Polarity-Driven Fluorescence Monitoring of Lipid Droplet Dynamics in Dry Eye Disease.

The emergence of lipid droplets (LDs) has been recognized as cellular markers of ocular surface hyperosmosis, which is recognized as a fundamental mechanism driving dry eye disease (DED), while their dynamics during DED progression and therapy remains unlocked. For this purpose, an LD-specific fluorescent probe P1 is presented in this work that exhibits highly selective and sensitive emission enhancement in response to a decreased ambient polarity (Δf) from 0.209 to 0.021. The hydrophobic nature of P1 enables specific staining of LDs, facilitating visualization of changes in polarity within these cellular structures. Utilizing P1, we observe a decrease in polarity accompanied by an increase in the size and number of LDs in hyperosmotic human corneal epithelial cells (HCECs). Furthermore, interplays between LDs and cellular organelles such as mitochondria and the Golgi apparatus are visualized, suggesting the underlying pathogenesis in DED. Notably, the variations of LDs are observed after the inhibition of ferroptosis or activation of autophagy in hyperosmotic HCECs, implying the great potential of LDs as indicators for the design and efficacy evaluation of DED drugs regarding ferroptosis or autophagy as targets. Finally, LDs are confirmed to be overproduced in corneal tissues from DED mice, and the application of clinical eye drops effectively impedes these changes. This detailed exploration underscores the significant roles of LDs as an indicator for the deep insight into DED advancement and therapy.

MiR-223 inhibits hyperosmolarity-induced inflammation through downregulating NLRP3 activation in human corneal epithelial cells and dry eye patients.

We previously showed that increases in reactive oxygen species (ROS) generation upregulate NLRP3 inflammasome and inflammation through increases in both caspase-1 activity and rises in IL-1ß expression levels in animal models of dry eye (DE). As changes in microRNA (miRNAs) expression levels can modulate inflammasome function, we determine here if there is a relationship in DE between changes in miR-223 expression levels and NLRP3 activation induced in an intelligent controlled environmental system (ICES) in mice. In parallel, ROS, miR-223 and NLRP3 expression levels were assessed in conjunctival impression cytology and tear fluid samples obtained from DE patients and normal subjects. MiR-223 expression levels were modulated by transfection of either a mimic or its negative control (NC) in a human corneal epithelial cell line (HCECs) exposed to a 500 mOsm hyperosmotic medium for 4 h. The dual-luciferase reporter assay confirmed that miR-223 controls NLRP3 gene expression readout through directly interacting with the 3' UTR of its mRNA. Hyperosmolarity-induced NLRP3 activation was confirmed based on recruitment and colocalization of NLRP3 with ASC as well as increases in IL-1ß expression. The miR-223 expression level decreased by 55% in the conjunctiva and cornea of the murine DE model from the level in the control group (P ≤ 0.047), while NLRP3 protein expression rose by 30% (P ≤ 0.017). In DE patients, miR-223 expression decreased in conjunctival impression cytology samples (P = 0.002), whereas IL-1ß tear content rose significantly (P < 0.001).The relevance of this decline was confirmed by showing that exposure to a 500 mOsm stress decreased the miR-223 expression level whereas ROS generation as well as the NLRP3, and IL-1ß expression levels rose in HCECs (P ≤ 0.037). In contrast, miR-223 mimic transfection reduced the NLRP3 protein expression level by 30% (P = 0.037), whereas both ROS generation and IL-1ß secretion rose compared to their corresponding levels in the control group (P ≤ 0.043). Thus, miR-223 negatively regulates NLRP3 inflammasome activity via suppressing NLRP3 translation in DE. This inverse regulation between miR-223 and NLRP3 expression levels suggests that selective upregulation of miR-223 expression may be a novel option to suppress chronic inflammation in DE.

Impact of cone base diameter on outcomes of deep anterior lamellar keratoplasty in keratoconus.

PURPOSE: To evaluate the outcomes of deep anterior lamellar keratoplasty (DALK) in treating keratoconus in relation to cone base diameter (CBD). METHODS: A retrospective study. Sixty-one eyes of 49 keratoconus patients who underwent DALK between 2009 and 2018 were enrolled. Preoperative and postoperative uncorrected visual acuity (UCVA), best-corrected visual acuity (BCVA), spherical equivalent, and astigmatism were measured. Scheimpflug tomography (Pentacam) was used to measure the cone base area (CBA) and CBD using MATLAB software. RESULTS: The mean age of the patients was 20.8 ± 6.1 years old, and the mean follow-up time was 27.3 ± 15.2 months. Mean UCVA improved from 1.23 ± 0.48 to 0.57 ± 0.27 (LogMAR, 95% CI [0.52, 0.80]; P < 0.001), whereas mean BCVA improved from 0.98 ± 0.55 to 0.18 ± 0.13 (95% CI [0.66, 0.94]; P < 0.001). The mean spherical equivalent decreased by 4.53 ± 5.65 D (95% CI [- 6.25, - 2.82]; P < 0.001), with little change in astigmatism (95% CI [- 1.39, 0.64]; P = 0.457). The postoperative BCVA in the patients with CBD < 5.07 mm and corneal curvature ≥ 55D was significantly better than those whose CBD ≥ 5.07 mm (0.14 ± 0.09 vs 0.25 ± 0.15, P = 0.001). The follow-up time was negatively correlated with the BCVA (P = 0.004). CONCLUSIONS: In this study, outcomes of DALK in keratoconus were related to CBD and corneal curvature. Patients with large CBD (≥ 5.07 mm) where the corneal curvature ≥ 55D are more likely to have poor visual outcomes after DALK.

Deep anterior lamellar keratoplasty with cross-linked acellular porcine corneal stroma to manage fungal keratitis.

PURPOSE: To evaluate the effects of deep anterior lamellar keratoplasty (DALK) with cross-linked acellular porcine corneal stroma (APCS) and post-operative topical tacrolimus treatment in patients with fungal keratitis. METHODS: This multicenter prospective study involved 25 cases of fungal keratitis that were treated by DALK with cross-linked APCSs and post-operative topical tacrolimus from December 2013 to November 2014 at the Wenzhou Eye Hospital and the Henan provincial Eye Hospital. Signs of post-operative inflammation, corneal reepithelialization, corneal neovascularization, and graft rejection were assessed, and best corrected visual acuity (BCVA), intraocular pressure (IOP), and APCS graft transparency were monitored for the 12-month follow-up period. RESULTS: All 25 patients underwent DALK without Descemet's membrane perforation. Corneal epithelium recovered completely in 17 patients in the first week, and APCS grafts maintained transparency in 18 patients at 1-year follow-up. The mean BCVA significantly improved from 2.16 ± 0.32 (LogMAR) at baseline to 1.56 ± 0.70 at 1-week (P < .001), 0.95 ± 0.57 at 1-month (P < .001), and 0.70 ± 0.51 at 3-month follow-ups (P < .001). The BCVA kept stable at 6-month and 12-month follow-ups. Post-operative topical tacrolimus alleviated the ciliary injection, except in one case which acute stromal rejection occurred. One patient developed fungal reinfection and underwent penetrating keratoplasty. Graft rejection occurred in three patients. No case was noted with graft splitting, elevated IOP or tacrolimus intolerance. CONCLUSIONS: DALK using cross-linked APCS combining topical tacrolimus treatment is safe and effective in managing fungal keratitis. It may ameliorate the shortage of corneal donation globally.

Substance P induces sympathetic immune response in the contralateral eye after the first eye cataract surgery in type 2 diabetic patients.

BACKGROUND: Substance P (SP) is a nociceptive tachykinin which regulates the immune inflammatory reactions including monocyte chemoattractant protein 1 (MCP-1) production. Sequential second-eye cataract surgery patients often suffer more pain than the first one partly because of the MCP-1 increase in aqueous humor (AH). This study aims to illustrate whether SP is involved in sympathetic inflammatory responses in the contralateral eye in patients with or without type 2 diabetes. METHODS: This prospective randomized clinical study included 51 cataract patients (22 with diabetes and 29 without). Bilateral sequential cataract surgeries were conducted with 1-day or 1-week interval randomly. More than 100 µl of AH were obtained before surgery and stored for later analysis using magnetic Luminex assays to detect interleukin (IL)-1ß, IL-1ra,IL-6, IL-8, IL-10, MCP-1, vascular endothelial growth factor, spinal macrophage inflammatory protein (MIP-1a), interferon-inducible protein 10 (IP-10), regulated on activation, normal T expressed and secreted (RANTES), as well as the enzyme-linked immunosorbent assay for SP. RESULTS: Among the 4 groups, no significant differences were found in age, sex distribution, the R/L ration of the first surgery eye, or the lens nucleus hardness (P ≥ 0.802). Over 100 µl of AH samples were collected safely in all cases without intraoperative complications. SP and MCP-1 levels were both increased significantly in the second eye of the diabetic patients with 1-day and 1-week interval (P ≤ 0.040). The SP increase in the second eye of the diabetic patients were significantly higher than that of the patients without diabetes (P ≤ 0.030) both in the groups with 1-day and 1-week interval. Similarly, the MCP-1 increase was significantly higher in the diabetic patients in the group with 1-week interval (P = 0.042). CONCLUSIONS: Substance P and MCP-1 productions elevated in the AH of the contralateral eye after the first-eye cataract surgery in diabetic patients, indicating that SP and MCP-1 were involved in the sympathetic inflammatory responses. Diabetic patients are susceptible to noninfectious inflammation after cataract surgery, and perceive more pain in the second-eye phacoemulsification. TRIAL REGISTRATION: Chinese Clinical Trial Registry, ChiCTR1900028374 , retrospectively registered on 20th December, 2019.

Insight into the distribution of metallic elements in membrane bioreactor: Influence of operational temperature and role of extracellular polymeric substances.

The distribution of metallic elements in a submerged membrane bioreactor (MBR) was revealed at different temperatures using inductively coupled plasma-optical emission spectrometry (ICP-OES), and the role of extracellular polymeric substances (EPS) was probed by integrating scanning electron microscopy (SEM) with confocal laser scanning microscopy (CLSM) over long-term operation. More metallic elements in the influent were captured by suspended sludge and built up in the fouling layer at lower temperature. The concentration of metallic elements in the effluent was 5.60mg/L at 10°C operational temperature, far lower than that in the influent (51.35mg/L). The total contents of metallic elements in suspended sludge and the membrane fouling layer increased to 40.20 and 52.19mg/g at 10°C compared to 35.14 and 32.45mg/g at 30°C, and were dominated by the organically bound fraction. The EPS contents in suspended sludge and membrane fouling layer sharply increased to 37.88 and 101.51mg/g at 10°C, compared to 16.87 and 30.03mg/g at 30°C. The increase in EPS content at lower temperature was responsible for the deposition of more metallic ions. The strong bridging between EPS and metallic elements at lower temperature enhanced the compactness of the fouling layer and further decreased membrane flux. This was helpful for understanding the mechanism of membrane fouling at different operational temperatures and the role of EPS, and also of significance for the design of cleaning strategies for fouled membranes after long-term operation.

Short-term effect of a developed warming moist chamber goggle for video display terminal-associated dry eye.

BACKGROUND: Video display terminal (VDT)-associated dry eye (DE) patients are the rising group worldwide, and moisture goggles are the preferable treatment since they are capable of improving tear film stability and DE discomfort. The current study aims to evaluate the short-term efficacy and safety of the developed warming moist chamber goggles (WMCGs) for VDT-associated DE patients. METHODS: In this prospective self-control study, 22 DE patients (22 eyes) working with VDTs over 4 h daily were enrolled and instructed to wear WMCGs for 15 min. Sodium hyaluronate (SH, 0.1%) eyedrops were applied as a control on another day on these same patients, however 4 subjects denied the eyedrop application. The symptomatology visual analog scale (VAS) score, tear meniscus height (TMH), noninvasive tear film break-up time (NI-BUT), tear film lipid layer thickness (LLT), and bulbar conjunctival redness were assessed with Keratograph 5 M at baseline, 5, 30 and 60 min after treatment. The WMCGs wearing comfort was also evaluated. RESULTS: The ocular discomfort evaluated by VAS decreased in the WMCGs group throughout 60 min (P<0.001), better than the control group levels (P ≤ 0.015). TMH, NI-BUT (including the first BUT and average BUT) increased than baseline level accross 60 min in the WMCG group (P ≤ 0.012), while those in the control group only showed temporary improvements in 5 min. LLT also increased obviously after WMCGs wear, while the change in the control group was nearly innoticeable. No adverse responses were detected. CONCLUSIONS: Temporary use of the WMCGs is able to relieve ocular discomfort, and improves tear film stability in DE patients for at least 1 h, making it a promising alternative to other treatments.

Ultrafast Interfacial Electron and Hole Transfer from CsPbBr3 Perovskite Quantum Dots.

Recently reported colloidal lead halide perovskite quantum dots (QDs) with tunable photoluminescence (PL) wavelengths covering the whole visible spectrum and exceptionally high PL quantum yields (QYs, 50-90%) constitute a new family of functional materials with potential applications in light-harvesting and -emitting devices. By transient absorption spectroscopy, we show that the high PL QYs (∼79%) can be attributed to negligible electron or hole trapping pathways in CsPbBr3 QDs: ∼94% of lowest excitonic states decayed with a single-exponential time constant of 4.5 ± 0.2 ns. Furthermore, excitons in CsPbBr3 QDs can be efficiently dissociated in the presence of electron or hole acceptors. The half-lives of electron transfer (ET) to benzoquinone and subsequent charge recombination are 65 ± 5 ps and 2.6 ± 0.4 ns, respectively. The half-lives for hole transfer (HT) to phenothiazine and the subsequent charge recombination are 49 ± 6 ps and 1.0 ± 0.2 ns, respectively. The lack of electron and hole traps and fast interfacial ET and HT rates are key properties that may enable the development of efficient lead halide perovskite QDs-based light-harvesting and -emitting devices.

Reactive oxygen species activated NLRP3 inflammasomes initiate inflammation in hyperosmolarity stressed human corneal epithelial cells and environment-induced dry eye patients.

In studies on dry eye (DE) disease, an association has been identified between tear film hyperosmolarity and inflammation severity elicited through receptor-induced increases in proinflammatory cytokine and chemokine release. These immune reactions might be mediated by inflammasomes, macromolecular complexes mounted around the NLRP3 protein and can be activated by reactive oxygen species (ROS) over-generation. Hence in this study we determine whether: a) ROS activated NLRP3 inflammasomes mediate hyperosmotic stress-induced inflammation in human corneal epithelial cells (HCECs); b) the ROS-NLRP3-IL-1ß axis activation is associated with environment-induced DE. Immortalized HCECs were exposed to 500 mOsm medium in the presence and absence of a ROS inhibitor, N-acetyl-l-cysteine (NAC). HCECs transfected with NLRP3 siRNA or a negative control (NC) siRNA. Intracellular ROS was measured by fluorometric analysis using the probe 2',7'-dichlorofluorescin diacetate (DCFH-DA). Real-time PCR evaluated NLRP3, ASC, pro-caspase-1 and pro-IL-1ß mRNA levels. Western blot analysis assessed NLRP3 protein expression whereas caspase-1 activity was determined with a fluorometric assay. Bioactive IL-1ß release was assessed by ELISA. ROS production, NLRP3 inflammasome and pro-IL-1ß gene expression as well as IL-1ß secretion were also evaluated in the conjunctival epithelial cells and tear fluid samples of environment-induced DE patients and normal subjects. NAC suppressed hyperosmolarity-induced rises in ROS levels, NLRP3 inflammasome formation and activation, caspase-1 activity and IL-1ß release. On the other hand, NLRP3 siRNA knockdown inhibited hyperosmotic stress-induced NLRP3 activation, which led to ASC, pro-caspase-1 and pro-IL-1ß mRNA down-regulation followed by suppression of associated caspase-1 activity and IL-1ß secretion. In addition, in ocular surface samples of environment-induced DE patients, ROS generation, NLRP3, ASC, pro-caspase-1 and pro-IL-1ß gene expression as well as IL-1ß secretion were upregulated. Taken together, NLRP3 mediated innate immune responses triggered by rises in ROS generation induce inflammation in hyperosmotic stressed HCECs. ROS-NLRP3-IL-1ß signaling pathway might play a priming role in environment-induced DE development.

Viscoelastic-Assisted Non-Descemet Stripping Automated Endothelial Keratoplasty in Vitrectomized and Iris-Lens Diaphragm Injured Eyes.

OBJECTIVES: To report the outcomes of viscoelastic-aided non-Descemet stripping automated endothelial keratoplasty (nDSAEK) to treat bullous keratopathy in vitrectomized and iris-lens diaphragm injured eyes. METHODS: A prospective consecutive case series of eyes with bullous keratopathy underwent nDSAEK. About 0.15 to 0.2 mL cohesive viscoelastic was injected into the anterior chamber in front of the iris defect to prevent air from entering the posterior chamber and vitreous cavity. Filtered air was injected until the intraocular pressure (IOP) rose to a mildly high level. Graft position and complications were monitored postoperatively. Endothelial cell density and best-corrected visual acuity (BCVA) were recorded at a 1-year follow-up. RESULTS: Viscoelastic-aided nDSAEK was performed in 21 cases (11 males, 10 females; mean age: 58.9±12.4 years). No intraoperative complications were noted. Effective graft adherence was achieved in all cases intraoperatively. Postoperatively, partial graft dislocation was seen in 2 cases because of hypotony (IOP, 6 and 5 mm Hg, respectively) due to postoperative cyclodialysis as demonstrated on anterior segment optical coherence tomography. Hypotony was successfully managed conservatively with spontaneous graft reattachment. One year postoperatively, the median BCVA improved to 20/50. The mean endothelial cell loss was 14.7±12.6% and 20.5±11.1% at 6 months and 1 year, respectively. CONCLUSIONS: In vitrectomized and iris-lens diaphragm injured eyes, viscoelastic-aided nDSAEK effectively facilitated graft adherence and reduced graft dislocation. Appropriate control of IOP was considered to play a vital role in supporting the graft adherence.

Reactive oxygen species activated NLRP3 inflammasomes prime environment-induced murine dry eye.

Tear film hyperosmolarity along with exposure to oxidant stress are factors that can induce chronic ocular surface inflammation and pain. However, there is limited information on how increases in reactive oxygen species (ROS) generated by oxidant exposure can induce inflammation. There is emerging evidence in other tissues that innate immune responses to a variety of environmental stresses stem from ROS-induced cytosolic NLRP3 inflammasome activation. Once this occurs, pro-caspase-1 is converted into its catalytic active form, which in turn cleaves pro-IL-1ß thereby generating its bioactive form. We determined the role of ROS generation in mediating increases in IL-1ß secretion through caspase-1 activation caused by NLRP3 inflammasome activation in an environment-induced murine dry eye (DE) model. An intelligently controlled environmental system (ICES) induced evaporative DE in female 4-6 week old C57BL/6J mice. Increases in ROS production preceded rises in corneal and conjunctival gene expression of NLRP3 inflammasome components and IL-1ß that were identified using real-time PCR. Confocal microscopy evaluated concomitant increases in NLPR3, caspase-1 and IL-1ß immunostaining. Increases in caspase-1 activity were used as an indicator of inflammasome activation. Rises in ROS generation occurred after 1 week of ICES exposure, which preceded increases in gene expression of three NLRP3 inflammasome components (i.e. NLRP3, ASC and caspase-1) leading to rises in bioactive IL-1ß release. Increases in caspase-1 activity occurred after 2 weeks of ICES exposure. Eyedrops containing 0.3% N-acetyl-l-cysteine (NAC) were applied to quench ROS generation by mice kept in the ICES for 2 weeks. This scavenger reduced corneal fluorescein staining and decreased ROS production. NAC also down-regulated both increases in NLRP3, ASC, caspase-1 and IL-1ß mRNA levels, along with their immunostaining. It robustly attenuated rises in inflammasome mediated increases in caspase-1 catalytic activity. We show in a dessicating DE disease murine model that rises in ROS generation trigger NLRP3 inflammasome complexation and activation leading to increases in bioactive IL-1ß secretion. These results prompt us to suggest that the ROS-NLRP3-IL-1ß signaling pathway might play a priming role in environment-induced DE progression. Finally, our findings provide a basis for developing novel strategies that may improve the management of patients requiring treatment for environment-induced dry eye disease.

Antiproliferative and apoptotic effects of indomethacin on human retinoblastoma cell line Y79 and the involvement of ß-catenin, nuclear factor-κB and Akt signaling pathways.

BACKGROUND: To determine in vitro if indomethacin inhibits proliferation and induces apoptosis in human retinoblastoma cell line Y79, and to explore possibly involved signaling pathways. METHODS: The human retinoblastoma cell line Y79 was cultured with indomethacin at various concentrations (0, 25, 50, 100, 200 and 400 µmol/l). The effect of indomethacin on cell proliferation and apoptosis was examined by the Cell Counting Kit-8 and TUNEL test, respectively. The mRNA level of survivin, ß-catenin and Bcl-2 was detected by RT-PCR. The protein level of survivin was measured by ELISA. Western blot was used to analyze ß-catenin, nuclear factor (NF)-κB/p65, phosphorylated Akt (pAkt) and total Akt (tAkt) expression in cultured cells. RESULTS: Indomethacin treatment inhibits proliferation (at concentrations from 25 to 400 µmol/l) and induces apoptosis (at concentrations from 100 to 400 µmol/l) of human retinoblastoma cell line Y79 in a dose-dependent manner. RT-PCR showed that the mRNA expression of Bcl-2 (F = 20.497; p < 0.001) and of ß-catenin (F = 14.835; p < 0.001) was significantly different among the treated groups. Survivin mRNA levels remained steady, but its protein levels decreased significantly as measured by ELISA (F = 67.633; p < 0.001). Western blot analysis showed a dose-dependent downregulation of ß-catenin (F = 37.411; p < 0.001), NF-κB/p65 (F = 16.302; p < 0.001) and of pAkt (F = 27.700; p < 0.001) after indomethacin treatment, while tAkt protein expression was steady among the groups. CONCLUSIONS: Treatment with indomethacin can potently suppress proliferation and induce apoptosis in the retinoblastoma Y79 cell line. Wnt/ß-catenin, NF-κB and Akt/PKB pathways might be implicated in the process.

Comparison of deep learning-assisted blinking analysis system and Lipiview interferometer in dry eye patients: a cross-sectional study.

BACKGROUND: Abnormal blinking pattern is associated with ocular surface diseases. However, blink is difficult to analyze due to the rapid movement of eyelids. Deep learning machine (DLM) has been proposed as an optional tool for blinking analysis, but its clinical practicability still needs to be proven. Therefore, the study aims to compare the DLM-assisted Keratograph 5M (K5M) as a novel method with the currently available Lipiview in the clinic and assess whether blinking parameters can be applied in the diagnosis of dry eye disease (DED). METHODS: Thirty-five DED participants and 35 normal subjects were recruited in this cross-sectional study. DED questionnaire and ocular surface signs were evaluated. Blinking parameters including number of blinks, number of incomplete blinking (IB), and IB rate were collected from the blinking videos recorded by the K5M and Lipiview. Blinking parameters were individually collected from the DLM analyzed K5M videos and Lipiview generated results. The agreement and consistency of blinking parameters were compared between the two devices. The association of blinking parameters to DED symptoms and signs were evaluated via heatmap. RESULTS: In total, 140 eyes of 70 participants were included in this study. Lipiview presented a higher number of IB and IB rate than those from DLM-assisted K5M (P ≤ 0.006). DLM-assisted K5M captured significant differences in number of blinks, number of IB and IB rate between DED and normal subjects (P ≤ 0.035). In all three parameters, DLM-assisted K5M also showed a better consistency in repeated measurements than Lipiview with higher intraclass correlation coefficients (number of blinks: 0.841 versus 0.665; number of IB: 0.750 versus 0.564; IB rate: 0.633 versus 0.589). More correlations between blinking parameters and DED symptoms and signs were found by DLM-assisted K5M. Moreover, the receiver operating characteristic analysis showed the number of IB from K5M exhibiting the highest area under curve of 0.773. CONCLUSIONS: DLM-assisted K5M is a useful tool to analyze blinking videos and detect abnormal blinking patterns, especially in distinguishing DED patients from normal subjects. Large sample investigations are therefore warranted to assess its clinical utility before implementation.

Elliptical deep anterior lamellar keratoplasty in severe Acanthamoeba keratitis.

Acanthamoeba keratitis (AK) is an intractable infection of the cornea. Penetrating keratoplasty is widely used for the management of severe AK but suffers from complications like graft rejection, endophthalmitis, and glaucoma. Herein, we aimed to describe the surgical technique and the results of elliptical deep anterior lamellar keratoplasty (eDALK) for the management of severe AK. In this retrospective case series, records of consecutive patients with AK poorly responsive to medical treatment who underwent eDALK from January 2012 to May 2020 were reviewed. The largest diameter of infiltration was ≥8 mm and did not involve the endothelium. The recipient bed was made by an elliptical trephine, and big bubble or wet-peeling technique was performed. Postoperative best spectacle-corrected visual acuity, endothelial cell density, corneal topographic data, and complications were evaluated. Thirteen eyes of thirteen patients (eight men and five women, 45.54 ± 11.78 years old) were included in this study. The mean follow-up interval was 21.31 ± 19.59 months (range, 12-82 months). At the last follow-up, the mean best spectacle-corrected visual acuity was 0.35 ± 0.27 logarithm of the minimum angle of resolution. The mean refractive and topographic astigmatism were - 3.21 ± 1.77 and 3.08 ± 1.14 D, respectively. Intraoperative perforation was encountered in one case and double anterior chambers occurred in two cases. One graft developed stromal rejection and one eye developed amoebic recurrence. eDALK can serve as the first-line surgical management of severe AK poorly responsive to medical treatment.

SIRT1 Inhibits High Glucose-Induced TXNIP/NLRP3 Inflammasome Activation and Cataract Formation.

Purpose: To determine whether SIRT1 regulates high glucose (HG)-induced inflammation and cataract formation through modulating TXNIP/NLRP3 inflammasome activation in human lens epithelial cells (HLECs) and rat lenses. Methods: HG stress from 25 to 150 mM was imposed on HLECs, with treatments using small interfering RNAs (siRNAs) targeting NLRP3, TXNIP, and SIRT1, as well as a lentiviral vector (LV) for SIRT1. Rat lenses were cultivated with HG media, with or without the addition of NLRP3 inhibitor MCC950 or SIRT1 agonist SRT1720. High mannitol groups were applied as the osmotic controls. Real-time PCR, Western blots, and immunofluorescent staining evaluated the mRNA and protein levels of SIRT1, TXNIP, NLRP3, ASC, and IL-1ß. Reactive oxygen species (ROS) generation, cell viability, and death were also assessed. Results: HG stress induced a decline in SIRT1 expression and caused TXNIP/NLRP3 inflammasome activation in a concentration-dependent manner in HLECs, which was not observed in the high mannitol-treated groups. Knocking down NLRP3 or TXNIP inhibited NLRP3 inflammasome-induced IL-1ß p17 secretion under HG stress. Transfections of si-SIRT1 and LV-SIRT1 exerted inverse effects on NLRP3 inflammasome activation, suggesting that SIRT1 acts as an upstream regulator of TXNIP/NLRP3 activity. HG stress induced lens opacity and cataract formation in cultivated rat lenses, which was prevented by MCC950 or SRT1720 treatment, with concomitant reductions in ROS production and TXNIP/NLRP3/IL-1ß expression levels. Conclusions: The TXNIP/NLRP3 inflammasome pathway promotes HG-induced inflammation and HLEC pyroptosis, which is negatively regulated by SIRT1. This suggests viable strategies for treating diabetic cataract.

Fluorescence Imaging of Diabetic Cataract-Associated Lipid Droplets in Living Cells and Patient-Derived Tissues.

Diabetic cataract (DC) surgery carries risks such as slow wound healing, macular edema, and progression of retinopathy and is faced with a deficiency of effective drugs. In this context, we proposed a protocol to evaluate the drug's efficacy using lipid droplets (LDs) as the marker. For this purpose, a fluorescent probe PTZ-LD for LDs detection is developed based on the phenothiazine unit. The probe displays polarity-dependent emission variations, i.e., lower polarity leading to stronger intensity. Especially, the probe exhibits photostability superior to that of Nile Red, a commercial LDs staining dye. Using the probe, the formation of LDs in DC-modeled human lens epithelial (HLE) cells is validated, and the interplay of LDs-LDs and LDs-others are investigated. Unexpectedly, lipid transfer between LDs is visualized. Moreover, the therapeutic efficacy of various drugs in DC-modeled HLE cells is assessed. Ultimately, more LDs were found in lens epithelial tissues from DC patients than in cataract tissues for the first time. We anticipate that this work can attract more attention to the important roles of LDs during DC progression.

Calcium-modified granular attapulgite removed phosphorus from synthetic wastewater containing low-strength phosphorus.

Traditional biological processes combined with chemical precipitation methods can effectively reduce phosphate concentration in wastewater. However, discharge standards required additional advanced treatment technologies, and the removal of low phosphorus concentration is complicated and expensive. This study proposes application of a simple and recyclable adsorbent to remove low-concentration phosphorus from water. The removal efficiency of phosphorus from low-strength synthetic wastewater was investigated and the adsorption mechanism was analyzed. When the initial phosphorus concentration was 2.0 mg/L, the phosphorus adsorption capacity of Ca-GAT increased to 0.891 mg/g from 0.074 mg/g for GAT at 298 K and pH of 7. Phosphorus adsorption on Ca-GAT performs well when the solution pH is in the range of 5-10, and it is not conducive to the adsorption reaction when the solution pH exceeds 11. The competing anions (such as NO3-, SO42-, HCO3- and F-) existed, Ca-GAT still performed better in removing phosphorus. Then, the saturated absorbents could be effectively regenerated with a 0.5 mos/L NaOH solution, while desorption efficiency was reduced from 97.11% to 33.06% after fifth regeneration cycle. Finally, Scanning electron microscopy-energy dispersive spectroscopy (SEM-EDS), Fourier-transform infrared spectroscopy (FTIR), and X-ray photoelectron spectroscopy (XPS) analysis demonstrated that the Ca2+ content on the Ca-GAT surface played an important role in capturing phosphate ions from wastewater. Phosphorus was mainly removed via the formation of Ca-phosphorus precipitation. To some extent, ligand exchanges of CO32- and OH- with HPO42- and H2PO4- were also beneficial for phosphorus removal. The present work shows that attapulgite has sustainable and beneficial potential in the removal of low-strength phosphorous in wastewater, and the phosphorus loaded adsorbent can be used in the agriculture as slow-release fertilizer.

Observation of HOCl generation associated with diabetic cataract using a highly sensitive fluorescent probe.

Diagnosis of diabetic cataract (DC) in the early stage is of great significance for drug intervention and surgery circumvention for DC patients. However, the lack of reliable imaging tools greatly limits the diagnosis of early DC. In this context, a fluorescent probe BBPy for hypochlorous acid (HOCl) is presented based on the oxidation of phenothiazine. The probe displays apparent emission enhancement at 562 nm toward HOCl with high selectivity, superb sensitivity (detection limit: 12.6 nM), and rapid response (within seconds). Using the probe, the HOCl generation in diabetic human lens epithelial cells was monitored, as well as the HOCl down-regulation during antioxidant treatment. Therefore, it is proposed that HOCl can be a promising biomarker for DC and fluorescence imaging technique can be regarded as a candidate tool for DC diagnosis.

iUMRG: multi-layered network-guided propagation modeling for the inference of susceptibility genes and potential drugs against uveal melanoma.

Uveal melanoma (UM) is the most common primary malignant intraocular tumor. The use of precision medicine for UM to enable personalized diagnosis, prognosis, and treatment require the development of computer-aided strategies and predictive tools that can identify novel high-confidence susceptibility genes (HSGs) and potential therapeutic drugs. In the present study, a computational framework via propagation modeling on integrated multi-layered molecular networks (abbreviated as iUMRG) was proposed for the systematic inference of HSGs in UM. Under the leave-one-out cross-validation experiments, the iUMRG achieved superior predictive performance and yielded a higher area under the receiver operating characteristic curve value (0.8825) for experimentally verified SGs. In addition, using the experimentally verified SGs as seeds, genome-wide screening was performed to detect candidate HSGs using the iUMRG. Multi-perspective validation analysis indicated that most of the top 50 candidate HSGs were indeed markedly associated with UM carcinogenesis, progression, and outcome. Finally, drug repositioning experiments performed on the HSGs revealed 17 potential targets and 10 potential drugs, of which six have been approved for UM treatment. In conclusion, the proposed iUMRG is an effective supplementary tool in UM precision medicine, which may assist the development of new medical therapies and discover new SGs.

Impact of Incomplete Blinking Analyzed Using a Deep Learning Model With the Keratograph 5M in Dry Eye Disease.

Purpose: To establish a deep learning model (DLM) for blink analysis, and investigate whether blink video frame sampling rate influences the accuracy of analysis. Methods: This case-controlled study recruited 50 dry eye disease (DED) participants and 50 normal subjects. Blink videos recorded by a Keratograph 5M, symptom questionnaires, and ocular surface assessments were collected. After processing the blink images as datasets, further training and evaluation of DLM was performed. Blink videos of 30 frames per second (FPS) under white light, eight FPS extracted from white light videos, and eight FPS under infrared light were processed by DLM to generate blink profiles, allowing comparison of blink parameters, and their association with DED symptoms and signs. Results: The blink parameters based on 30 FPS video presented higher sensitivity and accuracy than those based on eight FPS. The average relative interpalpebral height (IPH), the frequency and proportion of incomplete blinking (IB) were much higher in DED participants than in normal controls (P < 0.001). The IB frequency was closely associated with DED symptoms and signs (|R| ≥ 0.195, P ≤ 0.048), as was IB proportion and the average IPH (R ≥ 0.202, P ≤ 0.042). Conclusions: DLM is a powerful tool for analyzing blink videos with high accuracy and sensitivity, and a frame rate ≥ 30 FPS is recommended. The IB frequency is indicative of DED. Translational Relevance: The system of DLM-based blink analysis is of great potential for the assessment of IB and diagnosis of DED.