Search

Using demographic, clinical, visual field, and optical coherence tomography (OCT) variables to study the association of 5-year glaucomatous progression in glaucoma suspect eyes.This is a retrospective longitudinal clinical study. Inclusion criteria consisted of glaucoma suspect eyes (i.e., concerning cup-to-disk ratio and/or intraocular pressure (IOP)21 mm Hg), age ≥ 30 years old, follow-up time of 5 years, best-corrected visual acuity (BCVA) of 20/100 or better, spherical equivalent (SE) higher than 8 diopters and an astigmatism less than 3 diopters. Eyes with glaucoma-determined by two consecutive, reliable visual field tests-were excluded, as well as any eyes with any clinically significant retinal or neurological disease. The percentage of glaucoma suspect eyes, which progressed to glaucoma within a 5-year period, was calculated. Study subjects were divided into the following groups: eyes that progressed to glaucoma and those that did not.In the 288 patients which we looked at, 365 total eyes, 323 eyes had concerning cup-to-disk ratio and 42 had ocular hypertension. Bivariate analysis showed that the eyes which progressed to glaucoma had significantly worse mean deviation, increased pattern standard deviation (PSD), and less visual field index (VFI). Our bivariate analysis also showed a thinner average, superior and inferior retinal nerve fiber layer thickness (RNFL), and more severe average, superior, and inferior RNFL damages (i.e., color grading scale) at baseline. Logistic regression analysis showed that only PSD and severe inferior RNFL damage (i.e., red color) to be significantly associated with 5-year glaucomatous progression.Segmental RNFL damage and pattern standard deviation are associated with 5-year glaucomatous progression in glaucoma suspect eyes.Nassiri N, Das S, Patel V

"Consensual ophthalmotonic reaction" refers to changes in intraocular pressure (IOP) in one eye, which is accompanied by a corresponding change in IOP in the contralateral eye. This study evaluates whether monocular administration of selective laser trabeculoplasty (SLT) leads to a consensual ophthalmotonic reaction and how long this effect lasts.A retrospective chart review was performed on patients receiving SLT at Kresge Eye Institute in Detroit, MI, from January 2015 to August 2016. Patients were excluded if they had previous history of glaucoma incisional and/or laser procedures; required additional laser trabeculoplasty; had glaucoma medication changes during the follow-up period; experienced no decrease in IOP during the follow-up period; or had a diagnosis of angle closure on gonioscopy. Various demographic, clinical, and surgical data were collected. IOP measurements were collected at baseline and postoperatively at 1-3 months, 4-9 months, and 12-15 months.At all follow-up periods, the IOP of the treated eye was decreased from baseline IOP (This study showed that monocular administration of SLT results in a consensual ophthalmotonic reaction. The consensual ophthalmotonic reaction appears to last for up to 4-9 months.Therefore, although SLT does lead to a consensual ophthalmotonic reaction, monocular administration of SLT is not a reliable method of long-term IOP control for the contralateral non-SLT-treated eye.Nassiri N, Mei F, Tokko H

Analyze practice patterns of glaucoma surgeons with and without fellowship training.Physician-specific 2017 Medicare data were obtained from the Centers for MedicareMedicaid Services website. Current Procedural Terminology codes identified procedures including iStent, tube shunts, trabeculectomy, complicated trabeculectomy, endoscopic cyclophotocoagulation, internal Xen Implant, external Xen or Express shunt, Gonioscopy-Assisted Transluminal Trabeculotomy, and Kahook Dual Blade Goniotomy. Physicians with fellowship training were identified via the American Glaucoma Society website.A total of 1547 glaucoma surgeons were identified, of which 319 had completed fellowship training. Overall, fellowship-trained glaucoma surgeons performed more services (50.4 ± 47.0 vs 40.5 ± 35.3, P .001) with a larger variety of procedures (1.8 ± 1.0 vs 1.3 ± 0.6, P .001) than those without fellowship training. Surgeons without fellowship training were more likely to perform iStent and endoscopic cyclophotocoagulation and less likely to perform the remaining procedures than their fellowship-trained counterparts. Medicare payments did not differ between groups and the number of Medicare beneficiaries only differed for internal Xen implant (P = .03). Patient comorbidity burden was similar between groups with about one-third of patients being diagnosed with ischemic heart disease.Surgeons without fellowship training can treat a similar volume of glaucoma patients as those with fellowship training. However, based on surgical procedures employed, their practices are skewed towards mild and moderate glaucoma and they are more limited in their breadth of procedures. In addition, ischemic heart disease was prevalent in the glaucoma patient population.

Vision loss and blindness are among the top ten disabilities in the United States, yet access and utilization of eye care remains low. Vision Detroit aimed to address eye-care disparities via community-based screenings. By investigating burden of eye disease and barriers to eye-care utilization in an underserved urban community, we may direct efforts to improve access.Twenty-three screenings were conducted from March 2015-November 2017. Patient information gathered at screenings were demographics, medical and social history, eye exam/referral history, insurance status, primary care physician (PCP) status, and patient-perceived eye-care barriers.Three-hundred-eighty patients were screened, 42% African American and 51% Hispanic. Average age was 53 ± 16.4 years, 70% reported vision problems, 50% reported over two years of vision problems, and average habitual visual acuity in best-seeing eye was 20/37. Eye-care underutilization was reported in 61% of type-2 diabetics. Older age and PCP recommendations/referrals were associated with increased utilization in all patients. Insurance was the most common barrier (53%); of the 55% insured, 31% reported financial barriers. Employed patients were more likely than unemployed to report a time barrier (odds ratio = 1.76, 95% confidence interval 1.03-3.01). Those with high school or less education reported "unaware of need", "unsure where to go", "transportation", and "insurance" as barriers more often.Visual burden was pervasive, yet access was suboptimal. Financial, logistical, and awareness barriers were common. PCP referral and older age were associated with increased utilization. Those less educated reported more barriers, highlighting the need to address fiscal concerns and eye-health education.

Anion channels in the retinal pigment epithelium (RPE) play an essential role in the transport of Cl− between the outer retina and the choroidal blood to regulate the ionic composition and volume of the subretinal fluid that surrounds the photoreceptor outer segments. Recently, we reported that the anion conductance of the mouse RPE basolateral membrane is highly selective for the biologically active anion thiocyanate (SCN−), a property that does not correspond with any of the Cl− channels that have been found to be expressed in the RPE to date. The purpose of this study was to determine the extent to which SLC26A7, a SCN− permeable-anion exchanger/channel that was reported to be expressed in human RPE, contributes to the RPE basolateral anion conductance. We show by quantitative RT-PCR that Slc26a7 is highly expressed in mouse RPE compared with other members of the Slc26 gene family and Cl− channel genes known to be expressed in the RPE. By applying immunofluorescence microscopy to mouse retinal sections and isolated cells, we localized SLC26A7 to the RPE basolateral membrane. Finally, we performed whole cell and excised patch recordings from RPE cells acutely isolated from Slc26a7 knockout mice to show that the SCN− conductance and permeability of its basolateral membrane are dramatically smaller relative to wild-type mouse RPE cells. These findings establish SLC26A7 as the SCN−-selective conductance of the RPE basolateral membrane and provide new insight into the physiology of an anion channel that may participate in anion transport and pH regulation by the RPE.

To compare three-year surgical outcomes of trabeculectomy versus Ahmed valves in patients with prior failed trabeculectomy.This is a longitudinal retrospective comparative study of one-hundred twenty adult patients with prior failed trabeculectomy who underwent a repeat trabeculectomy or Ahmed valve implant. Demographic and clinical data were collected up to 3 years on all study participants at the Kresge Eye Institute from 2004 to 2016. Visual acuity, intraocular pressure, number of intraocular pressure reducing medications, and success rates at various time points up to 3 years after repeat surgery were the main outcome variables.Sixty-five and sixty eyes were included in the trabeculectomy and the Ahmed valve groups, respectively. Baseline intraocular pressure significantly decreased in both groups at 3 years (p 0.01). The number of medications was relatively similar to baseline in both study groups at 3 years (p 0.05). There was no statistically significant difference between the two groups in visual acuity, percentage of intraocular pressure reduction, number of medications, or success rates at any follow-up time points (p 0.05 for all).After 3 years, both trabeculectomy and Ahmed valves significantly reduced intraocular pressure from baseline, but with relatively similar number of medications compared to baseline. There was no significant difference in any outcome measure between trabeculectomy and Ahmed valves at any follow-up time points. These results may suggest neither trabeculectomy or Ahmed valves are superior in patients with previously failed trabeculectomies.

To describe opioid prescribing practices of ophthalmology subspecialties and determine whether opioid prescribing has decreased during the public health crisis.Retrospective cohort study.Ophthalmologists prescribing at least 11 medications billed to the Medicare Part D prescription drug plan.Publicly available Medicare data sets based on claims from the years 2016, 2017, and 2018 were used. Fellowship status was assumed based on subspecialty society membership or use of specified Current Procedural Terminology codes. The main outcome was the percentage of physicians in each subspecialty prescribing opioids.The database included 19,762, 19,790, and 19,840 ophthalmologists in the years 2016, 2017, and 2018, respectively. Only the subspecialties of comprehensive ophthalmology (43.5% vs 39.6% vs 35.7%; p0.001; φAll subspecialties experienced either a small reduction or no significant change in the percentage of opioid prescribers during the period analyzed. We hope to encourage collaboration between ophthalmology subspecialties in striving to reduce opioid prescribing. Further studies are needed to better fine-tune opioid prescribing practices.

Poor eye-health knowledge and health literacy are pervasive, contributing to worse outcomes. This study aims to examine short- and long-term eye-health knowledge retention following eye-health education interventions in adults. Vision Detroit was an outreach vision screening program that integrated a 5-Point Teaching Intervention (5PTI), at a Southwest Community Center (SW-CC) from 2015–2017. The 5PTI consists of eye-health learning points developed to verbally educate patients. During vision screenings, eye-health knowledge tests were administered before and after 5PTI (Test 1 and Test 2, respectively). In 2016, Community Eye-Health Education Interventions (CHEI) were initiated at the SW-CC. During CHEI, bilingual healthcare students taught voluntary SW-CC members the 5PTI learning points, regardless of participant interest to attend future screenings. CHEI sessions occurred on separate dates prior to vision screening events. Test 1 and Test 2 scores were compared for all participants. Test 1 scores were compared for those who underwent CHEI prior to vision screening (CHEI positive) versus those who did not (CHEI negative). Two-hundred-seventeen adult patients met inclusion criteria, with 75.8% women, 82.6% Hispanic, mean age 50.4 ± 16.2 years, 74.6% had high school or less education, and 49.2% had health insurance. Test 1 to Test 2 scores improved after 5PTI (71.2 ± 26.4% vs. 97.2 ± 9.9%, p < .00001). Forty-eight participants attended CHEI and subsequent vision screening. Test 1 scores were higher among those CHEI positive versus CHEI negative (81.1 ± 2.1% vs. 68.3 ± 3.4%, p = .0027). Simple eye-health education interventions, delivered during vision screenings and via community-based education, can improve eye-health knowledge.

Purpose: To investigate the frequency of conjunctival erosions in a series of patients who underwent Ahmed valve implantation in a tertiary referral center and to study risk factors associated with the conjunctival erosions. Methods: This is a single-center, retrospective case-control study of all patients who underwent Ahmed valve implantation between October 2006 and July 2016 at the Kresge Eye Institute in Detroit, Michigan. The series consisted of 306 eyes (277 patients) that underwent Ahmed valve implantation. The rate of conjunctival erosions was determined. Univariate, bivariate, and Cox-proportional hazard analyses were performed to identify factors associated with conjunctival erosions. Results: During the study period, 23 erosions occurred in 306 eyes (7.52%). Aphakia was significantly more common in eyes with erosions (P < 0.05). Aphakia (P = 0.02), uveitic glaucoma (P = 0.03), and longer post-operative use of topical steroids (P < 0.04) significantly increased the risk of erosions based on the Cox model. There were similar rates of erosions with each type of patch graft. No conjunctival erosion was observed after using the modified scleral tunnel method (n = 10). Conclusions: The overall erosion rate was 7.52% in our series. Uveitic glaucoma, aphakia, and longer post-operative use of topical steroids were significantly associated with conjunctival erosions.

Determining risk factors for posterior capsule opacification will allow for further interventions to reduce the risk of development and thus additional procedures.The purpose of this study was to investigate risk factors associated with development of clinically significant posterior capsule opacification requiring yttrium aluminum garnet (YAG) capsulotomy.Medical records of patients (≥18 years) who underwent cataract surgery between January 1, 2011, and March 31, 2014, at Kresge Eye Institute were reviewed. Three hundred eyes requiring YAG capsulotomy up to 3 years after cataract surgery were included in the YAG capsulotomy group. Three hundred eyes not requiring YAG capsulotomy up to 3 years after cataract surgery were selected via age-matched simple randomization (control group).The YAG capsulotomy group included patients with younger age (65.8 ± 11.3 vs. 70.1 ± 10.6 years, P.001), more men (42.67 vs. 34.67%, P = .04), fewer patients with hypertension (73.00 vs. 83.00%, P.001), and more patients with hydrophilic intraocular lenses (74.67 vs. 47.00%, P.001). Logistic regression analysis demonstrated a negative association between YAG capsulotomy and age (coefficient, -0.04; 95% confidence interval [CI], 0.95 to 0.98; P.001) and hydrophobic intraocular lenses (coefficient, -1.50; 95% CI, 0.15 to 0.33; P.001), and a positive association with presence of glaucoma (coefficient, 0.88; 95% CI, 1.39 to 4.17; P = .002). Elapsed time to YAG capsulotomy was sooner in patients with a history of uveitis (95% CI, 5.10 to 9.70 months; P = .02) and insertion of hydrophilic intraocular lenses (95% CI, 18.67 to 21.57 months; P.001).Results of this study suggest that development of visually significant posterior capsule opacification is associated with younger age, glaucoma, and hydrophilic intraocular lenses, and it occurs earlier among those with hydrophilic intraocular lenses and a history of uveitis.

Our recent electrophysiological analysis of mouse retinal pigment epithelial (RPE) cells revealed that in the presence of 10 mM external thiocyanate (SCN−), voltage steps generated large transient currents whose time-dependent decay most likely results from the accumulation or depletion of SCN− intracellularly. In the present study, we investigated the effects of more physiologically relevant concentrations of this biologically active anion. In whole cell recordings of C57BL/6J mouse RPE cells, we found that, over the range of 50 to 500 µM SCN−, the amplitude of transient currents evoked by voltage steps was proportional to the extracellular SCN− concentration. Transient currents were also produced in RPE cells when the membrane potential was held constant and the external SCN− concentration was rapidly increased by pressure-ejecting 500 µM SCN− from a second pipette. Other results indicate that the time dependence of currents produced by both approaches results from a change in driving force due to intracellular SCN− accumulation or depletion. Finally, by applying fluorescence imaging and voltage-clamping techniques to BALB/c mouse RPE cells loaded with the anion-sensitive dye MQAE, we demonstrated that in the presence of 200 or 500 µM extracellular SCN−, depolarizing voltage steps increased the cytoplasmic SCN− concentration to an elevated steady state within several seconds. Collectively, these results indicate that, in the presence of physiological concentrations of SCN− outside the RPE, the conductance and permeability of the RPE cell membranes for SCN− are sufficiently large that SCN− rapidly approaches electrochemical equilibrium within the cytoplasm when the membrane voltage or external SCN− concentration is perturbed.

The basolateral membrane anion conductance of the retinal pigment epithelium (RPE) is a key component of the transepithelial Cl− transport pathway. Although multiple Cl− channels have been found to be expressed in the RPE, the components of the resting Cl− conductance have not been identified. In this study, we used the patch-clamp method to characterize the ion selectivity of the anion conductance in isolated mouse RPE cells and in excised patches of RPE basolateral and apical membranes. Relative permeabilities ( PA/ PCl) calculated from reversal potentials measured in intact cells under bi-ionic conditions were as follows: SCN− >> ClO4− > [Formula: see text] > I− > Br− > Cl− >> gluconate. Relative conductances ( GA/ GCl) followed a similar trend of SCN− >> ClO4− > [Formula: see text] > I− > Br− ≈Cl− >> gluconate. Whole cell currents were highly time-dependent in 10 mM external SCN−, reflecting collapse of the electrochemical potential gradient due to SCN− accumulation or depletion intracellularly. When the membrane potential was held at −120 mV to minimize SCN− accumulation in cells exposed to 10 mM SCN−, the instantaneous current reversed at −90 mV, revealing that PSCN/ PCl is approximately 500. Macroscopic current recordings from outside-out patches demonstrated that both the basolateral and apical membranes exhibit SCN− conductances, with the basolateral membrane having a larger SCN− current density and higher relative permeability for SCN−. Our results suggest that the RPE basolateral and apical membranes contain previously unappreciated anion channels or electrogenic transporters that may mediate the transmembrane fluxes of SCN− and Cl−.

The daily shedding and renewal of photoreceptor outer segments (OS) is critical for maintaining vision. This process relies on the efficient uptake, degradation, and sorting of shed OS material by the retinal pigment epithelium (RPE). Poor OS degradation has been linked to retinal degenerations such as Stargardt disease and may contribute to macular degeneration. While primary human fetal RPE cultures have emerged as a valuable model of in vivo human RPE function, surprisingly few studies have utilized the model for tracking the degradation and fate of OS components in the RPE. Here, we establish an improved platform for studying this topic by modifying existing protocols and creating new methods. Our human fetal culture model facilitates studies of RPE secretion in response to OS ingestion, preserves RPE differentiation and polarization during live-cell imaging of OS phagocytosis, and minimizes costs. We optimize Mer tyrosine kinase-dependent OS phagocytosis assays specifically in human fetal cultures and provide a simple and accurate method for measuring total OS consumption by the RPE. Finally, we utilize chemical transfection, dextran labeling, and immunocytochemistry to evaluate key players in OS degradation, including lysosomes and autophagy proteins. To facilitate quantification of autophagy vesicles, we develop customized image analysis macros in the Fiji/ImageJ software environment. These protocols will facilitate a broad range of studies in human fetal RPE cultures aimed at determining the ultimate fate of OS components after ingestion, a critical step in understanding the pathogenesis of numerous retinal diseases.

Purpose To report a patient with retinal and choroidal vascular occlusion as a presenting sign of sickle cell trait following the development of aqueous misdirection syndrome. Methods Retrospective chart review. Results A patient treated for bilateral chronic angle-closure glaucoma with sequential EX-PRESS glaucoma filtration device surgery developed sequential bilateral aqueous misdirection syndrome. The left eye developed retinal arterial and localized choroidal vascular occlusions subsequent to an acute elevation in intraocular pressure and possibly the use of oral acetazolamide. The patient was subsequently found to have sickle cell trait. The right eye developed aqueous misdirection with acute elevation of intraocular pressure as well, but the patient was not treated with oral acetazolamide and did not develop vascular occlusion. Conclusion Retinal and choroidal vascular occlusions can be the presenting sign of a patient with sickle cell trait. Sickle cell screening may be beneficial in African American or Middle Eastern patients after an acute rise in intraocular pressure, particularly before initiation of treatment with oral carbonic anhydrase inhibitors.

Human, monkey, and bovine retinal pigment epithelial (RPE) cells exhibit an M-type K+ current, which in many other cell types is mediated by channels composed of KCNQ α-subunits and KCNE auxiliary subunits. Recently, we demonstrated the expression of KCNQ1, KCNQ4, and KCNQ5 in the monkey RPE. Here, we investigated the expression of KCNQ and KCNE subunits in native bovine RPE. RT-PCR analysis revealed the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in the RPE, but, in Western blot analysis of RPE plasma membranes, only KCNQ5 was detected. Among the five members of the KCNE gene family, transcripts for KCNE1, KCNE2, KCNE3, and KCNE4 were detected in bovine RPE, but only KCNE1 and KCNE2 proteins were detected. Immunohistochemistry of frozen bovine retinal sections revealed KCNE1 expression near the apical and basal membranes of the RPE, in cone outer segments, in the outer nuclear layer, and throughout the inner retina. The localization of KCNE1 in the RPE basal membrane, where KCNQ5 was previously found to be present, suggests that this β-subunit may contribute to M-type K(+) channels in this membrane.

Inwardly rectifying K+(Kir) channels in the apical membrane of the retinal pigment epithelium (RPE) contribute to extracellular K+homeostasis in the distal retina by mediating K+secretion. Multiple lines of evidence suggest that these channels are composed of Kir7.1. Previously, we showed that native Kirchannels in bovine RPE are modulated by changes in intracellular pH in the physiological range. In the present study, we used the Xenopus laevis oocyte expression system to investigate the pH dependence of cloned human Kir7.1 channels and several point mutants involving histidine residues in the NH2and COOH termini. Kir7.1 channels were inhibited by strong extracellular acidification and modulated by intracellular pH in a biphasic manner, with maximal activity at about intracellular pH (pHi) 7.0 and inhibition by acidification or alkalinization. Replacement of histidine 26 (H26) in the NH2terminus with alanine eliminated the requirement of protons for channel activity and increased sensitivity to proton-induced inhibition, resulting in maximal channel activity at alkaline pHiand smaller whole cell currents at resting pHicompared with wild-type Kir7.1. When H26 was replaced with arginine, the pHisensitivity profile was similar to that of the H26A mutant but with the p Kashifted to a more acidic value, giving rise to whole cell current amplitude at resting pHithat was comparable to that of wild-type Kir7.1. These results indicate that Kir7.1 channels are modulated by intracellular protons by diverse mechanisms and suggest that H26 is important for channel activation at physiological pHiand that it influences an unidentified proton-induced inhibitory mechanism.

Previous studies on bovine retinal pigment epithelium (RPE) established that Kir7.1 channels compose this epithelium's large apical membrane K + conductance. The purpose of this study was to determine whether Kir7.1 and potential Kir7.1 splice variants are expressed in native adult human RPE and, if so, to determine their function and how they are generated. RT-PCR analysis indicated that human RPE expresses full-length Kir7.1 and a novel Kir7.1 splice variant, designated Kir7.1S. Analysis of the human Kir7.1 gene ( KCNJ13 ) organization revealed that it contains three exons, two introns, and a novel alternative 5′ splice site in exon 2. In human RPE, the alternative usage of two competing 5′ splice sites in exon 2 gives rise to transcripts encoding full-length Kir7.1 and Kir7.1S, which is predicted to encode a truncated protein. Real-time PCR indicated that Kir7.1 transcript is nearly as abundant as GAPDH mRNA in human RPE whereas Kir7.1S transcript expression is 4-fold lower. Western blot analysis showed that the splice variant is translated in Xenopus oocytes injected with Kir7.1S cRNA and revealed the expression of full-length Kir7.1 but not Kir7.1S in adult human RPE. Co-expression of Kir7.1 with Kir7.1S in Xenopus oocytes had no effect on either the kinetics or amplitude of Kir7.1 currents. This study confirms the expression of Kir7.1 in human RPE, identifies a Kir7.1 splice variant resulting in predicted changes in protein sequence, and indicates that there is no functional interaction between this splice variant and full-length Kir7.1.

Reactive oxygen metabolites (ROMs) may contribute to several eye diseases, such as age-related macular degeneration, although the underlying mechanisms are unclear. The present study shows that human photoreceptor outer segments (POS) prime human retinal pigment epithelial (RPE) cells for massive ROM release in response to lipopolysaccharide (LPS) and interferon-γ. However, no ROM priming of human RPE cells is observed for bovine POS. ROM production appears to be linked with underlying metabolic oscillations involving the hexose monophosphate shunt.

Retinal pigment epithelial (RPE) cells mediate the recognition and clearance of effete photoreceptor outer segments (POS), a process central to the maintenance of normal vision. Given the emerging importance of Toll-like receptors (TLRs) in transmembrane signaling in response to invading pathogens as well as endogenous substances, we hypothesized that TLRs are associated with RPE cell management of POS. TLR4 clusters on human RPE cells in response to human, but not bovine, POS. However, TLR4 clustering could be inhibited by saturating concentrations of an inhibitory anti-TLR4 mAb. Furthermore, human POS binding to human RPE cells elicited transmembrane metabolic and calcium signals within RPE cells, which could be blocked by saturating doses of an inhibitory anti-TLR4 mAb. However, the heterologous combination of bovine POS and human RPE did not trigger these signals. The pattern recognition receptor CD36 collected at the POS–RPE cell interface for both homologous and heterologous samples, but human TLR4 only collected at the human POS–human RPE cell interface. Kinetic experiments of human POS binding to human RPE cells revealed that CD36 arrives at the POS–RPE interface followed by TLR4 accumulation within 2 min. Metabolic and calcium signals immediately follow. Similarly, the production of reactive oxygen metabolites (ROMs) was observed for the homologous human system, but not the heterologous bovine POS–human RPE cell system. As (a) the bovine POS/human RPE combination did not elicit TLR4 accumulation, RPE signaling, or ROM release, (b) TLR4 arrives at the POS–RPE cell interface just before signaling, (c) TLR4 blockade with an inhibitory anti-TLR4 mAb inhibited TLR4 clustering, signaling, and ROM release in the human POS–human RPE system, and (d) TLR4 demonstrates similar clustering and signaling responses to POS in confluent RPE monolayers, we suggest that TLR4 of RPE cells participates in transmembrane signaling events that contribute to the management of human POS.

Purpose. To determine the expression and functional activity of proton-coupled oligopeptide transporters (POT) in retinal pigment epithelial (RPE) cells.

To evaluate the long-term effectiveness of glaucoma management in patients undergoing primary glaucoma triple procedure (PGTP) with and without adjunctive subconjunctival mitomycin-C (MMC).Case-controlled study.Of the 203 eyes of 203 primary open-angle glaucoma (POAG) patients who had undergone PGTP and in whom reliable Humphrey visual fields had been obtained both before and after surgery at 13.5 +/- 8.9 and 27.9 +/- 8.9 months, 124 of the 144 eyes that received MMC during surgery were matched to the other 59 eyes that did not with respect to cup-to-disc ratio and risk factors for filtration failure in addition to other variables.Both preoperative and postoperative intraocular pressure (IOP), Humphrey visual fields and their global indices, number of glaucoma medications, and best-corrected visual acuity (BCVA).There were no significant differences in demographics between the two groups (P0.05 for each). Whereas both the control and the MMC groups attained significant decreases of mean IOP (18.5 +/- 5.7 mmHg-;15.6 +/- 4.6 mmHg, P = 0.0014; 19.3 +/- 7.0 mmHg-13.7 +/- 4.9 mmHg, P = 0.0001) and mean number of medications (2.1 +/- 1.3-1.3 +/- 1.3, P = 0.0001; 2.3 +/- 1.2-1.0 +/- 1.3, P = 0.0001) at 36 months after surgery, the MMC group had significantly lower mean IOP than the control group at all postoperative visits (P0.05 for each). The MMC group also tended to have less medical dependency after surgery than the control group. There was no significant difference in postoperative BCVA between the two groups. Patients in both groups had mean visual acuity of 20/30 or better. There was a significant worsening of corrected pattern standard deviation (CPSD) in the control group (3.97 +/- 3.18-5.17 +/- 3.36, P = 0.001) compared with no significant change in the MMC group (5.07 +/- 4.11-5.23 +/- 3.36, P = 0.93). The mean deviation did not change significantly in either group.The long-term glaucoma management in POAG patients with cataract undergoing PGTP indicates a successful outcome in final IOP, medical dependency, and BCVA. Furthermore, the MMC group had better IOP control and stable visual fields (CPSD), whereas the control group had a significant worsening of CPSD.

Human, monkey, and bovine retinal pigment epithelial (RPE) cells exhibit an M-type K+ current, which in many other cell types is mediated by channels composed of KCNQ α-subunits and KCNE auxiliary subunits. Recently, we demonstrated the expression of KCNQ1, KCNQ4, and KCNQ5 in the monkey RPE. Here, we investigated the expression of KCNQ and KCNE subunits in native bovine RPE. RT-PCR analysis revealed the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in the RPE, but, in Western blot analysis of RPE plasma membranes, only KCNQ5 was detected. Among the five members of the KCNE gene family, transcripts for KCNE1, KCNE2, KCNE3, and KCNE4 were detected in bovine RPE, but only KCNE1 and KCNE2 proteins were detected. Immunohistochemistry of frozen bovine retinal sections revealed KCNE1 expression near the apical and basal membranes of the RPE, in cone outer segments, in the outer nuclear layer, and throughout the inner retina. The localization of KCNE1 in the RPE basal membrane, where KCNQ5 was previously found to be present, suggests that this β-subunit may contribute to M-type K(+) channels in this membrane.

We report a novel 3-dimensional model for visualizing tumor cell migration across a nylon mesh-supported gelatin matrix. To visualize migration across these model barriers, cell proteolytic activity of the pericellular matrix was detected using Bodipy-BSA (fluorescent upon proteolysis) and DQ collagen (fluorescent upon collagenase activity). For 3-dimensional image reconstruction, multiple optical images at sequential z axis positions were deconvoluted by computer analysis. Specificity was indicated using well-known inhibitors. Using these fluorescent proteolysis markers and imaging methods, we have directly demonstrated proteolytic and collagenolytic activity during tumor cell invasion. Moreover, it is possible to visualize migratory pathways followed by tumor cells during matrix invasion. Using cells of differing invasive potentials (uPAR-negative T-47D wild-type and uPAR-positive T-47D A2--1 cells), we show that the presence of the T-47D-A2--1 cells facilitates the entry of T-47D wild-type cells into the matrix. In some cases, wild-type cells follow T-47D A2--1 cells into the matrix whereas other T-47D-wild-type cells appear to enter without the direct intervention of T-47D A2--1 cells. Thus, we have developed a new 3-dimensional model of tumor cell invasion, demonstrated protein and collagen disruption, mapped the pathways followed by tumor cells during migration through an extracellular matrix, and illustrated cross-talk among tumor cell populations during invasion.

Bovine Kir7.1 clones were obtained from a retinal pigment epithelium (RPE)-subtracted cDNA library. Human RPE cDNA library screening resulted in clones encoding full-length human Kir7.1. Northern blot analysis indicated that bovine Kir7.1 is highly expressed in the RPE. Human Kir7.1 channels were expressed in Xenopus oocytes and studied using the two-electrode voltage-clamp technique. The macroscopic Kir7.1 conductance exhibited mild inward rectification and an inverse dependence on extracellular K+ concentration ([K+]o). The selectivity sequence based on permeability ratios was K+ (1.0) approximately Rb+ (0.89) > Cs+ (0.013) > Na+ (0.003) approximately Li+ (0.001) and the sequence based on conductance ratios was Rb+ (9.5) >> K+ (1.0) > Na+ (0.458) > Cs+ (0.331) > Li+ (0.139). Non-stationary noise analysis of Rb+ currents in cell-attached patches yielded a unitary conductance for Kir7.1 of approximately 2 pS. In whole-cell recordings from freshly isolated bovine RPE cells, the predominant current was a mild inwardly rectifying K+ current that exhibited an inverse dependence of conductance on [K+]o. The selectivity sequence based on permeability ratios was K+ (1.0) approximately Rb+ (0.89) > Cs+ (0.021) > Na+ (0.003) approximately Li+ (0.002) and the sequence based on conductance ratios was Rb+ (8.9) >> K+ (1.0) > Na+ (0.59) > Cs+ (0.23) > Li+ (0.08). In cell-attached recordings with Rb+ in the pipette, inwardly rectifying currents were observed in nine of 12 patches of RPE apical membrane but in only one of 13 basolateral membrane patches. Non-stationary noise analysis of Rb+ currents in cell-attached apical membrane patches yielded a unitary conductance for RPE Kir of approximately 2 pS. On the basis of this molecular and electrophysiological evidence, we conclude that Kir7.1 channel subunits comprise the K+ conductance of the RPE apical membrane.

To identify novel potassium channel genes expressed in the retina, we screened a human retina cDNA library with an EST sequence showing partial homology to inwardly rectifying potassium (Kir) channel genes. The isolated cDNA yielded a 2,961-base pair sequence with the predicted open reading frame showing strong homology to the rat Kir2.4 (rKir2.4). Northern analysis of mRNA from human and bovine tissues showed preferential expression of Kir2.4 in the neural retina. In situ hybridization to sections of monkey retina detected Kir2.4 transcript in most retinal neurons. Somatic hybridization analysis and dual-color in situ hybridization to metaphase chromosomes mapped Kir2.4 to human chromosome 19 q13.1–q13.3. Expression of human Kir2.4 cRNA in Xenopus oocytes generated strong, inwardly rectifying K+ currents that were enhanced by extracellular alkalinization. We conclude that human Kir2.4 encodes an inwardly rectifying K+ channel that is preferentially expressed in the neural retina and that is sensitive to physiological changes in extracellular pH.

The authors compared the efficacy of apraclonidine 1% versus pilocarpine 4% prophylaxis of post-argon laser trabeculoplasty (ALT) intraocular pressure (IOP) spike.Prospective randomized clinical trial.Two hundred twenty-eight eyes of 228 patients with primary open-angle glaucoma undergoing ALT were studied.Patients were given 1 drop of either apraclonidine 1% (n = 114) or pilocarpine 4% (n = 114) 15 minutes before ALT.Peri-ALT IOPs and incidences of post-ALT IOP spikes at 5 minutes, 1 hour, and 24 hours were compared between the two groups.The two groups were similar in age, race, and medical dependency. Post-ALT mean IOPs at 5 minutes, 1 hour, and 24 hours were significantly lower than pre-ALT mean IOPs in both apraclonidine (P0.001) and pilocarpine (P0.001) groups. Incidences of IOP spikes greater than 1, 3, and 5 mmHg at 1 hour post-ALT were 21.1%, 14.9%, and 8.8% for the apraclonidine group and 12.3%, 5.3%, and 4.4% for the pilocarpine group (P = 0.076, 0.015, and 0.18 chi-square test). In the apraclonidine prophylaxis group, patients on long-term apraclonidine showed significantly higher incidence of post-ALT IOP spike than the patients without such long-term apraclonidine use (35.7%, 15 of 42 eyes, vs. 12.5%, 9 of 72 eyes; P = 0.003). In addition, peri-ALT pilocarpine prophylaxis tended to be less effective in patients undergoing long-term pilocarpine therapy but without statistical significance (17.4%, 8 of 46 eyes, vs. 9.4%, 6 of 64 eyes; P = 0.17).Peri-ALT pilocarpine 4% was at least as effective as, if not more effective than, apraclonidine 1% in post-ALT IOP spike prophylaxis. Peri-ALT apraclonidine prophylaxis was not effective in patients on long-term apraclonidine, and peri-ALT pilocarpine prophylaxis tended to be less effective in patients undergoing long-term pilocarpine therapy. Pilocarpine 4% can be considered as a first-choice drug for post-ALT IOP spike prophylaxis, especially in patients under treatment with apraclonidine.

Objective To assess the efficacy of latanoprost additive therapy in patients with intraocular pressure (IOP) out of control while taking maximum-tolerated medications and to determine whether pilocarpine therapy has a dose-dependent adverse effect on the efficacy of latanoprost therapy. Design Noncomparative case series. Participants Sixty-one eyes of 61 patients with chronic glaucoma with IOP out of control while receiving maximum-tolerated medications were treated with latanoprost additive therapy on a compassionate basis. Main outcome measures Follow-up was up to 22 months with a mean of 13.9 ± 5.7 months. Kaplan-Meier survival analysis with Mantel-Cox log-rank test was performed to determine the overall success of latanoprost additive therapy and to compare the success rates of high-dose pilocarpine, low-dose pilocarpine, and no pilocarpine therapies. The criterion for success was avoiding glaucoma surgery with IOP decrease of 20% or greater and final IOP less than 22 mmHg. The IOP change and its significance for patients satisfying and failing the criterion for success also were determined to assess the latanoprost additive therapy. In addition, a number of pretreatment variables, including pilocarpine therapy, were analyzed for a significant effect on the efficacy of latanoprost additive therapy using Cox proportional hazards regression analysis. Results Latanoprost additive therapy significantly lowered mean IOP by 3.9 ± 5.5 mmHg at 3 months and by 3.5 ± 5.8 mmHg at 12 months. The cumulative success rate of the latanoprost additive therapy was 70% at 1 month, 42% at 3 months, 40% at 6 months, and 30% at 12 months. Of the variables studied, only increased number of previous incisional glaucoma surgeries and IOP greater than 24 mmHg before latanoprost additive therapy were significant prognostic factors for failure of latanoprost additive therapy. Pilocarpine therapy in any dose had no significant effect. Conclusion This study supports a trial of latanoprost additive therapy before glaucoma surgery in patients with IOP out of control while receiving maximum-tolerated medications irrespective of pilocarpine therapy and the pilocarpine dosage, especially when the number of previous incisional glaucoma surgery is less than three and the IOP is less than 25 mmHg.

To investigate the effect of adjunctive mitomycin C on primary glaucoma triple procedure in patients with primary open-angle glaucoma with and without one or more of the prognostic factors for filtration failure of primary glaucoma triple procedure. Those factors include being of African-American race, having a preoperative intraocular pressure of 20 mm Hg or more on maximum tolerated medications, and being on two or more medications preoperatively.Study patients consisted of 197 consecutive patients with primary open-angle glaucoma who were randomly assigned to receive either no adjunctive mitomycin C (101 eyes of 101 patients) or to receive adjunctive subconjunctival mitomycin C (96 eyes of 96 patients) during the primary glaucoma triple procedure. Kaplan-Meier survival analysis comparisons were made between respective subgroups with and without prognostic indicators for filtration failures using a relatively stringent set of criteria for filtration success of primary glaucoma triple procedure.There was no statistically significant (P = .117) difference in filtration success of primary glaucoma triple procedure between the control and mitomycin C groups. Adjunctive mitomycin C significantly (P.05) improved the filtration outcome of the primary glaucoma triple procedure in the subgroups with each of the three prognostic factors for filtration failure of primary glaucoma triple procedure. On the other hand, in the subgroups without the prognostic factors, adjunctive mitomycin C did not significantly (P.05) change the filtration outcome of the primary glaucoma triple procedure.These findings establish the basis for selective use of mitomycin C in patients with primary open-angle glaucoma undergoing primary glaucoma triple procedure.

The translocation of ions, fluid and macromolecules across epithelia is made possible by the asymmetric distribution of transport proteins, enzymes and receptors in two physically distinct plasma membrane domains that form the apical and basolateral sides of the cell. Each side faces a different extracellular environment. In the back of the vertebrate eye, the retinal pigment epithelium (RPE) apical membrane receives a continuous stream of paracrine signals that are generated by a variety of retinal neurons in the light and dark. These signals help regulate RPE function, and conversely, alterations in RPE function can modify the activity of retinal neurons. At the basolateral surface, there is a continual exchange of nutrients and waste products, along with a flow of hormonal signals from the choroidal blood supply, all of which serve to maintain the health and integrity of the distal retina and in particular, the photoreceptors. This review provides an integrated summary of the apical and basolateral membrane and intracellular signaling mechanisms that mediate the vectorial traffic of ions and fluid across the RPE. These same mechanisms help regulate the chemical milieu within the cell and in the extracellular spaces that surround the cell. They also generate specific components of the electrical signals that are recorded clinically across the intact human eye, the electroretinogram (ERG) and the electrooculogram (EOG). The last part of this review is focused on the light-induced photoreceptor-dependent decrease in subretinal potassium concentration ([K] 0 ) that occurs in the intact eye and serves as a paracrine signal for the RPE. This signal plays a central role in regulating RPE physiology and in mediating retina/RPE interactions, following transitions between light and dark; it is mimicked in vitro by a small (3 m m ) change in [K] 0 on the apical side of the epithelium. The clinical implications are discussed in terms of the transport mechanisms that regulate hydration of the subretinal space and that potentially mediate fluid absorption out of the retina.

Objective: To compare silicone versus acrylic foldable intraocular lens (IOL) implantation in primary glaucoma triple procedure (PGTP). Design: Prospective, randomized. Participants: A total of 79 eyes of 79 primary open-angle glaucoma (POAG) patients in need of combined surgery were randomized to a silicone IOL group (36 eyes) and acrylic IOL group (43 eyes). Intervention: The study eyes underwent PGTP, which consisted of primary trabeculectomy, phacoemulsification, and posterior chamber IOL implantation. Adjunctive mitomycin C (MMC) (0.5 mg/ml for 1 minute) was used selectively only in patients with one or more risk factors for filtration failure of PGTP. Main Outcome Measures: Snellen visual acuity, intraocular pressure (IOP), slitlamp biomicroscopy, and number of glaucoma medications were measured, performed, or determined preoperatively and at regular intervals postoperatively. Results: There were no significant differences in the mean number of postoperative glaucoma medications at 1, 2, 3, 4–6, and 9–12 months and at last follow-up ( P > 0.05); mean change in corrected visual acuity best attained ( P = 0.315) or at last followup ( P = 0.223) between the silicone and acrylic groups. Both groups had significant decreases in mean IOP and mean number of medications postoperatively at all times ( P 25 mmHg and IOP spike > 5 mmHg above preoperative IOP during the first month were significantly higher in the acrylic group ( P = 0.026). The mean postoperative IOP at 1 month in the acrylic group was also significantly higher than the silicone group (14.1 ± 5.0, 11.2 ± 3.9, P = 0.005). Conversely, there were no significant differences in mean postoperative IOP at 2, 3, 4–6, and 9–12 months and at last follow-up between the silicone and acrylic groups ( P > 0.05). Suture removal or release occurred significantly more frequently in the acrylic IOL group during the first month and the first 2 months (48.8% and 60.5%) than the silicone group (25.0% and 36.1%, P = 0.030 and 0.031, respectively). There were no significant differences in postoperative complications or surgical interventions between the two groups ( P > 0.05). Conclusions: During the first year following the PGTP with selective use of MMC, there were no significant differences in the medical dependency or visual outcomes or complications between the silicone and acrylic groups. Both groups attained significant decreases in IOP postoperatively. However, the mean IOP was significantly higher in the acrylic than the silicone group at 1 month postoperatively, and postoperative IOP > 25 mmHg and IOP spike > 5 mmHg above preoperative IOP were significantly greater in the acrylic group. There were significantly more suture releases in the acrylic IOL group than the silicone IOL group in the first 2 months postoperatively.

Outwardly rectifying K+ currents in freshly isolated bovine retinal pigment epithelial (RPE) cells were characterized using the whole cell and perforated-patch configurations of the patch-clamp technique. All cells exhibited a delayed rectifier type K+ current. This current had an activation threshold voltage of approximately -40 mV, activated with a sigmoidal trajectory, and inactivated completely over a period of several seconds. External tetraethylammonium (TEA) was an effective blocker of the delayed rectifier current [apparent dissociation constant (Kd) = 5.1 mM], but external Ba2+ was relatively ineffective. Approximately 24% of the cells also exhibited a sustained outwardly rectifying K+ current that became activated at voltages positive to approximately -80 mV. This current resembled the neuronal M-current. External Ba2+ was a potent blocker of this current (apparent Kd = 1.1 mM), but external TEA and Cs+ were relatively ineffective. These results indicate that freshly isolated bovine RPE cells express K+ currents of both the delayed rectifier and M types. The latter may contribute to the resting K+ conductances of the apical and basolateral membranes.

Purpose: The prerequisite for selective use of adjunctive mitomycin C (MMC) in primary glaucoma triple procedure (PGTP) is a better understanding of the prognostic factors. Therefore, the authors carried out the current study on the outcome of PGTP with and without adjunctive MMC to determine the prognostic factors for filtration failure of PGTP. Methods: The study patients consisted of 174 consecutive primary open-angle glaucoma (POAG) patients undergoing PGTP. They were assigned randomly to either no adjunctive MMC (93 eyes of 93 patients) or adjunctive subconjunctival MMC (81 eyes of 81 patients) during the PGTP. After surgery, the patients were examined at regular intervals for visual acuity, intraocular pressure (IOP) control, medical therapy requirements, and complications. Filtration failure was determined according to two different criteria: (1) a less stringent set of criteria (criterion I) and (2) a more stringent set of criteria (criterion II). Results: There were no statistically significant differences between the groups with and without adjunctive MMC with respect to postoperative IOP, number of medications, and visual acuity outcome during mean follow-up (± standard deviation) of 25.1 ± 5.5 months ( P > 0.05 for each). However, there were prognostic factors for filtration failure in the control group but not in the MMC group. Black race, diabetes mellitus, preoperative IOP greater than or equal to 20 mmHg, and number of preoperative medications greater than two were found to be significant prognostic factors for filtration failure by criterion I. Black race, preoperative IOP greater than or equal to 20 mmHg and number of preoperative medications greater than 1 were significant prognostic factors for filtration failure by criterion II. Conclusion: There was no statistically significant difference in the overall outcome of PGTP between control and MMC groups of nonselected patients with primary openangle glaucoma. Black race, diabetes mellitus (by criterion I only), preoperative IOP greater than or equal to 20 mmHg, and number of preoperative medications greater than 2 (by criterion I) or greater than 1 (by criterion II) were found to be significant independent prognostic factors for filtration failure of PGTP without adjunctive MMC but not with adjunctive MMC. The use of adjunctive subconjunctival MMC in PGTP may have to be selective, primarily in those patients with primary open-angle glaucoma with one or more of the prognostic factors for filtration failure.

KCNJ13 encodes Kir7.1, an inwardly rectifying K+channel that is expressed in multiple ion-transporting epithelia. A mutation in KCNJ13 resulting in an arginine-to-tryptophan change at residue 162 (R162W) of Kir7.1 was associated with snowflake vitreoretinal degeneration, an inherited autosomal-dominant disease characterized by vitreous degeneration and mild retinal degeneration. We used the Xenopus laevis oocyte expression system to assess the functional properties of the R162W (mutant) Kir7.1 channel and determine how wild-type (WT) Kir7.1 is affected by the presence of the mutant subunit. Recordings obtained via the two-electrode voltage-clamp technique revealed that injection of oocytes with mutant Kir7.1 cRNA resulted in currents and cation selectivity that were indistinguishable from those in water-injected oocytes, suggesting that the mutant protein does not form functional channels in the plasma membrane. Coinjection of oocytes with equal amounts of mutant and WT Kir7.1 cRNAs resulted in inward K+and Rb+currents with amplitudes that were ∼17% of those in oocytes injected with WT Kir7.1 cRNA alone, demonstrating a dominant-negative effect of the mutant subunit. Similar to oocytes injected with WT Kir7.1 cRNA alone, coinjected oocytes exhibited inwardly rectifying Rb+currents that were more than seven times larger than K+currents, indicating that mutant subunits did not alter Kir7.1 channel selectivity. Immunostaining of Xenopus oocytes or Madin-Darby canine kidney cells expressing mutant or WT Kir7.1 demonstrated distribution of both proteins primarily in the plasma membrane. Our data suggest that the R162W mutation suppresses Kir7.1 channel activity, possibly by negatively impacting gating by membrane phosphadidylinositol 4,5-bisphosphate.

Currents in freshly dissociated adult human retinal pigment epithelial (RPE) cells were studied using the perforated patch-clamp technique. The zero-current potential (V0) averaged -48.9 +/- 7.7 mV (n = 50). Depolarizing voltage pulses from -70 mV evoked an outward current that activated with first-order kinetics and that did not inactivate during prolonged depolarizations. Repolarizing the membrane potential produced tail currents that reversed near the K+ equilibrium potential, indicating that the sustained outward current was carried mainly by K+. The outwardly rectifying K+ conductance (gK) had an activation threshold voltage near -60 mV and was half-maximal at -37 mV. Approximately 25% of gK was active at the average V0. The K+ current was nearly completely blocked by 2 mM Ba2+ but was relatively insensitive to 20 mM tetraethylammonium. The kinetics, voltage dependence, and blocker sensitivity of this current clearly distinguish it from delayed rectifier K+ currents previously identified in RPE cells. We conclude that the sustained outward K+ current may help establish the resting potential of the apical and/or basolateral membranes and may also participate in K+ transport across the RPE.

Intracellular microelectrode techniques were employed to characterize the blocker sensitivity of the K+ conductance (gK) at the apical membrane of the toad retinal pigment epithelium (RPE). Increasing the K+ concentration in the apical bath ([K+]o) from 2 to 5 mM produced a rapid depolarization of the apical membrane potential (VA). The addition of 0.5 mM Ba2+ or 5 mM Cs+ to the apical bath rapidly depolarized VA and increased the transepithelial resistance and ratio of apical-to-basolateral membrane resistance. In the presence of apical Ba2+ or Cs+, the response of VA to delta [K+]o was markedly reduced, indicating that these ions are effective blockers of apical gK. The Ba(2+)- and Cs(+)-induced decreases in the apparent apical-to-basolateral membrane conductance ratio were concentration dependent, with apparent dissociation constants of 17 microM and 0.5 mM, respectively. The apparent blocker sensitivity of apical gK is similar to that previously demonstrated for the inwardly rectifying K+ conductance in isolated toad RPE cells, suggesting that the inwardly rectifying K+ conductance comprises much of apical gK.

Purpose: In 1987, the authors reported the successful eradication of infection in 16 patients with culture-proven acute exogenous bacterial endophthalmitis using intravitreal but no systemic antibiotics. They retrospectively reviewed additional consecutive cases since then to determine if the initial omission of systemic antibiotics remained reasonable. Methods: Twenty patients had culture-proven endophthalmitis. Four patients initially received systemic antibiotics for orbital cellulitis (2 patients), prevention of a possible scleral buckle infection (1 patient), and ascending cholangitis (1 patient). The remaining 16 patients were treated initially with intravitreal antibiotics only. Findings: Three of these additional 16 patients ultimately required systemic antibiotics for orbital cellulitis (1 patient), infectious scleritis (1 patient), and prevention of central nervous system infection with Neisseria meningitidis (1 patient). Only in one patient who had a neglected endophthalmitis and in whom an orbital cellulitis ultimately developed were we unable to clear the intraocular infection. In the overall series of 32 patients, cultures yielded staphylococcal species in 16 eyes, gram-positive bacilli in 3, streptococcal infection in 5, gram-negative cocci in 1, and gram-negative bacilli in 7. Half of the 14 specimens (1 aqueous and 13 vitreal) collected at the time of 16 reinjections in 13 eyes yielded organisms. Half (16/32) of the eyes attained visual acuity of 20/40 or better; 87.5% (28/32) attained visual acuity of 20/400 or better. Conclusions: Therapy with intravitreal antibiotics without systemic antibiotics is reasonable, unless the infection has extended (or is at risk to extend) beyond the globe. Such evidence includes an elevated temperature or leukocyte count, corneal ring abscess, proptosis, loss of extraocular movements, scleral abscesses or infectious scleritis, and, perhaps, the presence of a scleral buckle.

PURPOSE To study the impact of ocular hypotensive lipids (OHL) on the incidence, progression, and response to treatment of clinically significant diabetic macular edema (CSDME). METHODS A total of 379 patients (232 female, 147 male) with a history of diabetes mellitus (DM) and primary open-angle glaucoma (POAG) were identified and included in the study. Patients were stratified into groups based on CSDME development and OHL exposure. Main outcome measures included time to development of CSDME, total duration of OHL exposure, and duration of DM and POAG. RESULTS Seven patients (1.8%) developed CSDME after OHL exposure (group 1A), 15 (4.0%) developed CSDME prior to OHL exposure (group 1B), and 197 (52.0%) were treated with OHL but never developed CSDME (group 2). Of patients not exposed to OHL, 22 (5.8%) developed CSDME (group 3) and 138 (36.4%) did not (group 4). Mean duration of DM was longer (p

Recently, we demonstrated the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in monkey retinal pigment epithelium (RPE) and showed that the M-type current in RPE cells is blocked by the specific KCNQ channel blocker XE991. Using patch-clamp electrophysiology, we investigated the pharmacological sensitivity of the M-type current in isolated monkey RPE cells to elucidate the subunit composition of the channel. Most RPE cells exhibited an M-type current with a voltage for half-maximal activation of approximately −35 mV. The M-type current activation followed a double-exponential time course and was essentially complete within 1 s. The M-type current was inhibited by micromolar concentrations of the nonselective KCNQ channel blockers linopirdine and XE991 but was relatively insensitive to block by 10 μM chromanol 293B or 135 mM tetraethylammonium (TEA), two KCNQ1 channel blockers. The M-type current was activated by 1) 10 μM retigabine, an opener of all KCNQ channels except KCNQ1, 2) 10 μM zinc pyrithione, which augments all KCNQ channels except KCNQ3, and 3) 50 μM N-ethylmaleimide, which activates KCNQ2, KCNQ4, and KCNQ5, but not KCNQ1 or KCNQ3, channels. Application of cAMP, which activates KCNQ1 and KCNQ4 channels, had no significant effect on the M-type current. Finally, diclofenac, which activates KCNQ2/3 and KCNQ4 channels but inhibits KCNQ5 channels, inhibited the M-type current in the majority of RPE cells but activated it in others. The results indicate that the M-type current in monkey RPE is likely mediated by channels encoded by KCNQ4 and KCNQ5 subunits.

Previous studies identified in retinal pigment epithelial (RPE) cells an M-type K+ current, which in many other cell types is mediated by channels encoded by KCNQ genes. The aim of this study was to assess the expression of KCNQ genes in the monkey RPE and neural retina. Application of the specific KCNQ channel blocker XE991 eliminated the M-type current in freshly isolated monkey RPE cells, indicating that KCNQ subunits contribute to the underlying channels. RT-PCR analysis revealed the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in the RPE and all five KCNQ transcripts in the neural retina. At the protein level, KCNQ5 was detected in the RPE, whereas both KCNQ4 and KCNQ5 were found in neural retina. In situ hybridization in frozen monkey retinal sections revealed KCNQ5 gene expression in the ganglion cell layer and the inner and outer nuclear layers of the neural retina, but results in the RPE were inconclusive due to the presence of melanin. Immunohistochemistry revealed KCNQ5 in the inner and outer plexiform layers, in cone and rod photoreceptor inner segments, and near the basal membrane of the RPE. The data suggest that KCNQ5 channels contribute to the RPE basal membrane K+ conductance and, thus, likely play an important role in active K+ absorption. The distribution of KCNQ5 in neural retina suggests that these channels may function in the shaping of the photoresponses of cone and rod photoreceptors and the processing of visual information by retinal neurons.

The inwardly rectifying K+ (Kir) current in mammalian retinal pigment epithelial (RPE) cells, which is largely mediated by Kir7.1 channels, is stable in cells dialyzed with MgATP but runs down when intracellular ATP is depleted. A potential mechanism for this rundown is a decrease in phosphatidylinositol 4,5-bisphosphate (PIP2) regeneration by ATP-dependent lipid kinases. Here, we used the whole cell voltage-clamp technique to investigate the membrane PIP2 dependence of Kir channels in isolated bovine RPE cells. When RPE cells were dialyzed with ATP-free solution containing PIP2 (25–50 μM), rundown persisted but was markedly reduced. Removal of Mg2+ from the pipette solution also slowed rundown, indicating that elevated intracellular Mg2+ concentration contributes to rundown. Cell dialysis with the PIP2 scavenger neomycin in MgATP solution diminished Kir current in a voltage-dependent manner, suggesting that it acted at least in part by blocking the Kir channel. Kir current in MgATP-loaded cells was partially inhibited by bath application of quercetin (100 μM), phenylarsine oxide (100 μM), or wortmannin (50 μM), inhibitors of phosphatidylinositol (PI) kinases, and was completely inhibited by cell dialysis with 2 mM adenosine, a PI4 kinase inhibitor. Both LY-294002 (100 μM), an inhibitor of PI3 kinases, and its inactive analog LY-303511 (100 μM) rapidly and reversibly inhibited Kir current, suggesting that these compounds act as direct channel blockers. We conclude that the activity of Kir channels in the RPE is critically dependent on the regeneration of membrane PIP2 by PI4 kinases and that this may explain the dependence of these channels on hydrolyzable ATP.

Previously, we demonstrated that the inwardly rectifying K+ (Kir) channel subunit Kir7.1 is highly expressed in bovine and human retinal pigment epithelium (RPE). The purpose of this study was to determine whether any of the 14 other members of the Kir gene family are expressed in native human RPE. Conventional reverse transcription-polymerase chain reaction (RT-PCR) analysis indicated that in addition to Kir7.1, seven other Kir channel subunits (Kir1.1, Kir2.1, Kir2.2, Kir3.1, Kir3.4, Kir4.2 and Kir6.1) are expressed in the RPE, whereas in neural retina, all 14 of the Kir channel subunits examined are expressed. The identities of RT-PCR products in the RPE were confirmed by DNA sequencing. Real-time RT-PCR analysis showed, however, that transcripts of these channels are significantly less abundant than Kir7.1 in the RPE. Western blot analysis of the Kir channel subunits detected in the RPE by RT-PCR revealed the expression of Kir2.1, Kir3.1, Kir3.4, Kir4.2, Kir6.1, and possibly Kir2.2, but not Kir1.1, in both human RPE and neural retina. Our results indicate that human RPE expresses at least five other Kir channel subtypes in addition to Kir7.1, suggesting that multiple members of the Kir channel family may function in this epithelium.

PURPOSE. Mutations in the membrane frizzled-related protein (MFRP) gene cause nanophthalmos in humans, and a splice site mutation causes recessive retinal degeneration in the rd6 mouse. In human and mouse genomes, the MFRP gene lies adjoining to the complement 1q tumor necrosis factor-related protein 5 (CTRP5/C1QTNF5) gene involved in causing retinal degeneration and abnormal lens zonules in human. The purpose of this study was to characterize the spatial and temporal expression of the mouse Mfrp gene, determine tissue and subcellular localization of MFRP protein, and study its interaction with CTRP5. METHODS. Expression of the Mfrp gene in the mouse was studied by quantitative (q)RT-PCR. MFRP protein expression and distribution were studied by Western blot analysis, immunohistochemistry, and immunoelectron microscopy. Interaction with CTRP5 was studied by immunoprecipitation and immunoblot analysis, using mouse eye and human retinal pigmented epithelium (RPE) choroid extracts and by expressing full-length CTRP5 and MFRP in a heterologous system. RESULTS. The Mfrp gene is specifically expressed in RPE and ciliary body (CB), and its expression starts during early stages of embryogenesis. In the albino mouse eye, MFRP is localized to the apical and basal membranes of RPE and ciliary epithelium (CE). In addition, MFRP and CTRP5 were found to colocalize in RPE, CE, and MDCK cells, a general model of polarized epithelia. These proteins interact with each other in ocular tissues and also in a heterologous system. CONCLUSIONS. MFRP is localized to the plasma membrane of CE and RPE, and colocalizes and interacts with CTRP5 indicating a functional relationship between these two proteins.

The primary objective of this study was to compare the intraocular pressure (IOP)-lowering efficacy of travoprost 0.004%/timolol 0.5% fixed combination to the concomitant administration of travoprost 0.004% (TRAVATAN) and timolol 0.5% in subjects with open angle glaucoma or ocular hypertension.This was a randomized, multicenter, double-masked, active-controlled, parallel group study. Three hundred sixteen patients with open angle glaucoma or ocular hypertension were randomly assigned to travoprost 0.004%/timolol 0.5% ophthalmic solution fixed combination once daily in the morning or concomitant administration of timolol 0.5% once daily in the morning and travoprost 0.004% ophthalmic solution once daily in the evening. The efficacy and safety of the fixed combination were compared with concomitant therapy over three months. The primary efficacy outcome measure was mean intraocular pressure.Both travoprost 0.004%/timolol 0.5% fixed combination and the concomitant administration of travoprost 0.004% and timolol 0.5% produced statistically significant reductions from baseline in IOP, with mean IOP ranging from 15.2 to 16.5 mm Hg in the patients using travoprost 0.004%/timolol 0.5% fixed combination compared with 14.7 to 16.1 mm Hg in the concomitant group. The upper 95.1% confidence limit for the differences in mean IOP (fixed combination minus concomitant) wasor =1.5 mm Hg at 7 of 9 visits, including all three 8 AM time points, 24-hours post-dose. Mean IOP reductions from baseline ranged from 7.4 to 9.4 mm Hg in the fixed combination group compared with 8.4 to 9.4 mm Hg with concomitant therapy. Safety analysis demonstrated equivalent safety between the two treatment groups.A fixed combination of travoprost 0.004% and timolol 0.5% produced clinically relevant IOP reductions in patients with open angle glaucoma or ocular hypertension that were comparable to concomitant therapy with its components. Safety and tolerability of the fixed combination were also equivalent to concomitant therapy. Travoprost 0.004%/timolol 0.5% fixed combination offers IOP reduction equivalent to concomitant therapy, with potential benefits that include convenience (fewer bottles and drops per day), improved compliance, cost savings (based on fewer co-payments), and elimination of potential washout effects.

To investigate the effect of the timing of the scleral flap suture release on the safety and long-term success of phacotrabeculectomy.Retrospective, nonrandomized chart review.We studied 173 eyes of 173 consecutive patients who underwent primary phacotrabeculectomy, with or without intraoperative mitomycin-C, and subsequent scleral flap suture release.Early suture release (3 weeks) had a lower long-term success rate than late suture release (3 weeks), with an average follow-up period of 44.8 months. Patients who used mitomycin-C had statistically later suture release than did patients without mitomycin-C. There were nine cases of hypotony, for which independent risk factors were early suture release and the use of mitomycin-C.Greater long-term success with minimal complications was achieved when scleral flap suture release was performed three weeks after phacotrabeculectomy. Suture release within three weeks after surgery was associated with an increased risk of filtration failure and a higher risk of postoperative complications.

To compare the intraocular pressure (IOP) reduction between dorzolamide 2% and brimonidine 0.2% in primary open-angle glaucoma (POAG) or ocular hypertension (OHT).This study was a prospective, double-masked, randomized, crossover comparison of dorzolamide 2% (Trusopt) and brimonidine 0.2% (Alphagan), three times daily during two six-week study periods. The primary endpoint was mean change from baseline in trough IOP and secondary endpoints were mean change from baseline in IOP one and three hours after dosing. T-tests and a repeated-measures ANOVA were used to statistically evaluate the data.Of 43 patients enrolled, 41 completed the first treatment and 38 completed both treatments. Baseline IOP for dorzolamide was 24.3 mm Hg and brimonidine, 24.6 mm Hg (P = 0.9). Mean IOP reduction at trough was similar for both agents, 3.0 mm Hg (P = 0.96). Reductions at one and three hours were comparable (P = ns). Both agents were well tolerated with adverse events consistent with the package inserts. Dorzolamide was associated with more frequent stinging (P = 0.017) and burning (P0.001), while brimonidine was associated with more frequent dry eye (P = 0.04).Dorzolamide and brimonidine, as monotherapy, produced equivalent IOP-lowering efficacy at trough and at one and three hours after instillation, and both were well tolerated.

To determine the expression and functional activity of proton-coupled oligopeptide transporters (POT) in retinal pigment epithelial (RPE) cells.RT-PCR was used to probe the presence of POT mRNA in freshly isolated bovine RPE (BRPE) and human RPE (HRPE) cells, a human RPE cell line (ARPE-19), and human and bovine neural retina. [14C]GlySar uptake was used to characterize POT activity in cultured ARPE-19 cells and freshly isolated BRPE cell sheet suspensions.PHT1 mRNA was expressed in BRPE, HRPE, ARPE-19, and bovine and human neural retina. In contrast, PEPT2 and PHT2 were expressed only in bovine and human retina, and PEPT1 could not be detected. GlySar exhibited a linear uptake over 6 h at pH values of 6.0 and 7.4, with greater uptake at pH 7.4 (p0.01). GlySar uptake did not exhibit saturability (5-2000 microM) and was unchanged when studied in the presence of 1 mM L-histidine. In contrast, GlySar uptake was significantly decreased when studied at 4 degrees C or in the presence of endocytic inhibitors at 37 degrees C (p0.01). Studies in BRPE cell sheet suspensions validated the results obtained in ARPE-19 cells and strongly suggested the absence of POT on the apical and basolateral membranes of RPE.PHT1 mRNA is present in native bovine and human RPE and a human RPE cell line. However, the data argue against PHT1 being expressed on plasma membranes of RPE. Overall, GlySar appears to be taken up by RPE cells via a low-affinity, endocytic process.

PURPOSE The purpose of this study was to identify the molecular basis of the apical membrane K(+) conductance in native bovine retinal pigment epithelium (RPE). METHODS RT-PCR, Northern blot, and Western blot analyses were used to detect the expression of the inwardly rectifying K(+) (Kir) channel subunits Kir7.1 and Kir4.1 in native bovine RPE and neural retina. The distribution of Kir7.1 protein was determined in frozen sections of bovine retina-RPE-choroid by indirect immunofluorescence analysis. RESULTS RT-PCR analysis revealed Kir7.1 transcript in both RPE and neural retina, but Kir4.1 transcript only in the neural retina. In Northern blot analysis, Kir4.1 probe hybridized to an appropriately sized-transcript in neural retina but not in RPE. Kir7.1 probe hybridized to a major transcript of approximately 1.5 kb in both RPE and neural retina, but with greater expression in RPE. In Western blot analysis, Kir7.1 antibody recognized a major monomer of approximately 53 kDa in RPE, whereas Kir4.1 antibody recognized a monomer of approximately 60 kDa in neural retina but not in RPE. Intense Kir7.1 immunolabeling was present on the apical surface of all RPE cells and appeared to extend over the length of the apical processes. Na(+),K(+)-ATPase expression varied among RPE cells, but in highly expressing cells, it colocalized with Kir7.1. CONCLUSIONS These results indicate that the Kir7.1 channel subunit, but not Kir4.1, is a major component of the apical K(+) conductance in bovine RPE. Kir7.1 is distributed over the length of apical processes, where it probably functions in the regulation of K(+) transport and the electrical response of the RPE to light-evoked changes in subretinal K(+) concentration.

Purpose To compare the long-term success rate of selective laser trabeculoplasty (SLT) versus argon laser trabeculoplasty (ALT). Design Retrospective chart review. Participants One hundred ninety-five eyes of 195 patients with uncontrolled open-angle glaucoma (OAG), of which 154 eyes underwent ALT and 41 eyes underwent SLT and were followed up for a maximum of 5 years. Intervention The SLT patients were treated with the frequency-doubled q-switched neodymium:yytrium–aluminum–garnet laser (532 nm). Approximately 50 to 55 nonoverlapping spots were placed over 180° of the trabecular meshwork at energy levels ranging from 0.6 to 1.0 mJ per pulse. The ALT patients were treated with the argon blue–green laser with between 45 to 55 adjacent, nonoverlapping spots over 180° of the trabecular meshwork at 470 to 1150 mW of energy per pulse. Main outcome measures The success rates were defined by criterion I and criterion II. Success by criterion I was defined as a decrease in intraocular pressure (IOP) of 3 mmHg or more with no additional medications, laser, or glaucoma surgery. Criterion II had the same requirements as criterion I, except that a 20% or more IOP reduction was required for success. Results The mean follow-up time was 37.4±14.7 months for patients in the SLT group and 33.6±17.0 months for patients in the ALT group. The long-term success rate was not significantly different between the ALT and SLT groups by either criterion (Kaplan-Meier survival analysis log-rank P = 0.20 by criterion I and P = 0.12 by criterion II). When comparing patients with and without previous ALT, there was not a statistically significant difference in the patients treated with SLT by either criterion (log-rank P = 0.37 by criterion I and P = 0.39 by criterion II). Conclusions In eyes with primary OAG that are receiving maximally tolerated medical therapy, SLT was found to be as effective as ALT in lowering IOP over a 5-year period. However, long-term data reveal that many of the glaucoma patients treated with SLT and ALT required further medical or surgical intervention. Whether SLT has better long-term success than ALT in repeat laser trabeculoplasty treatments remains unclear.

Inwardly rectifying K+ (Kir) channels in the apical membrane of the retinal pigment epithelium (RPE) play a key role in the transport of K+ into and out of the subretinal space (SRS), a small extracellular compartment surrounding photoreceptor outer segments. Recent molecular and functional evidence indicates that these channels comprise Kir7.1 channel subunits. The purpose of this study was to determine whether Kir channels in the RPE are modulated by extracellular (pHo) or intracellular pH (pHi), both of which change upon illumination of the dark-adapted retina. The Kir current (IKir) in acutely dissociated bovine RPE cells was recorded in the whole-cell configuration while altering pHo or pHi. In cells dialysed with pipette solution buffered to pH 7.2, step changes in pHo from 7.4 to 8.0, 7.0 or 6.5 had little effect on IKir. Acidification to pHo 6.0, however, caused a transient activation of IKir followed by a slower inhibition. To determine the dependence of IKir on pHi, we altered pHi within individual RPE cells at constant pHo by imposing transmembrane acetate concentration gradients. These experiments revealed a biphasic relationship between IKir and pHi: IKir was maximal at about pHi 7.1, but decreased sharply at more acidic or alkaline levels. To evaluate the role of Kir7.1 channels in the pHi-dependent changes in IKir, we tested the effect of transmembrane acetate concentration gradients on Rb+ currents, which are 10-fold larger than K+ currents for this channel subtype. Inwardly rectifying Rb+ currents were maximal at about pHi 7.0 and were inhibited by intracellular alkalinization or acidification. We conclude that the Kir conductance in the RPE is modulated by intracellular pH in the physiological range and that this reflects the behaviour of Kir7.1 channels. This sensitivity to pHi may provide an important mechanism linking photoreceptor activity and RPE function.