Continuous Corneal Endothelial Damage by Chlorhexidine Alcohol Used for Disinfection in Nonophthalmic Surgery.

PURPOSE: To report a case of corneal endothelial damage caused by alcohol-containing chlorhexidine gluconate (CG-A) and its progression over time. METHODS: This was a case report. RESULTS: A 22-year-old man underwent neurosurgery under general anesthesia. CG-A (1%) was used for disinfection after the application of corneal protection tape. Postoperatively, the patient presented with hyperemia and swelling of the left conjunctiva and was referred to our department. Initial examination revealed left corneal epithelial erosion and corneal edema, which improved on postoperative day 14. The corneal endothelial cell density (ECD) was 3,345 cells/mm 2 on day 14, decreased rapidly to 2,090 cells/mm 2 on day 42, and slowly reduced to 1,122 cells/mm 2 on day 168. Thereafter, no decrease in ECD was observed. CONCLUSIONS: CG formulations can lead to a persistent decrease in ECD over several months, even after improvement of acute corneal edema.

Association between aqueous humor cytokines and postoperative corneal endothelial cell loss after Descemet stripping automated endothelial keratoplasty.

This study measured the intraoperative anterior aqueous humor concentrations of various cytokines during corneal endothelial transplantation and searched for relationships between these concentrations and postoperative corneal endothelial cell (CEC) depletion. We recruited 30 consecutive patients who underwent corneal endothelial transplantation with Descemet's stripping automated endothelial keratoplasty (DSAEK) at Tohoku University Hospital between February 2014 and July 2017. During surgery, we obtained aqueous humor samples and later measured the concentrations of 27 cytokines with a Multiplex Bead Assay (Bio-Plex Pro). We counted CECs 1, 6 and 12 months after surgery, and used Spearman's rank correlation coefficient to identify relationships between CEC depletion and the concentrations of detected cytokines. The loss of CECs 1-6 months after surgery was significantly correlated with IL-7, IP-10, MIP-1a and MIP-1b concentrations (-0.67, -0.48, -0.39, and -0.45, respectively, all P <0.01). CEC loss 1-12 months after surgery was significantly correlated with IL-1b, IL-7, IP-10 and RANTES concentrations (-0.46, -0.52, -0.48, and -0.43, respectively). Multiple regression analysis showed that IL-7 concentration was significantly associated with CEC loss 1-6 months after surgery (b = -0.65, P < 0.01) and IP-10 concentration was associated with CEC loss 1-12 months after surgery (ß = -0.38, P < 0.05). These results suggest that not only inflammatory cytokines but also IL-7, a cytokine related to lymphocytes, may be involved in the depletion of CECs after DSAEK, particularly depletion that occurs relatively early.

Ascorbic acid ameliorates corneal endothelial dysfunction and enhances cell proliferation via the noncanonical GLUT1-ERK axis.

BACKGROUND: The pumping function of corneal endothelial cells (CECs) plays a pivotal role in the maintenance of corneal water homeostasis. Corneal endothelial dysfunction (CED) leads to corneal edema and opacity, but with the exception of keratoplasty, no optimal therapeutic strategies have been established for CED. In this study, we aimed to investigate the ameliorative effect of ascorbic acid (AA) on CED and the underlying mechanism of action in the corneal endothelium. METHODS: Rabbit corneal endothelial damage was induced by anterior chamber injection of benzalkonium chloride (BAK). AA was topically administered to the corneal surface, and the transparency and thickness of the cornea were assessed by external eye photography, slit-lamp photography, and ultrasonic pachymetry. To further analyze the mechanism, rabbit CECs and immortalized human CECs (B4G12 cells) were cultured. A ferric reducing/antioxidant and AA (FRASC) assay was performed to measure the AA concentration. Cell proliferation was evaluated by cell counting and bromodeoxyuridine (BrdU) labeling assays, and protein expression was examined by liquid chromatography-mass spectrometry (LC/MS) and immunoblotting. The involvement of glucose transporter 1 (GLUT1) and phospho-ERK was evaluated via GLUT1-siRNA and phospho-ERK inhibitor (PD98059) treatment. INTERPRETATION: We observed that topical AA ameliorates BAK-induced rabbit corneal endothelial damage. Furthermore, we demonstrated that AA is transported into B4G12 cells via GLUT1, and afterward, AA increases ERK phosphorylation and promotes cell proliferation. Our findings indicate that CEC proliferation stimulated via the noncanonical AA-GLUT1-ERK axis contributes to AA-enhanced healing of CED.

Myopia, corneal endothelial cell density and morphology in a Japanese population-based cross-sectional study: the JPHC-NEXT Eye Study.

This population-based cross-sectional study was performed to determine the mean corneal endothelial cell density (ECD), coefficient of variation (CV), and hexagonality (HEX), and their associations with myopia in Japanese adults living in Chikusei city. Of 7109 participants with available data, 5713 (2331 male and 3382 female) participants were eligible for analysis. After assessing the relationship between participant characteristics and spherical equivalent refraction (SER), the association of SER with the abnormal value of ECD (< 2000 cells/mm), CV (≥ 0.40), and HEX (≤ 50%) were determined using the logistic regression models adjusting for potential confounders (age, intraocular pressure, keratometric power, height, and antihypertensive drug use). In male participants, there was no statistically significant relationships between SER and endothelial parameters. In female participants, compared to emmetropia, SER ≤ - 6 D had significantly higher odds ratio (OR) of having the abnormal value of CV (OR = 2.07, 95% confidence interval [CI] 1.39-3.10) and HEX (OR = 2.04, 95% CI 1.29-3.23), adjusted for potential confounders, indicating that the high myopia was associated with the abnormal values of CV and HEX. Further adjustment for contact lenses wear partly attenuated these associations. Association between the SER and ECD was not detected.

Cigarette Smoke Triggers Loss of Corneal Endothelial Cells and Disruption of Descemet's Membrane Proteins in Mice.

Purpose: To investigate changes at a molecular level in the mouse corneal endothelium (CE) exposed to chronic cigarette smoke (CS). Methods: Pregnant mice (gestation days 18-20) were placed in a whole-body exposure smoking chamber, and a few days later pups were born. After 3.5 months of CS exposure, a ConfoScan4 scanning microscope was used to examine the corneal endothelial cells (CECs) of CS-exposed and control (Ct) mice. The CE was peeled under a microscope and maintained as four biological replicates (two male and two female) for CS-exposed and Ct mice; each replicate consisted of 16 CEs. The proteome of the CE was investigated through mass spectrometry. Results: The CE images of CS-exposed and Ct mice revealed a difference in the shape of CECs accompanied by a nearly 10% decrease in CEC density (P < 0.00003) following CS exposure. Proteome profiling identified a total of 524 proteins exhibiting statistically significant changes in CE from CS-exposed mice. Importantly, proteins associated with Descemet's membrane (DM), including COL4α1, COL4α2, COL4α3, COL4α4, COL4α5, COL4α6, COL8α1, COL8α2, and FN1, among others, exhibited diminished protein levels in the CE of CS-exposed mice. Conclusions: Our data confirm that exposure to CS results in reduced CEC density accompanied by diminished levels of multiple collagen and extracellular matrix proteins associated with DM.

Effect of Down syndrome and keratoconus on corneal density and volume: a triple comparative study.

Keratoconus (KCN) and Down syndrome affect the corneal density and volume. In this study included Down syndrome patients with and without KCN (24 Down-KCN and 204 Down-nonKCN eyes) and normal age- and gender-matched individuals (184 eyes). Studied parameters were the corneal density measured with Pentacam HR in 5 concentric zones and annuli (0-2 mm, 2-6 mm, 6-10 mm, 10-12 mm, and 0-12 mm) in 4 different depth layers (anterior 120 µm, posterior 60 µm, middle layer, and the full thickness of the cornea), and the 10 mm zone corneal volume. In Down-KCN, Down-nonKCN, and control groups, respectively, mean full thickness density in the 0-12 mm zone was 19.35 ± 2.92, 17.85 ± 2.55, and 15.78 ± 2.67 GSU, and mean corneal volume was 57.45 ± 4.37, 56.99 ± 3.46, and 61.43 ± 3.42mm3. All density readings were significantly different between the three studied groups (all P < 0.01) except full thickness density in 0-2 mm and 2-6 mm (P > 0.05) and corneal volume (P = 0.519) between Down-KCN and Down-nonKCN groups; these inter-group densitometry differences within the 6 mm zone were only in the middle layer, and not the anterior or posterior thickness layers (all P > 0.05). Corneal density increased with age and corneal thickness, but there was no significant relationship with gender. Overall, Down syndrome is associated with increased density and light scatter in all corneal layers up to the 12 mm diameter. In Down patients with KCN, the increased light scatter and density in the 6 mm zone is only in the middle thickness layer. Corneal volume is reduced in Down syndrome irrespective of the presence or absence of KCN.

The Influence of Speed During Stripping in Descemet Membrane Endothelial Keratoplasty Tissue Preparation.

PURPOSE: To evaluate whether the speed of stripping a Descemet membrane endothelial keratoplasty graft influences the graft scroll width. METHODS: Human corneas suitable for research were selected for the study. Pairs of corneas were randomly divided into 2 groups: 1 cornea was stripped with a slow speed (group 1) and the contralateral with a fast speed (group 2). Slow speed was defined as the total time greater than 150 seconds or speed <0.057 mm/s. Fast peeling was defined as less than 75 seconds or speed >0.11 mm/s. The grafts acquired were evaluated by microscopy for the graft scroll width and endothelial cell density change pre- and post-preparation. RESULTS: Twenty corneas of 10 donors were included in the analysis. The mean donor age was 68.6 ± 7.58 years. The mean total time of the tissue preparation in group 1 was 282.7 ± 28 seconds and in group 2 was 126 ± 50 seconds (P-value = 0.00000047). The mean speed of stripping in group 1 was 0.045 ± 0.006 mm/s and in group 2 was 0.266 ± 0.093 mm/s (P-value = 0.000027). The graft width in group 1 was 6.4 ± 0.92 mm and in group 2 was 2.87 ± 0.32 mm (P-value = 0.00000014). The mean endothelial cell loss in group 1 was 389 ± 149 cells/mm and in group 2 was 186 ± 63.44 cells/mm (P-value = 0.00134). CONCLUSION: We found a correlation between the speed of stripping, scroll width, and endothelial cell loss. Slow-peeled Descemet membrane endothelial keratoplasty grafts result in a wider scroll width but were associated with a greater reduction in endothelial cell density.

Comparison of changes in corneal endothelial cell density and central corneal thickness between conventional and femtosecond laser-assisted cataract surgery: a randomised, controlled clinical trial.

BACKGROUND/AIM: To identify changes in endothelial cell density (ECD) and central corneal thickness (CCT) in eyes undergoing femtosecond laser-assisted cataract surgery (FLACS) compared with conventional phacoemulsification surgery (CPS). METHODS: This is an intraindividual randomised, controlled clinical trial. One eye was randomised to receive FLACS, while the contralateral eye of the same patient received CPS. The femtosecond laser pretreatment included creating main and side-port corneal incisions, capsulotomy and lens fragmentation. Non-contact endothelial cell microscopy and pachymetry were performed preoperatively and at postoperative day 1, week 1, month 1 and month 3. RESULTS: A total of 134 paired eyes from 67 patients were included in the analysis. ECD was not significantly different between the two groups at either postoperative month 1 (2370±580 cells/mm2 and 2467±564 cells/mm2 in FLACS and CPS groups, respectively; p=0.18) or at postoperative month 3 (2374±527 cells/mm2 and 2433±526 cells/mm2 in FLACS and CPS groups, respectively; p=0.19). No significant difference was observed in the mean CCT values between the two groups over the follow-up period (p>0.05). CONCLUSION: Postoperative corneal ECD and CCT were comparable between FLACS and CPS during the 3 months' follow-up period.

Effects of ultrasound energy on the porcine corneal endothelium - Establishment of a phacoemulsification damage model.

PURPOSE: The purpose of this study was to establish a standardized in vitro phacoemulsification damage model for future investigations of the effects of phacoemulsification, surgical devices, protective ophthalmic viscoelastic devices (OVDs), irrigation solutions and other aspects related to cataract phacoemulsification surgery on the corneal endothelium using porcine eyes. METHODS: Thirty-four porcine eyes were randomly assigned to three groups (phacoemulsification (n = 13), irrigation (n = 9), control (n = 12)). A total of 5 min of ultrasound energy with intermittent irrigation/aspiration was applied in the eyes of the phacoemulsification group. The eyes of the irrigation group received the identical treatment, but without the application of ultrasound energy. The control group was left untreated. All eyes were then prepared to split corneal buttons followed by 15 days of cultivation. Endothelial cell density (ECD) was assessed blinded on day 15. RESULTS: Endothelial cell density declined significantly more until day 15 in the phacoemulsification group (2567 ± 317/267 cells/mm² (median ± 25%/75%-quartiles), -32.5 ± 7.0/6.4%) compared to the irrigation (3450 ± 350/383 cells/mm², -11.8 ± 5.3/2.6%; p < 0.001) and the control group (3650 ± 288/258 cells/mm², -10.2 ± 3.2/4.6%; p < 0.001). CONCLUSION: The phacoemulsification damage model presented in this study is sensitive to phacoemulsification energy and may reliably be used to investigate various factors involved in phacoemulsification with regard to their influence on corneal endothelial cells. This method is able to replace animal experiments or in vitro cell culture experiments that often do not translate well to the in vivo situation in humans.

Quantitative and Qualitative Differences in Endothelial Cell Loss Between Endothelium-In Versus Endothelium-Out Loading in Descemet Membrane Endothelial Keratoplasty.

PURPOSE: This study compares endothelial cell loss (ECL) between donor grafts loaded in the Geuder Glass Cannula and the Coronet EndoGlide made specifically for Descemet membrane endothelial keratoplasty (DMEK). METHODS: Prestripped tissues using the submerged cornea using backgrounds away (SCUBA) technique were obtained from an eye bank. Donor grafts were loaded with the Geuder Glass Cannula or Coronet DMEK EndoGlide. Grafts were then ejected directly onto a glass slide, stained, and imaged with confocal fluorescent microscopy. Trainable Fiji (ImageJ) segmentation software was used to quantify ECL. RESULTS: Twenty-three total tissue samples were tested. Eleven grafts were loaded using the Geuder, and 12 were loaded with the Coronet. Preloading graft endothelial cell density was comparable between Geuder (2436 ± 581 cells/mm) and Coronet (2577 ± 483 cells/mm) groups with P = 0.56. Other baseline characteristics including donor age and time from death to loading were comparable (P = 0.73 and P = 0.66, respectively). Average ECL was 11.3% ± 3.0% in the Geuder group versus 7.2% ± 7.0% in the Coronet group and not significantly different (P = 0.07). When observing patterns of cell loss, grafts loaded in the Coronet DMEK EndoGlide resulted in more focal forceps grabs and trifold patterns. Grafts loaded with the Geuder Glass Cannula yielded more diffuse stippled stress lines. CONCLUSIONS: Our results suggest comparable ECL between Geuder (endothelium-out) and Coronet (endothelium-in) DMEK EndoGlide tissue injection techniques. Patterns of ECL in the Coronet group correlated with forceps grabs, but this study suggests that additional manipulation of tissue does not result in higher ECL.

Parameters Associated With Endothelial Cell Density Variability After Descemet Membrane Endothelial Keratoplasty.

PURPOSE: To evaluate which parameters may affect endothelial cell loss after Descemet membrane endothelial keratoplasty (DMEK) by comparing eyes in the low vs high quartile of endothelial cell loss over a follow-up period of 4 years. DESIGN: Retrospective cohort study. METHODS: Donor endothelial cell density (ECD) decline was evaluated for 351 eyes of 275 patients up to 4 years after DMEK for Fuchs endothelial corneal dystrophy (FECD). Eyes with a postoperative endothelial cell loss in the lower quartile at all available follow-up moments were assigned to Group 1 (n = 51) and those in the upper quartile to Group 2 (n = 42). Multinomial regression was used to assess which covariates were related to greater ECD decline. RESULTS: Mean endothelial cell loss as compared to preoperative donor ECD for the entire study group was 33 (±16)%, 36 (±17)%, and 52 (±18)% at 1, 6, and 48 months postoperatively. Endothelial cell loss of Group 1 was 12 (±7)%, 13 (±6)%, and 26 (±8)% at, respectively, 1, 6, and 48 months postoperatively, and 59 (±10)%, 64 (±9)%, and 75 (±5)% in Group 2. Partial graft detachment, donor death cause cardiovascular/stroke (vs cancer), postoperative complications other than graft detachment, and severity of preoperative FECD (all P < .01) showed the strongest relation with greater ECD decline. CONCLUSIONS: DMEK eyes with a completely attached graft and operated in an early stage of FECD may show the lowest endothelial cell loss postoperatively.

Comparison of Perioperative Parameters in Femtosecond Laser-Assisted Cataract Surgery Using 3 Nuclear Fragmentation Patterns.

PURPOSE: The purpose of this study was to compare the perioperative parameters of quadrant, sextant, and grid lens fragmentation patterns in femtosecond laser-assisted cataract surgery (FLACS). DESIGN: Prospective randomized clinical trial. METHODS: Setting: Eye Center of the Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China. STUDY POPULATION: A total of 894 eyes in 661 patients with cataracts were enrolled. Intervention or observation procedures: the nuclear density was graded according to the Emery-Little classification. Patients received lens fragmentation using a quadrant, sextant, or grid pattern after random allocation. Evaluations included intraoperative parameters, complications, and postoperative outcomes. MAIN OUTCOME MEASUREMENTS: effective phacoemulsification time (EPT), intraoperative complications, visual acuity and intraocular pressure at one day postoperatively, as well as endothelial cell density, endothelial cell loss, and central corneal thickness at 1 week postoperatively. RESULTS: In grade 1 nuclei, the mean EPT in the grid group was the shortest compared to those in the quadrant (P = 0.011) and sextant (P = 0.001) groups. In grade 2 nuclei, all 3 patterns showed no significant differences in the mean EPT (P > 0.05). In grade 3 nuclei, the sextant group revealed shorter mean EPT than the grid (P = 0.017) and quadrant (P > 0.05) groups. In grades 4 and 5 nuclei, the quadrant pattern had the shortest mean EPT among all 3 patterns (P < 0.05). The grid pattern is associated with higher intraocular pressure in hard nuclei (grades 4 and 5) than the other 2 patterns (P < 0.05). CONCLUSIONS: The grid and quadrant patterns allow for shorter EPT in soft (grade 1) and hard (grades 4 and 5) nuclei, respectively. All 3 patterns can be selected for treating grade 2 nuclei. The sextant pattern may be the best option when treating grade 3 nuclei. The grid pattern should be avoided in hard nuclei combined with glaucoma or glaucoma suspect.

Corneal endothelial cell changes in diabetics versus age group matched nondiabetics after manual small incision cataract surgery.

Purpose: To assess and compare the endothelial cell changes after manual small incision cataract surgery (SICS) in diabetic patients versus age group matched non-diabetic patients. Methods: This comparative prospective observational follow-up study included 54 diabetic patients and 52 control patients without diabetes who underwent manual SICS. Preoperative, one day, one week, one month and three months post-surgery assessments of corneal endothelial cell changes were done using specular microscopy. Data analysis was performed using SPSS software (version 20.0, SPSS, Inc.). Mann-Whitney U test was used to compare the data between the test group and control group. Results: There was drop in the endothelial density in both the groups postoperatively, with the mean percentage of endothelial loss at three months post- surgery being 27.5% in diabetics and 18.3% in controls. There was also a significant increase in central corneal thickness and coefficient of variance in diabetics as compared to controls at every follow up one day, one week, one month and three months. The percentage of hexagonality was statistically significant at post-operative three months. Conclusion: The diabetic endothelium was found to be under greater metabolic stress and had less functional reserve after manual SICS than the normal corneal endothelium.

Biophysical and microstructural changes of swelling cornea caused by endothelial cells damage.

Biophysical properties and microstructural changes of swelling cornea which caused by endothelial cells damage will be evaluated. Swelling cornea models were established by endothelial cells damage in 114 Sprague Dawley rats. Relative gray value, swelling rate and light transmittance were measured to evaluated the biophysical properties and microstructure changes were observed by transmission electron microscopy. Relative gray value decreased while swelling rate rose along with time and both of them reached relative stability after 7 days. Light transmittance showed a decline trend with time even after corneal thickness had reached stable stage. Observed by transmission electron microscopy, interfibrillar distance increased, fewer proteoglycans coating appeared and remnants proteoglycan branches became thinner and longer in 7 days. Diameter of fibrils didn't change obviously with time. In cornea edema models caused by endothelial cells damage, the changes of biophysical property and microstructure can help us evaluate corneal edema accurately and objectively.

Descemet's membrane endothelial keratoplasty for pseudoexfoliation syndrome: a case series.

BACKGROUND: To evaluate the clinical outcomes and features of Descemet's membrane endothelial keratoplasty (DMEK) for eyes with pseudoexfoliation syndrome (PEX). METHODS: In this retrospective study, 37 DMEK cases were reviewed from available medical records. Patients who exhibited endothelial dysfunction derived from PEX or Fuchs endothelial corneal dystrophy (FECD) and successfully underwent cataract surgery about four weeks before DMEK were enrolled. The best spectacle-corrected visual acuity (BSCVA), central corneal thickness (CCT), endothelial cell density (ECD), and incidence of intra-operative/post-operative complications of DMEK were analyzed. RESULTS: This study included 14 eyes of 14 patients (PEX: n = 6, FECD: n = 8). There was no primary graft failure. In the PEX group, BSCVA improved from 0.67 ± 0.28 at the preoperative point to 0.43 ± 0.14 at 1 month, 0.27 ± 0.10 at 3 months, and 0.19 ± 0.08 at 6 months after DMEK. The donor corneal ECD was 2704 ± 225 cells/mm2 at the preoperative point and decreased to 1691 ± 498 cells/mm2 at 1 month, 1425 ± 366 cells/mm2 at 3 months, and 1281 ± 340 cells/mm2 (52.7 ± 11.7% less than ECD of the donor graft) at 6 months after DMEK. None of the patients required rebubbling. When compared with the FECD group, no statistical difference was observed in CCT (p = 0.821); BSCVA (p = 0.001) and the reduction rate of ECD (p = 0.010) were relatively worse. CONCLUSIONS: DMEK is effective for the treatment of endothelial dysfunction due to PEX.

Risk Factors for Cystoid Macular Edema After Descemet Membrane Endothelial Keratoplasty.

PURPOSE: To investigate factors associated with cystoid macular edema (CME) after Descemet membrane endothelial keratoplasty (DMEK) in Asian eyes. METHODS: In this retrospective, interventional, consecutive case series, 77 eyes of 65 patients who underwent DMEK were evaluated; in 53 eyes, cataract surgery was performed 1 month before DMEK (staged DMEK), and 24 eyes underwent DMEK alone (simple DMEK). Central retinal thickness, incidence of CME, postoperative best-corrected visual acuity, central corneal thickness, and corneal endothelial cell density were assessed at 1, 3, and 6 months after surgery. Multiple regression analysis and stepwise variable selection were performed for parameters such as type of surgery, iris damage scores, age, sex, axial length, preoperative visual acuity, rebubbling, air volume in the anterior chamber on postoperative day 1, history of diabetes, and endothelial cell density loss rates at 6 months after surgery. RESULTS: CME occurred in 12 (15.6%) of 77 eyes. There was no significant difference in best-corrected visual acuity between eyes with and without CME (P = 0.27). Multivariable analysis revealed that the difference in iris damage scores between before and after DMEK (P < 0.001), air volume in the anterior chamber (P = 0.012), simple DMEK (P = 0.020), and rebubbling (P = 0.036) were significantly associated with CME. Stepwise variable selection indicated that iris damage (P < 0.001) was the most important risk factor for CME. CONCLUSIONS: Iris damage due to DMEK might be a possible risk and aggravating factor for the development of CME after DMEK. Surgeons should attempt to minimize damage to the iris.

Calcium Oscillatory Behavior and Its Possible Role during Wound Healing in Bovine Corneal Endothelial Cells in Culture.

In epithelial layers in culture, immediately after an injury a fast calcium wave (FCW) propagates from the wound borders toward the rest of the monolayer. We show here that similarly to other tissues, during the FCW in bovine corneal endothelial (BCE) cells in culture many cells exhibit calcium oscillations mediated by IP3 signaling. In this study we perform a detailed characterization of this oscillatory behavior and explore its possible role in the process of wound healing. In previous work we showed that, in BCE cells in culture, the healing cells undergo two stages of caspase-dependent apoptosis, at approximately two and eight hours after wounding. We determined that inhibition of the FCW greatly increases the apoptotic rate of the two stages, suggesting that the wave prevents excessive apoptosis of the healing cells. Taking this into account, we investigated the possible participation of the calcium oscillations during the FCW in apoptosis of the healing cells. For this, we employed ARL-67156 (ARL), a weak competitive inhibitor of ecto-ATPases, and the calcium chelator EGTA. We show here that, in healing BCE cells, ARL enhances cellular calcium oscillations during the FCW, while EGTA decreases oscillations. We found that ARL produces a significant decrease (to about half the control value) in the apoptotic index of the first stage of apoptosis, while EGTA increases it. Neither drug noticeably affects the second stage. We have interpreted the effect of ARL on apoptosis as due to the maintenance of moderately risen ATP levels during the FCW, which is in turn the cause for the enhancement of ATP-dependent calcium oscillations. Correspondingly, EGTA would increase the apoptotic index of the first stage by promoting a decrease in the calcium oscillatory rate. The fact that the second stage of apoptosis is not affected by the drugs suggests that the two stages are at least partially subject to different signaling pathways.

Effect of the retention ring-assisted continuous application of riboflavin in pulsed-light accelerated corneal collagen cross-linking on the progression of keratoconus.

BACKGROUND: To investigate the efficacy and safety of the retention ring-assisted continuous application of 0.1% riboflavin in pulsed-light accelerated corneal collagen cross-linking on the progression of keratoconus. METHODS: The medical records of 20 eyes of 18 patients with progressive keratoconus who received collagen cross-linking at Seoul National University Hospital were retrospectively reviewed. Isotonic 0.1% riboflavin was continuously applied for 10 min using an 8.0-mm retention ring before the irradiation and accelerated cross-linking was applied with 30-mW pulsed-ultraviolet light at a wavelength 365 nm for eight minutes (1 s on/1 s off; 30 mW/cm2, cumulative dose of 7 .2J/cm2) without further intermittent application of riboflavin. Visual acuity, refractive error, topographic index, corneal thickness, and endothelial cell density were evaluated before the operation and at 1, 3, 6, and 12 months. RESULTS: The best corrected visual acuity in logMAR improved from preoperative 0.43 to 0.17 in 12 months (p = 0.050). Maximum keratometry decreased from 51.8 D to 50.4 D at 6 months (p = 0.015) and 50.1 D at 12 months (p = 0.0003). Astigmatism decreased from preoperative 5.5 D to 4.1 D at 12 months (p < 0.0001). Thinnest corneal thickness decreased at three and 6 months but recovered in 12 months (p > 0.05). Endothelial cell density decreased at postoperative 1 month (p = 0.02) but gradually recovered in 12 months (p > 0.05). CONCLUSIONS: Retention ring-assisted continuous application of riboflavin for 10 minutes in pulsed-light accelerated cross-linking is a comparably safe and effective treatment for halting the progression of keratoconus in 12 months when compared to outcomes of the standard Dresden protocol shown in previous reports.