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Background: The CorneaL GrAft Thickness Evaluation (COLGATE) system was recently developed to facilitate the evaluation of corneal graft thickness from OCT images. Graft thickness measurement can be a surrogate indicator for detecting graft failure or success. The purpose of this study was to determine the reproducibility of the COLGATE system in measuring DSAEK graft area between two observers. Methods: This was a prospective case series in which 50 anterior segment OCT images of patients who had undergone DSAEK in either eye were analysed. Two observers (MW, AC) independently obtained the image analysis for the graft area using both semi automated and automated method. One week later, each observer repeated the analysis for the same set of images. Bland-Altman analysis was performed to analyze inter and intra observer agreement. Results: There was strong intraobserver correlation between the 2 semi automated readings obtained by both observers. (r= 0.936 and r = 0.962). Intraobserver ICC for observer 1 was 0.936 (95% CI 0.890 to 0.963) and 0.967 (95% CI 0.942 to 0.981) for observer 2. Likewise, there was also strong interobserver correlation (r= 0.913 and r = 0.969). The interobserver ICC for the first measurements was 0.911 (95% CI 0.849 to 0.949) and 0.968 (95% CI 0.945 to 0.982) for the second. There was statistical difference between the automatic and the semi automated readings for both observers (p= 0.006, p = 0.003). The automatic readings gave consistently higher values than the semi automated readings especially in thin grafts. Conclusion: The analysis from the COLGATE programme can be reproducible between different observers. Care must be taken when interpreting the automated analysis as they tend to over estimate measurements. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effectiveness of slitlamp optical coherence tomography (SL-OCT) and Scanning Peripheral Anterior Chamber depth analyzer (SPAC) in detecting angle closure, using gonioscopy as the reference standard. Methods: A total of 153 subjects underwent gonioscopy, SL-OCT, and SPAC. The anterior chamber angle (ACA) was classified as closed on gonioscopy if the posterior trabecular meshwork could not be seen; with SLOCT, closure was determined by contact between the iris and angle wall anterior to the scleral spur; and with SPAC by a numerical grade of 5 or fewer and/or a categorical grade of suspect or potential. Results: A closed ACA was identified in 51 eyes with gonioscopy, 86 eyes with SL-OCT, and 61 eyes with SPAC (gonioscopy vs SL-OCT, P<.001; gonioscopy vs SPAC, P=.10; SL-OCT vs SPAC, P<.001; McNemar test). Of the 51 eyes with a closed ACA on gonioscopy, SL-OCT detected a closed ACA in 43, whereas SPAC identified 41 (P=.79). An open angle in all 4 quadrants was observed in 102 eyes with gonioscopy, but SL-OCT and SPAC identified 43 and 20 of these eyes, respectively, as having angle closure. The overall sensitivity and specificity for SL-OCT were 84% and 58% vs 80% and 80% for SPAC. Conclusion: Using gonioscopy as the reference, SLOCT and SPAC showed good sensitivity for detecting eyes at risk of angle closure. [ABSTRACT FROM AUTHOR]

Background: Although the lymphatic system arises as an extension of venous vessels in the embryo, little is known about the role of circulating progenitors in the maintenance or development of lymphatic endothelium. Here, we investigated whether hematopoietic stem cells (HSCs) have the potential to give rise to lymphatic endothelial cells (LEC). Methodology/Principal Findings: Following the transfer of marked HSCs into irradiated recipients, donor-derived LEC that co-express the lymphatic endothelial markers Lyve-1 and VEGFR-3 were identified in several tissues. HSC-derived LEC persisted for more than 12 months and contributed to ∼3-4% of lymphatic vessels. Donor-derived LECs were not detected in mice transplanted with common myeloid progenitors and granulocyte/macrophage progenitors, suggesting that myeloid lineage commitment is not a requisite step in HSC contribution to lymphatic endothelium. Analysis of parabiotic mice revealed direct evidence for the existence of functional, circulating lymphatic progenitors in the absence of acute injury. Furthermore, the transplantation of HSCs into ApcMin/+ mice resulted in the incorporation of donor-derived LEC into the lymphatic vessels of spontaneously arising intestinal tumors. Conclusions/Significance: Our results indicate that HSCs can contribute to normal and tumor associated lymphatic endothelium. These findings suggest that the modification of HSCs may be a novel approach for targeting tumor metastasis and attenuating diseases of the lymphatic system. [ABSTRACT FROM AUTHOR]

Presents an approach for conducting pathogenesis of mosaic diseases based on patterns of Cre recombinase protein expression in intestinal epithelium and laser capture microdissection. Utilization of transcriptional regulatory elements that direct region-specific and mosaic patterns; Method for retrieval of epithelial cells from crypts with or without recombination.

Discusses the important roles of the cell adhesion and cell signaling through the association with the cadherins on mouse intestinal epithelial homeostasis. How the protein influences adhesion; Stimulation of cell division; Indication of results.

Purpose: To compare the benefit of femtosecond laser-assisted cataract surgery (FLACS) versus phacoemulsification (PE) and 2 fragmentation patterns in managing dense cataracts., Design: Randomized controlled trial., Methods: Patients with nuclear opacity (NO) grade >5 (Lens Opacities Classification System III) were enrolled at the Singapore National Eye Centre. Patients who were unsuitable for FLACS, whose corneal endothelial cell count (ECC) was <1,500 cells/mm 2 , or had cataracts with additional complexities were excluded from the study. Eyes were randomized to PE, 600 μm grid (FLACSg), or 16-segment fragmentation (FLACS16) in 2:1:1 ratio. The Victus (Bausch & Lomb) laser platform and in situ phacoemulsification chop technique was used. Data for patient demographics, preoperative, and 1 month postoperative best-corrected visual acuity (BCVA), ECC, effective phacoemulsification time (EPT), and perioperative complications were collected. Outcome measurements were the loss of ECC at 1 month and EPT., Results: Ninety-three patients were randomized to PE (48), FLACSg (22), and FLACS16 (23). Majority were Chinese (87; 93.5%). Mean age was 74.3 ± 8.8 years of age. Cataracts were mostly graded as NO 5-6 (49; 61.3%). EPT among treatment arms was not different (P = .097, one-way ANOVA) but was significantly higher for NO >6 than NO <6 (P < .001, general linear model). ECC loss was significantly less in FLACSg than in PE (P = .018, Bonferroni correction). Mean 1-month postoperative LogMAR BCVA (0.23 ± 0.20) was significantly better than preoperative BCVA (1.02 ± 0.85; P < .001, paired t test) but not different between PE and FLACS., Conclusions: FLACSg but not FLACS16 significantly lowered the mean ECC loss during phacoemulsification in dense cataracts., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Purpose: To assess the role of femtosecond laser technology in the management of severely subluxated cataracts., Design: Retrospective, interventional case series., Methods: All eyes with subluxated cataract seen between July 2012 and June 2015 were assessed for suitability for femtosecond laser-assisted cataract surgery, with the use of capsular tension devices. Participants with subluxated cataracts of at least 6 clock hours of zonular weakness were included in the study. Data collected included patient demographics, preoperative and postoperative best-corrected visual acuity (BCVA), nuclear density, extent of zonular weakness, completeness of capsulotomy, and complications. Poor visual outcome was defined as BCVA of worse than 20/40. Main outcome measure was the retention of the capsular bag., Results: Of the 72 eyes with subluxated cataracts undergoing surgery during the study period, 47 eyes of 47 patients were eligible for analysis. Mean age of the patients was 60.7 years (standard deviation [SD] 13.2 years). The majority were male (32, 68.1%) and Chinese (38, 80.8%). The mean duration of follow-up was 8 months (SD 5.6 months). The main identifiable cause of lens subluxation was trauma (11 eyes). Almost two thirds (30 eyes) had more than 9 clock hours of zonular weakness. Seventy percent of cataracts (33) were nuclear sclerosis grade 3 and above. The capsular bag was preserved in 43 eyes (91.5%). The intraocular lens was stable and centered at the last follow-up in all these 43 eyes. An anterior capsule tear occurred in 6 eyes, all of which had cataracts of nuclear sclerosis grade 3 and above, with posterior extension occurring in 3 eyes. Primary posterior capsule rupture occurred in 1 eye. At 1 month 37 eyes (80.4%) had a BCVA of 20/40 or better. There was significant improvement in BCVA at 1 month (mean of 0.92 logMAR units [SD 0.88] to 0.22 [SD 0.38] [P < .001, paired samples t test]), which was maintained at 1 year., Conclusions: Selected cases of severely subluxated cataracts may be managed using femtosecond laser technology to perform the capsulotomy and nuclear fragmentation, with successful preservation of the capsular bag in 90% of eligible cases, especially in eyes with soft cataracts., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Objective: We report a case series of brown discoloration of Abbott Medical Optics, USA, acrylic intraocular lens (IOLs; AABOO, ZCBOO, and ZMBOO) after cataract operation., Design: Retrospective case series., Participants: Patients with brown discoloration of their IOLs postoperatively during the period from March 2013 to October 2013 at the Singapore National Eye Centre were consecutively identified., Methods: Distance best-corrected visual acuity (BCVA), intraocular pressure, slit-lamp examination, colour fundus photographs, flare meter reading, endothelial cell count, dilated fundus examination, as well as colour discrimination test on the Lanthony D 15 Hue test were performed., Results: Sixteen eyes of 14 patients were found to have evenly brown discoloration of varying degrees of acrylic IOL postphacoemulsification. This included 11 AABOO IOLs, 3 ZCBOO IOLs, and 2 ZMBOO IOLs. All had uneventful surgery except one who required anterior vitrectomy for posterior capsule rupture. One patient had uneventful operation but subsequently developed pigment dispersion glaucoma with tunnel vision and required a trabeculectomy. Mean time from operation to reporting date was 143 days (range 1-327 days). Pre- and postoperative month 6 BCVA was logMAR 0.54 and logMAR 0.13, respectively. None of the patients had loss of lines of BCVA. The desaturated Lanthony D 15 Hue test was abnormal in 8 out of the 16 eyes. None required IOL explantation., Conclusions: Varying degrees of brown discoloration may occur in today's modern hydrophobic acrylic IOLs., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Correlated imaging is the process of imaging a specimen with two complementary modalities and then registering and overlaying the fields obtained in each modality to create a composite view. One of the images is made somewhat transparent, allowing detail in the underlying image to be visible and assisting in the registration of the two images. As an example, an image localizing a specific tissue component by fluorescence may be overlaid atop a TEM image of the same field. The resulting composite image would demonstrate specific ultrastructural features in the high-resolution TEM field, which are colorized in the overlay. Other examples include composites from MicroCT or soft X-ray images overlaid atop light microscopy or TEM images. Automated image registration may be facilitated by a variety of sophisticated computer programs utilized by high-throughput laboratories. This chapter is meant for the more occasional user wishing to align images manually. ImageJ is a public domain, image processing program developed at the National Institutes of Health and is available to anyone as a free download. ImageJ performs marvelously well for the purpose of image registration; therefore, step-by-step instructions are included here. Specimen handling, including fixation and choice of embedding media, is not straightforward for correlative imaging. A step-by-step description of the protocols which work in our laboratory is included for simultaneous localization in LM, EM and micro-CT, as well as maintaining GFP emission in tissue embedded for TEM., (© 2014 Elsevier Inc. All rights reserved.)

Fluorescence-activated cell sorting (FACS) is an essential tool for studies requiring isolation of distinct intestinal epithelial cell populations. Inconsistent or lack of reporting of the critical parameters associated with FACS methodologies has complicated interpretation, comparison, and reproduction of important findings. To address this problem a comprehensive multicenter study was designed to develop guidelines that limit experimental and data reporting variability and provide a foundation for accurate comparison of data between studies. Common methodologies and data reporting protocols for tissue dissociation, cell yield, cell viability, FACS, and postsort purity were established. Seven centers tested the standardized methods by FACS-isolating a specific crypt-based epithelial population (EpCAM+/CD44+) from murine small intestine. Genetic biomarkers for stem/progenitor (Lgr5 and Atoh 1) and differentiated cell lineages (lysozyme, mucin2, chromogranin A, and sucrase isomaltase) were interrogated in target and control populations to assess intra- and intercenter variability. Wilcoxon's rank sum test on gene expression levels showed limited intracenter variability between biological replicates. Principal component analysis demonstrated significant intercenter reproducibility among four centers. Analysis of data collected by standardized cell isolation methods and data reporting requirements readily identified methodological problems, indicating that standard reporting parameters facilitate post hoc error identification. These results indicate that the complexity of FACS isolation of target intestinal epithelial populations can be highly reproducible between biological replicates and different institutions by adherence to common cell isolation methods and FACS gating strategies. This study can be considered a foundation for continued method development and a starting point for investigators that are developing cell isolation expertise to study physiology and pathophysiology of the intestinal epithelium.

Objective: To report the 8-year outcomes of Asian subjects who underwent trabeculectomy augmented by intraoperative 5-fluorouracil (5-FU) or placebo., Design: Retrospective review of a randomized controlled trial., Participants: Subjects with primary open-angle or angle-closure glaucoma., Methods: Study subjects were prospectively followed up for 3 years. After the last subject recruited had completed 8 years of follow-up, the charts of all subjects were reviewed to capture data from the year 3 visit onward. Kaplan-Meier survival function with Cox regression was performed to identify risk factors associated with trabeculectomy failure at 8 years., Main Outcome Measures: The primary outcome was trabeculectomy failure defined as intraocular pressure (IOP) >21 or <6 mmHg on 2 consecutive occasions after the first 6 weeks after surgery, repeat glaucoma surgery, or loss of light perception. Further end points were defined at IOP levels >17 and >14 mmHg. Secondary outcomes included IOP at 8 years and number of glaucoma medications. Complete success was defined using IOP end points ≤ 21, ≤ 17, or ≤ 14 mmHg at 8 years without medications., Results: Of the 243 enrolled subjects, 170 (70.0%) completed 8 years follow-up, 86 in the 5-FU and 84 in the placebo group. There was no significant difference in failure rates at 8 years for the failure definitions of IOP >21 mmHg (11.6% of the 5-FU group vs. 16.7% of the placebo group; P = 1.00), IOP >17 mmHg (23.3% of the 5-FU group vs. 31% of the placebo group; P = 0.78), and IOP >14 mmHg (46.5% of the 5-FU group vs. 58.3% of the placebo group; P = 0.37). Mean IOP at 8 years was 13.7 mmHg in the 5-FU versus 14.4 mmHg in the placebo group (P = 0.24). Mean number of medications was 0.65 drops in the 5-FU versus 0.93 drops in the placebo group (P = 0.06). Complete success with IOP ≤ 21 mmHg at 8 years was achieved in 48 subjects (55.8%) in the 5-FU and 33 subjects (39.3%) in the placebo group (P = 0.09). Absence of bleb microcysts at 1 year, preoperative IOP, and number of bleb needlings performed within the first year were significantly associated with failure., Conclusions: There was no significant difference in IOP between the 5-FU and the placebo group at 8 years., Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article., (Copyright © 2013 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Many advances have been reported in the long-term culture of intestinal mucosal cells in recent years. A significant number of publications have described new culture media, cell formations, and growth patterns. Furthermore, it is now possible to study, e.g., the capabilities of isolated stem cells or the interactions between stem cells and mesenchyme. However, at the moment there is significant variation in the way these structures are described and named. A standardized nomenclature would benefit the ability to communicate and compare findings from different laboratories using the different culture systems. To address this issue, members of the NIH Intestinal Stem Cell Consortium herein propose a systematic nomenclature for in vitro cultures of the small and large intestine. We begin by describing the structures that are generated by preparative steps. We then define and describe structures produced in vitro, specifically: enterosphere, enteroid, reconstituted intestinal organoid, induced intestinal organoid, colonosphere, colonoid, and colonic organoid.