81 mmol/L) were significantly different, with a p value of 1.16E-06. It is important to note, however, that the baseline levels of the parameters showed no statistical differences. In total, 397 aqueous humour proteins were identified; of these, 137 showed significant differences, with 63 upregulated ones and 74 down-regulated ones. The differential proteins play important roles in numerous biological processes and pathways, such as complement and coagulation cascades (p = 1.71E-09). Some of these differential proteins are associated with diabetic retinal degeneration and other diabetic complications. Differential proteins, such as HP, GC, and TTR, have high node degree in the protein interaction network. Western blot results further confirmed that GC were down-regulated while TTR was up-regulated in aqueous humour under diabetic condition. CONCLUSION: A list of differential proteins in the human aqueous humour of diabetic patients was established. Proteins with high interaction scores as per protein interaction analysis, such as GC and TTR, were further verified and could potentially be used as early diagnostic markers for diabetic eye complications in clinical practice.

Intraocular complement activation is related to retinal vascular and neuronal degeneration in myopic retinopathy.

Purpose: To investigate the relationship between the intraocular levels of complement proteins and myopia-related retinal neuronal and vascular degeneration. Methods: Aqueous humour from 147 myopic patients, including 60 low-myopia and 87 high-myopia were collected during Implantable Collamer Lens implantation surgery. All participants received comprehensive ophthalmic examinations, including logMAR best corrected visual acuity, axial length measurement, fundus photography and ocular B-scan ultrasonography. The myopic eyes were further classified into simple myopia (SM, n = 78), myopic posterior staphyloma (PS, n = 39) and PS with myopic chorioretinal atrophy (PS + CA, n = 30). Retinal thickness and vascular density in the macula (6 mm × 6 mm) and optic nerve head (4.5 mm × 4.5 mm) were measured using Optical Coherence Tomography (OCT) and OCT angiography (OCTA). The levels of complement proteins including C1q, C3, C3b/iC3b, C4, CFB, CFH, C2, C4b, C5, C5a, CFD, MBL and CFI in the aqueous humour were measured using the Luminex Multiplexing system. The real-time RT-PCR was conducted to examine the expression of complement genes (C1q, C2, C3, C4, CFI and CFD) in the guinea pig model of long-term form deprivation-induced myopic retinal degeneration. Results: OCTA showed that retinal neuronal thickness and vascular density in superficial and deep layers of the macular zone as well as vascular density in the optic nerve head were progressively decreased from SM to PS and PS + CA (p < 0.05). The aqueous humour levels of C1q, C3, C3b/iC3b, C4, CFB, CFH, C2, C4b, C5 and CFI were significantly higher in high-myopic eyes compared to those in low-myopic eyes. Further subgroup analysis revealed the highest levels of complement components/fragments in the PS + CA group. The intraocular levels of complement factors particularly C3b/iC3b and C4 were negatively correlated with macular zone deep layer retinal thickness and vascular density and optic nerve head vascular density. The expression of C2, C3 and C4 genes was significantly higher in guinea pig eyes with myopic retinal degeneration compared to control eyes. Conclusions: The intraocular classical pathway and alternative pathway of the complement system are partially activated in pathological myopia. Their activation is related to the degeneration of retinal neurons and the vasculature in the macula and the vasculature in the optic nerve head.

Copper, Lysyl Oxidase Activity, and Collagen in Aqueous Humour of Primary Glaucoma: An Association with Clinical Parameters.

INTRODUCTION: To measure copper (Cu), lysyl oxidase (LOX) activity, and collagen levels in aqueous humour (AH) of primary glaucoma patients and correlate with clinical parameters. METHODS: 120 patients with 40 each of primary open angle glaucoma (POAG), primary angle closure glaucoma (PACG), and cataract controls were recruited in this case-control study. AH samples were collected during the trabeculectomy and cataract surgeries. Cu levels were measured using an atomic absorption spectrophotometer. LOX unit activity was determined by Amplex Red assay and collagen concentration by Sirius red assay. RESULTS: Significantly higher levels of Cu expressed as median (IQR) µmol/L were observed in POAG (p = 0.008) and PACG (p = 0.005) compared to controls. The LOX activity was increased in POAG and PACG (p = 0.04) compared to controls represented as median (IQR) µmol/min. The collagen levels given as median (IQR) mg/ml showed an insignificant increase in POAG and PACG compared to controls (p = 0.78). The LOX unit activity was correlated with visual field index (VFI), which showed a significant increase with the progression of the diseases (p < 0.05), whereas Cu levels were negatively correlated with LOX activity in AH. Cu and LOX activity showed weak correlation with YAG peripheral iridotomy (YAGPI), duration of anti-glaucoma medications, and highest preoperative intraocular pressure. CONCLUSION: Elevated Cu and LOX activity was observed in both POAG and PACG groups compared to controls. LOX activity showed notable increase with VFI as the severity of the disease. Although Cu levels are increased in glaucoma, it's insufficient to significantly increase the activity of LOX.

Concentrations of glucose metabolites in the aqueous humour of diabetic cataract eyes.

PURPOSE: Glucose metabolism underpins diabetic cataracts (DCs), but the relationship between the two remains unclear. Here, we tested the aqueous humour (AH) of patients with DCs to elucidate glucose metabolite levels. METHODS: In this study, aqueous humour (AH) samples were collected preoperatively from DC eyes (n = 37) and age-related cataract eyes (n = 37) from 74 patients (74 eyes) undergoing uncomplicated cataract surgery. The content of glucose, pyruvate, L-lactate were detected by biochemical methods and advanced glycation end-products (AGEs) was detemined using enzyme-linked immunosorbent assay (ELISA) technique. Furthermore, the ratios of glucose/pyruvate and L-lactate/pyruvate in the AH were calculated. In addition, we calculated the correlation between glucose levels and AGEs in the AH. RESULTS: The concentrations of glucose, pyruvate and AGEs in the DC group were higher than those in the control group. Significantly lower levels of L-lactate in the AH were found in the DC group. We calculated the glucose/pyruvate ratio and the L-lactate/pyruvate ratio in the AH, which showed that glucose metabolism was changed in the AH from DC patients. Interestingly, we observed that AGEs in the AH were significantly correlated with increased anterior chamber glucose permeability. A stronger correlation was found in the subgroups of male patients, younger patients, and patients with poor glycaemic control status. CONCLUSIONS: Comparison of the levels of glucose metabolism-related products in the AH in the DC group highlight a potential pathological mechanism for DC from a glucose metabolism perspective. The findings indicated an alteration in the metabolic pathways of energy metabolism and amino acids in the AH of DC patients.

Changes in Macular Thickness after Cataract Surgery in Patients with Open Angle Glaucoma.

BACKGROUND: The purpose of this study was to examine the changes in IOP, total macular and RNFL, ganglion cell layer (GCL) thickness, and aqueous humour flare in open angle glaucoma (OAG) patients before and 6 months after cataract surgery. METHODS: This was a prospective observational case-control age- and gender-matched study. Groups: 40 subjects in a controlled OAG (OAGc) group, 20 subjects in an uncontrolled OAG (OAGu) group, and 60 control group subjects. EXAMINATION: complete ophthalmic evaluation, IOP measurement, anterior and posterior segment Optical Coherence Tomography (OCT), and laser flare photometry before and 6 months postoperatively. RESULTS: Six months postoperatively IOP decreased in all groups. An increase in macular thickness was found postoperatively in all groups. Preoperative aqueous humour flare was higher in the OAGc group than in the control group. After cataract surgery, aqueous humour flare was higher in the control group compared to the preoperative result. CONCLUSIONS: Changes in IOP following cataract surgery were strongly negatively correlated with preoperative IOP. An increase in macular thickness was observed 6 months postoperatively in all groups. Aqueous humour flare did not differ in OAGc and OAGu groups pre- and postoperatively but significantly increased in the control group postoperatively.