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DNA origami nanostructures (DOs) are promising tools for applications including drug delivery, biosensing, detecting biomolecules, and probing chromatin substructures. Targeting these nanodevices to mammalian cell nuclei could provide impactful approaches for probing, visualizing, and controlling biomolecular processes within live cells. We present an approach to deliver DOs into live-cell nuclei. We show that these DOs do not undergo detectable structural degradation in cell culture media or cell extracts for 24 hours. To deliver DOs into the nuclei of human U2OS cells, we conjugated 30-nanometer DO nanorods with an antibody raised against a nuclear factor, specifically the largest subunit of RNA polymerase II (Pol II). We find that DOs remain structurally intact in cells for 24 hours, including inside the nucleus. We demonstrate that electroporated anti-Pol II antibody-conjugated DOs are piggybacked into nuclei and exhibit subdiffusive motion inside the nucleus. Our results establish interfacing DOs with a nuclear factor as an effective method to deliver nanodevices into live-cell nuclei.

Background: Multiple pregnancy is associated with higher risk of uterine atony, postpartum hemorrhage (PPH), blood transfusion, hysterectomy, and death. The optimal dose of oxytocin at cesarean delivery in people with twin pregnancy is unknown. We sought to determine the effective bolus dose of oxytocin required to initiate adequate uterine tone in 90% of people (ED90) with twin pregnancy undergoing elective cesarean delivery. Our hypothesis was that the dose of oxytocin would be higher than 0.5 international units (IU) but lower than 5 IU., Methods: A double-blind dose-finding study using the biased coin up-down method was undertaken in people with twin pregnancy ≥36 weeks gestational age undergoing elective cesarean delivery under neuraxial anesthesia. Those with additional risk factors for PPH, apart from twin pregnancy, were excluded. Oxytocin was administered as an intravenous bolus over 1 minute on delivery of the second fetus. The first patient received 0.5 IU, and subsequent oxytocin doses were administered according to a sequential allocation scheme. The actual doses administered were 0.5, 1, 2, 3, 4, and 5 IU of oxytocin. The primary outcome was the response defined as the satisfactory uterine tone at 2 minutes after completion of administration of the oxytocin bolus, as assessed by the operating obstetrician. Secondary outcomes included need for rescue uterotonic drugs, adverse effects, and estimated blood loss. The ED90 was estimated using the Dixon-Mood and the isotonic regression methods., Results: Thirty patients were included in study. The estimated ED90 of oxytocin was 4.38 IU (95% confidence interval [CI], 3.68-4.86 IU) and 3.41 IU (95% CI, 2.83-3.98 IU) by the isotonic regression and Dixon-Mood methods, respectively. Seven patients had inadequate tone at the 2-minute evaluation point and required rescue uterotonic drugs. The median (interquartile range [IQR]) estimated blood loss was 1031 mL (732-1462 mL) calculated by the change in 24-hour hematocrit. Incidence of hypotension after oxytocin administration was 27%, nausea 30%, and vomiting 17%., Conclusions: Our results demonstrated that people with twin pregnancy require a much higher dose of oxytocin than those with singleton pregnancies. We recommended people with twin pregnancies should receive an initial 5 IU bolus over at least 1 minute when undergoing elective cesarean delivery under neuraxial anesthesia., Competing Interests: Conflicts of Interest: See Disclosures at the end of the article., (Copyright © 2022 International Anesthesia Research Society.)

DNA origami (DO) are promising tools for in vitro or in vivo applications including drug delivery; biosensing, detecting biomolecules; and probing chromatin sub-structures. Targeting these nanodevices to mammalian cell nuclei could provide impactful approaches for probing visualizing and controlling important biological processes in live cells. Here we present an approach to deliver DO strucures into live cell nuclei. We show that labelled DOs do not undergo detectable structural degradation in cell culture media or human cell extracts for 24 hr. To deliver DO platforms into the nuclei of human U2OS cells, we conjugated 30 nm long DO nanorods with an antibody raised against the largest subunit of RNA Polymerase II (Pol II), a key enzyme involved in gene transcription. We find that DOs remain structurally intact in cells for 24hr, including within the nucleus. Using fluorescence microscopy we demonstrate that the electroporated anti-Pol II antibody conjugated DOs are efficiently piggybacked into nuclei and exihibit sub-diffusive motion inside the nucleus. Our results reveal that functionalizing DOs with an antibody raised against a nuclear factor is a highly effective method for the delivery of nanodevices into live cell nuclei., Competing Interests: Declaration of Competing Interest The authors declare no competing interests.

Unilateral congenital cataracts present multiple barriers in the development of vision and stereoacuity despite the improved visual optics that early surgery, contact lenses and intraocular lenses (IOL) have provided. With better understanding of the latent period (the timeframe in which the abnormal event has no long-term effect on visual development in the deprived eye) and the critical periods (the age range during which developing brains can be altered in a profound and permanent way by abnormal experience) for stereoacuity and amblyopia we can focus our treatment methods to not only improve vision but also develop binocularity. Fifty years ago, it was believed that it was almost impossible for an eye with a unilateral congenital cataract to achieve good visual acuity. Twenty-five years ago, we believed that it was almost impossible for an eye with a unilateral cataract to achieve stereoacuity. It is time to expand our belief that the best that we can do with the eye in unilateral congenital cataract is to create a spare.

Mouse models of CLN3 Batten disease, a rare lysosomal storage disorder with no cure, have improved our understanding of CLN3 biology and therapeutics through their ease of use and a consistent display of cellular pathology. However, the translatability of murine models is limited by disparities in anatomy, body size, life span and inconsistent subtle behavior deficits that can be difficult to detect in CLN3 mutant mouse models, thereby limiting their use in preclinical studies. Here, we present a longitudinal characterization of a novel miniswine model of CLN3 disease that recapitulates the most common human pathogenic variant, an exon 7-8 deletion (CLN3Δex7/8). Progressive pathology and neuron loss is observed in various regions of the CLN3Δex7/8 miniswine brain and retina. Additionally, mutant miniswine present with retinal degeneration and motor abnormalities, similar to deficits seen in humans diagnosed with the disease. Taken together, the CLN3Δex7/8 miniswine model shows consistent and progressive Batten disease pathology, and behavioral impairment mirroring clinical presentation, demonstrating its value in studying the role of CLN3 and safety/efficacy of novel disease-modifying therapeutics., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Background: Dystroglycanopathies are a heterogeneous group of membrane-related muscular dystrophies. The dystroglycanopathy phenotype includes a spectrum of severity ranging from severe congenital muscular dystrophy to adult-onset limb-girdle muscular dystrophy (LGMD). LGMDR9 is a dystroglycanopathy caused by mutations in the FKRP gene. Previous studies have characterized electroretinogram findings of dystroglycanopathy mouse models but have not been reported in humans., Purpose: This study set out to characterize the electroretinogram in eight participants with LGMDR9., Methods: Eight participants were recruited from an ongoing dystroglycanopathy natural history study at the University of Iowa (NCT00313677). Inclusion criteria for the current study were children and adults > 6 years old with confirmed LGMDR9. Age similar controls were identified from our electrophysiology service normative control database. Full-field electroretinograms were recorded using ISCEV standards. Six of the eight participants underwent light-adapted ON/OFF testing., Results: The electronegative electroretinogram was not seen in any participants with LGMDR9. An unusual sawtooth pattern in the 30 Hz flicker with faster rise than descent was noted in all 8 participants. Our cases showed a decreased b-wave amplitude in light-adapted ON responses (p = 0.011) and decreased d-wave amplitude in light-adapted OFF responses (p = 0.015). Decreased b-wave amplitude in light-adapted 3.0 testing (p = 0.015) and decreased flicker ERG amplitudes were also detected (p = 0.0018). Additionally, compared to controls, participants with LGMDR9 had decreased a-wave amplitudes on dark-adapted 10 testing (p = 0.026)., Conclusions: Abnormal ON/OFF bipolar cell responses and sawtooth 30 Hz flicker waveforms on full-field electroretinogram may be specific for LGMDR9. If confirmed in a larger population and if related to disease stage, these tests are potential biomarkers which could be useful as endpoints in clinical treatment trials., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Molecular dynamics simulations are often used to provide feedback in the design workflow of DNA nanostructures. However, even with coarse-grained models, the convergence of distributions from unbiased simulation is slow, limiting applications to equilibrium structural properties. Given the increasing interest in dynamic, reconfigurable, and deformable devices, methods that enable efficient quantification of large ranges of motion, conformational transitions, and mechanical deformation are critically needed. Metadynamics is an automated biasing technique that enables the rapid acquisition of molecular conformational distributions by flattening free energy landscapes. Here we leveraged this approach to sample the free energy landscapes of DNA nanostructures whose unbiased dynamics are nonergodic, including bistable Holliday junctions and part of a bistable DNA origami structure. Taking a DNA origami-compliant joint as a case study, we further demonstrate that metadynamics can predict the mechanical response of a full DNA origami device to an applied force, showing good agreement with experiments. Our results exemplify the efficient computation of free energy landscapes and force response in DNA nanodevices, which could be applied for rapid feedback in iterative design workflows and generally facilitate the integration of simulation and experiments. Metadynamics will be particularly useful to guide the design of dynamic devices for nanorobotics, biosensing, or nanomanufacturing applications.

Purpose: The purpose of this study was to compare the natural history of visual function change in cohorts of patients affected with retinal degeneration due to biallelic variants in Bardet-Biedl syndrome genes: BBS1 and BBS10., Methods: Patients were recruited from nine academic centers from six countries (Belgium, Canada, France, New Zealand, Switzerland, and the United States). Inclusion criteria were: (1) female or male patients with a clinical diagnosis of retinal dystrophy, (2) biallelic disease-causing variants in BBS1 or BBS10, and (3) measures of visual function for at least one visit. Retrospective data collected included genotypes, age, onset of symptoms, and best corrected visual acuity (VA). When possible, data on refractive error, fundus images and autofluorescence (FAF), optical coherence tomography (OCT), Goldmann kinetic perimetry (VF), electroretinography (ERG), and the systemic phenotype were collected., Results: Sixty-seven individuals had variants in BBS1 (n = 38; 20 female patients and 18 male patients); or BBS10 (n = 29; 14 female patients and 15 male patients). Missense variants were the most common type of variants for patients with BBS1, whereas frameshift variants were most common for BBS10. When ERGs were recordable, rod-cone dystrophy (RCD) was observed in 82% (23/28) of patients with BBS1 and 73% (8/11) of patients with BBS10; cone-rod dystrophy (CORD) was seen in 18% of patients with BBS1 only, and cone dystrophy (COD) was only seen in 3 patients with BBS10 (27%). ERGs were nondetectable earlier in patients with BBS10 than in patients with BBS1. Similarly, VA and VF declined more rapidly in patients with BBS10 compared to patients with BBS1., Conclusions: Retinal degeneration appears earlier and is more severe in BBS10 cases as compared to those with BBS1 variants. The course of change of visual function appears to relate to genetic subtypes of BBS.

Background: Aniridia patients have poor visual acuity and iris malformation. The fovea in these patients is underdeveloped, but the relationship between structure and electrophysiologic function remains incompletely understood. This study correlates electrophysiology, visual acuity and optical coherence tomography (OCT) in patients with aniridia secondary to mutations in the PAX6 gene and compares with age-similar controls., Methods: Patients were recruited from clinical practice. The mfERG protocol was a 4-min 103-hexagon protocol covering approximately 40° in diameter of central retina. Diagnosys full-field ERG (ffERG) and VERIS multifocal ERG (mfERG) were obtained using standard International Society for Clinical Electrophysiology of Vision protocols. OCT central thickness was recorded, and an OCT foveal score was calculated. Nonparametric permutation testing was utilized to determine the statistical significance., Results: A total of 6 aniridia patients and 25 control patients were recruited. On mfERG, aniridia patients had significantly lower amplitudes in rings 1-3 (p = 0.0006, 0.0013, 0.0132), shorter latencies in ring 1 (p = 0.0312) and longer latencies in rings 5 and 6 (p = 0.0026, p = 0.0042) than controls. There was a significantly positive relationship in aniridia patients between logMAR visual acuity and mfERG amplitude in ring 4 (p = 0.0392) and ring 5 (p = 0.0489). On ffERG, there was no difference in amplitudes, though photopic 3.0 a- and b-wave latency, 30 Hz flicker latency and scotopic 0.01 b-wave latency were significantly longer in aniridia patients versus control (p = 0.0018, 0.0.0005, 1.00 x [Formula: see text], 0.0198). Thicker central macula on OCT correlated with lower mfERG amplitudes in rings 4-6 (p = 0.0369, 0.0292, 0.0255). There was no correlation between visual acuity and central macular thickness or foveal hypoplasia score as determined by OCT., Conclusions: Higher amplitude on mfERG correlated with poorer visual acuity in rings 4 and 5 in patients with PAX6 mutations. The slope of the change in amplitude from central to peripheral rings on the mfERG is significantly different in aniridia patients compared to controls, with a slower drop-off of amplitude from center to periphery. Additionally, mfERG in aniridia showed lower amplitudes than controls in rings 1-3. These changes along with the lack of correlation between visual acuity and central macular thickness/OCT score suggest that changes in electrical topography may be important to visual deficits in patients with PAX6 gene mutations., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Purpose: To correlate clinical features, molecular genetic findings, and visual acuity in a cohort of patients clinically diagnosed with oculocutaneous albinism. Design: Retrospective chart review Methods: 58 charts met the inclusion criteria. Clinical examination, ancillary testing, and molecular genetic diagnoses were extracted. A novel clinical albinism score (CAS) was developed. Results: A least one likely pathogenic mutation was found in 44/58 (75.9%) patients. Mutations in the OCA1 gene were the most common (52.3%), followed by OCA2 (34%), OCA4 (2.3%), OA1 (6.8%), and HPS (4.5%). Thirty-four percentage of patients had a complete genotype, 41% had one mutation found and 24% had negative genetic testing. CAS was statistically significantly higher in patients with complete genotype, versus patients with one or no mutations found ( p < .01). Better visual acuity was associated with lower CAS and fewer disease-causing mutations ( p < .01). Foveal defects and iris transillumination were associated with a higher number of mutations ( p < .01). Patients with nystagmus or anomalous optic nerves had worse visual acuity than those who did not ( p < .01, p < .05). Conclusions: Patients with a complete genotype were more likely to have higher CAS. Vision loss correlated with complete phenotype and higher CAS, the presence of nystagmus and anomalous optic nerves. Patients with features of albinism in whom an incomplete genotype was found had better vision than those with complete genotype, suggesting a mild occult mutation or modifier variant. Genetic diagnosis is vital for complete diagnosis, counseling, and family planning.

Manipulation of temperature can be used to actuate DNA origami nano-hinges containing gold nanoparticles. We develop a physical model of this system that uses partition function analysis of the interaction between the nano-hinge and nanoparticle to predict the probability that the nano-hinge is open at a given temperature. The model agrees well with experimental data and predicts experimental conditions that allow the actuation temperature of the nano-hinge to be tuned over a range of temperatures from 30 °C to 45 °C. Additionally, the model identifies microscopic interactions that are important to the macroscopic behavior of the system, revealing surprising features of the system. This combination of physical insight and predictive potential is likely to inform future designs that integrate nanoparticles into dynamic DNA origami structures or use strand binding interactions to control dynamic DNA origami behavior. Furthermore, our modeling approach could be expanded to consider the incorporation, stability, and actuation of other types of functional elements or actuation mechanisms integrated into nucleic acid devices.

Background: Albinism patients have poor visual acuity in addition to hypopigmentation. Their foveal anatomy is abnormal, but correlation with function is incompletely understood. This study correlates retinal electrophysiology, visual acuity and optical coherence tomography (OCT) anatomy in albinism patients and compares with age-similar controls., Methods: Institutional Review Board approval was obtained (IRB# 201408782). Patients were recruited from clinical practice. Inclusion criteria were at least three clinical features of albinism including iris transillumination, nystagmus, fundus hypopigmentation, or foveal hypoplasia on OCT and/or molecular genetic confirmation. Diagnosys (Lowell, Mass) full-field ERG (ffERG) and VERIS multifocal ERG (mfERG; Electro-Diagnostic Imaging, Milpitas, California) were obtained using standard International Society for Clinical Electrophysiology of Vision protocols. The mfERG protocol was a 4-min 103-hexagon protocol covering approximately 40° in diameter of central retina. Control subjects without albinism were recruited by in-hospital notices and invitations in clinic. OCT central thickness was recorded, and an OCT foveal score was calculated. Nonparametric permutation testing was utilized to determine the statistical significance., Results: A total of 16 albinism patients and 19 age-similar controls were recruited. Four of 16 albinism patients had no nystagmus. Seventeen non-albinism controls had no ocular disorder other than refractive error. Two controls had infantile nystagmus with normal maculas on OCT. There was no statistically significant difference in mfERG amplitude or latency between albinism patients with or without nystagmus (lowest p = 0.68; 0.54, respectively). mfERG: 12 of 16 (75%) albinism patients had average ring 1 amplitudes within one standard deviation of controls despite having abnormal foveal anatomy on OCT. Patients averaged shorter latencies in rings 1 and 2 than controls (p = 0.005, p = 0.02). Patients averaged higher amplitudes than controls in rings 4, 5 and 6 (p = 0.03, p = 0.006, p = 0.004). There was no significant correlation between visual acuity and mfERG amplitudes in any ring (smallest p = 0.15). ffERG: Patients averaged higher amplitudes on 30 Hz flicker (p = 0.008). In all conditions, albinism patients had higher amplitude a-waves (p ≤ 0.03). B-waves were higher amplitude than controls in light-adapted 3.0 (p = 0.03). There was no statistical correlation between ffERG amplitudes and visual acuity (smallest p = 0.45). OCT: In albinism patients, thicker central macula on OCT correlated with lower mfERG amplitudes in all rings except for ring 1 (p < 0.05) and lower ffERG a-wave amplitudes on dark-adapted 0.01 (p = 0.003). Macular thickness on OCT did not correlate with visual acuity (p = 0.51); OCT foveal score did (p = 0.0004)., Conclusions: Amplitude of mfERG does not correlate with visual acuity in any ring in patients with albinism. The slope of the change in amplitude from central to peripheral rings on the mferg is significantly different in albinism patients versus controls whether or not nystagmus is present. The decreased slope of change in amplitudes from center to periphery of the macula in albinism patients indicates changes in macular topography that are more important to visual deficits than the foveal depression.

Background : Inherited retinal dystrophies are a leading cause of irreversible blindness in children in the United States. Topical carbonic anhydrase inhibitors have improved central vision and cystoid macular edema in patients with retinal dystrophies, but few studies have assessed their efficacy in children. Materials and Methods : A retrospective chart review was performed with Institutional Review Board approval to identify pediatric patients with inherited retinal dystrophies who received topical brinzolamide at a single university center between 2008 and 2015. Serial visual acuity and central macular thicknesses were compared to assess the efficacy of brinzolamide. Results : Seven subjects were identified who met the inclusion criteria. Four had juvenile X-linked retinoschisis, two had retinitis pigmentosa, and one had Leber congenital amaurosis. All were prescribed brinzolamide thrice daily; however, one patient was completely non-compliant. Four of the six treated patients exhibited a mild decrease in central macular thickness in both eyes during the study with all six treated patients having significantly improved vision at the first endpoint, 33.2 ± 8.2 months after treatment initiation. For treated patients, average visual acuity (LogMAR) ± standard error of the mean improved from 0.5 ± 0.04 pre-treatment to 0.3 ± 0.1 at the second endpoint, 50.2 ± 7.3 months after treatment initiation. Conclusions : Mild anatomic improvement of macular cysts was seen in pediatric patients using brinzolamide. Visual acuity improvement occurred even without significant reduction in macular cysts. Further studies are needed to determine whether the beneficial effects of carbonic anhydrase inhibitors are sustained in children with inherited retinal degenerations.

The self-assembly of a DNA origami structure, although mostly feasible, represents indeed a rather complex folding problem. Entropy-driven folding and nucleation seeds formation may provide possible solutions; however, until now, a unified view of the energetic factors in play is missing. Here, by analyzing the self-assembly of origami domains with identical structure but different nucleobase composition, in function of variable design and experimental parameters, we identify the role played by sequence-dependent forces at the edges of the structure, where topological constraint is higher. Our data show that the degree of mechanical stress experienced by these regions during initial folding reshapes the energy landscape profile, defining the ratio between two possible global conformations. We thus propose a dynamic model of DNA origami assembly that relies on the capability of the system to escape high structural frustration at nucleation sites, eventually resulting in the emergence of a more favorable but previously hidden state.

Many challenges in biosensing originate from the fact that the all-important nanoarchitecture of the biosensor surface, including precise density and orientation of bioreceptors, is not entirely comprehended. Here, we introduced a three-dimensional DNA origami as a bioreceptor carrier to functionalize the fiber optic surface plasmon resonance (FO-SPR) sensor with nanoscale precision. Starting from a 24-helix bundle, two distinct DNA origami structures were designed to position thrombin-specific aptamers with different densities and distances (27 and 113 nm) from the FO-SPR surface. The origami-based biosensors not only proved to be capable of reproducible, label-free thrombin detection but revealed also valuable innovative features: (1) a significantly better performance in the absence of backfilling, known as essential in the biosensing field, suggesting improved bioreceptor orientation and accessibility, and (2) a wider linear range compared to previously reported thrombin biosensors. We envisage that our method will be beneficial for both scientists and clinicians looking for new surface (bio)chemistry and improved diagnostics.

Natural filaments, such as microtubules and actin filaments, are fundamental components of the cell. Despite their relatively simple linear structure, filaments play a number of crucial roles in living organisms, from scaffolding to cellular adhesion and motility. The mechanical properties of natural filaments mostly rely on the structural features of the component units and on the way they are connected together, thus providing an ideal molecular model for emulation purposes. In this review, we describe the progresses done in this field using DNA for the rational design of synthetic filamentous-like materials with tailored structural and physical characteristics. We firstly survey the strategies that have been adopted until now for the construction of individual DNA building components and their programmable self-assembly into linear oligomeric structures. We then describe the theoretical models of polymer elasticity applied to calculate the bending strength of DNA filaments, expressed in terms of persistence length. Finally, we report some of the most exciting examples of truly biomimetic DNA filaments, which are capable of mimicking not only the sophisticated structural features of their natural counterparts but also their responsiveness to external stimuli, thus resulting in active motion and growing networks between distant loci.

Importance: Congenital stationary night blindness (CSNB) implies a stable condition, with the major symptom being nyctalopia present at birth. Pediatric clinical presentation and the course of different genetic subtypes of CSNB have not, to our knowledge, been well described in the era of molecular genetic diagnosis., Objective: To describe the presentation and longitudinal clinical characteristics of pediatric patients with molecularly confirmed TRPM1-associated complete CSNB (cCSNB)., Design, Setting, Participants: This study was conducted at the University of Iowa from January 1, 1990, to July 1, 2015, and was a retrospective, longitudinal case series of 7 children (5 [71.4%] female) with TRPM1-associated cCSNB followed up for a mean (SD) of 11.1 (2.8) years., Main Outcomes and Measures: History, ophthalmologic examination findings, full-field electroretinogram (ffERG) results, full-field stimulus threshold testing results, Goldmann visual field results, optical coherence tomography results, and molecular genetic results were evaluated. Presenting symptoms and signs, the correlation of refractive error with electroretinography, and clinical evolution were analyzed., Results: Seven patients (5 [71.4%] female) presented early in childhood with strabismus (n = 6 [86%]), myopia (n = 5 [71%]), and/or nystagmus (n = 3 [43%]). The mean (SD) age at presentation was 8 (4) months and for receiving a diagnosis by ffERG was 7.3 years, with molecular diagnosis at 9.7 years. The mean (SD) length of follow-up was 11 (2.8) years. The best-corrected visual acuity at the most recent visit averaged 20/30 in the better-seeing eye (range, 20/20-20/60). The mean (SD) initial refraction was -2.80 (4.42) diopters (D) and the mean refraction at the most recent visit was -8.75 (3.53) D (range, -4.00 to -13.75 D), with the greatest rate of myopic shift before age 5 years. Full-field electroretinogram results were electronegative, consistent with cCSNB, without a significant change in amplitude over time. No patient or parent noted night blindness at presentation; however, subjective nyctalopia was eventually reported in 5 of 7 patients (71%). The full-field stimulus threshold testing results were moderately subnormal (-29.7 [3.8] dB; normal -59.8 [4.0] dB). Goldmann visual field results were significant for full I-4e, but constricted I-2e isopter. Eight different mutations or rare variants in TRPM1 predicted to be pathogenic were detected, with 3 novel variants., Conclusions and Relevance: Children with TRPM1-associated cCSNB presented before school age with progressive myopia as well as strabismus and nystagmus (but not nyctalopia), with stable, electronegative ffERG results, mildly subnormal full-field stimulus threshold testing results, and a constricted I2e isopter on perimetry. These findings suggest that ffERG and cCSNB genetic testing should be considered for children who present with early-onset myopia, especially in the presence of strabismus and/or nystagmus, and that TRPM1-associated cCSNB is a channelopathy that may present without complaints of night blindness in childhood.

The elastic features of protein filaments are encoded in their component units and in the way they are connected, thus defining a biunivocal relationship between the monomer and the result of its self-assembly. Using DNA origami approaches, we constructed a reconfigurable module, composed of two quasi-independent domains and four possible interfaces, capable of facial and lateral growing through specific recognition patterns. Whereas the flexibility of the intra-domains region can be regulated by switchable DNA motifs, the inter-domain interfaces feature mutually and self-complementary shapes, whose pairwise association leads to filaments of programmable periodicity and variable persistence length. Thus, we show here that the assembly pathway leading to oligomeric chains can be finely tuned and fully controlled, enabling the emulation of protein-like filaments using a single construction principle. Our approach results in artificial materials with a large variety of ultrastructures and bending strengths comparable, or even superior, to their natural counterparts.

Background: Phase 1 studies have shown potential benefit of gene replacement in RPE65-mediated inherited retinal dystrophy. This phase 3 study assessed the efficacy and safety of voretigene neparvovec in participants whose inherited retinal dystrophy would otherwise progress to complete blindness., Methods: In this open-label, randomised, controlled phase 3 trial done at two sites in the USA, individuals aged 3 years or older with, in each eye, best corrected visual acuity of 20/60 or worse, or visual field less than 20 degrees in any meridian, or both, with confirmed genetic diagnosis of biallelic RPE65 mutations, sufficient viable retina, and ability to perform standardised multi-luminance mobility testing (MLMT) within the luminance range evaluated, were eligible. Participants were randomly assigned (2:1) to intervention or control using a permuted block design, stratified by age (<10 years and ≥10 years) and baseline mobility testing passing level (pass at ≥125 lux vs <125 lux). Graders assessing primary outcome were masked to treatment group. Intervention was bilateral, subretinal injection of 1·5 × 10 11 vector genomes of voretigene neparvovec in 0·3 mL total volume. The primary efficacy endpoint was 1-year change in MLMT performance, measuring functional vision at specified light levels. The intention-to-treat (ITT) and modified ITT populations were included in primary and safety analyses. This trial is registered with ClinicalTrials.gov, number NCT00999609, and enrolment is complete., Findings: Between Nov 15, 2012, and Nov 21, 2013, 31 individuals were enrolled and randomly assigned to intervention (n=21) or control (n=10). One participant from each group withdrew after consent, before intervention, leaving an mITT population of 20 intervention and nine control participants. At 1 year, mean bilateral MLMT change score was 1·8 (SD 1·1) light levels in the intervention group versus 0·2 (1·0) in the control group (difference of 1·6, 95% CI 0·72-2·41, p=0·0013). 13 (65%) of 20 intervention participants, but no control participants, passed MLMT at the lowest luminance level tested (1 lux), demonstrating maximum possible improvement. No product-related serious adverse events or deleterious immune responses occurred. Two intervention participants, one with a pre-existing complex seizure disorder and another who experienced oral surgery complications, had serious adverse events unrelated to study participation. Most ocular events were mild in severity., Interpretation: Voretigene neparvovec gene replacement improved functional vision in RPE65-mediated inherited retinal dystrophy previously medically untreatable., Funding: Spark Therapeutics., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Baraitser-Winter syndrome (cerebrofrontofacial syndrome, type 3) is a rare developmental disorder typified by hypertelorism, ptosis, high-arched eyebrows, ocular coloboma, and brain malformations. Other common manifestations include hearing loss, short stature, seizures, intellectual impairment, muscle dysfunction, and abnormalities of the kidney and urinary system. This syndrome is caused by missense mutations in the genes ACTB or ACTG1, both of which encode for cytoplasmic actin proteins crucial for proper development of many organs in the human body. There are no reports of familial transmission; all reported cases have been new mutations. However, different mutations in ACTG1 have been reported to cause isolated non-syndromic hearing loss, with many reported cases of autosomal dominant (AD) inheritance. We have identified a three-generation pedigree segregating a novel mutation in the ACTG1 gene that causes Baraitser-Winter Syndrome with extremely variable expressivity, leading to an initial diagnosis of isolated AD hearing loss in two members. Subtle optic nerve signs not previously reported in this syndrome are also documented in one patient.

Introduction: Infantile nystagmus has many causes, some life threatening. We determined the most common diagnoses in order to develop a testing algorithm., Methods: Retrospective chart review. Exclusion criteria were no nystagmus, acquired after 6 months, or lack of examination., Data Collected: pediatric eye examination findings, ancillary testing, order of testing, referral, and final diagnoses. Final diagnosis was defined as meeting published clinical criteria and/or confirmed by diagnostic testing. Patients with a diagnosis not meeting the definition were "unknown." Patients with incomplete testing were "incomplete." Patients with multiple plausible etiologies were "multifactorial." Patients with negative complete workup were "motor.", Results: A total of 284 charts were identified; 202 met inclusion criteria. The three most common causes were Albinism (19%), Leber Congenital Amaurosis (LCA; 14%), and Non-LCA retinal dystrophy (13%). Anatomic retinal disorders comprised 10%, motor another 10%. The most common first test was MRI (74/202) with a diagnostic yield of 16%. For 28 MRI-first patients, nystagmus alone was the indication; for 46 MRI-first patients other neurologic signs were present. 0/28 nystagmus-only patients had a diagnostic MRI while 14/46 (30%) with neurologic signs did. The yield of ERG as first test was 56%, OCT 55%, and molecular genetic testing 47%. Overall, 90% of patients had an etiology identified., Conclusion: The most common causes of infantile nystagmus were retinal disorders (56%), however the most common first test was brain MRI. For patients without other neurologic stigmata complete pediatric eye examination, ERG, OCT, and molecular genetic testing had a higher yield than MRI scan. If MRI is not diagnostic, a complete ophthalmologic workup should be pursued.

From atoms to molecules and bio-macromolecules, from organelles to cells, tissues, to the whole living system, nature shows us that the formation of complex systems with emergent properties originates from the hierarchical self-assembly of single components in guided bottom-up processes. By using DNA as a fundamental building block with well-known self-recognition properties, scientists have developed design rules and physical-chemical approaches for the fully programmable construction of highly organized structures with nanosized features. This review highlights the basic principles of hierarchical self-assembly in terms of type and number of distinguishable components and their interaction energies. Such general concepts are then applied to DNA-based systems. After a brief overview of the strategies used until now for the construction of individual DNA units, such as DNA tile motifs and origami structures, their self-association into assemblies of higher order is discussed. Particular emphasis is given to the forces involved in the self-assembly process, understanding and rational combination of which might help to coordinate the single elements of hierarchical structures both in space and time, thus advancing our efforts towards the creation of devices that mimic the complexity and functionality of natural systems., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Purpose: To determine the sensitivity and specificity of the Web-based vision-screening test (WBT) VisionForKids.org when administered by an untrained layperson in a controlled environment., Methods: Visual acuities were obtained by an untrained layperson using the WBT under observation and by an ophthalmic professional using the Electronic Visual Acuity (EVA) Tester. Subjects were randomized to which method was used first. Subjects were considered to have failed by the following criteria: in children <48 months, 20/50 vision; in children between 48 and 59 months, 20/40 or worse; in children ≥60 months, 20/30 or worse. The results of the vision examinations by the WBT and the EVA Tester were compared; sensitivity and specificity of the WBT and the correlation coefficient to detect normal and abnormal vision compared to EVA testing were determined., Results: Visual acuities were obtained on 203 children between ages 3 and 12 years (average age, 7.4 years). Sensitivity of the WBT was 78.7% (95% CI, 66.0%-87.7%), specificity was 89.4% (95% CI, 82.9%-93.8%). The correlation coefficient of EVA testing with the WBT was 0.89 (P < 0.001)., Conclusions: This layperson-administered WBT, VisionForKids.org, is valid for identifying amblyopia in a controlled environment, making it possible for cost-effective and easily accessible vision screening to be performed by laypersons., (Copyright © 2015 American Association for Pediatric Ophthalmology and Strabismus. Published by Elsevier Inc. All rights reserved.)

Breast carcinomas caused by inheritance of cancer-predisposing germ-line mutations have specific bioclinical features. This study aimed to analyze the efficacy of conventional cytotoxic treatment in BRCA1 and CHEK2 mutation carriers and non-carriers. The study included 415 Russian breast cancer patients aged 50 years or younger, who were subjected to various standard schemes of neoadjuvant therapy. The choice of therapy was done without the knowledge of the mutations status, because DNA testing was performed retrospectively using the archival tissue samples. 19 BRCA1 (4.6%) and 8 CHEK2 (1.9%) heterozygous genotypes were identified. BRCA1 mutation carriers achieved pathological complete response more frequently than non-carriers [6/19 (31.6%) vs. 46/388 (11.9%), p = 0.024]; this effect was limited to women treated by anthracycline-based therapy without taxanes [5/9 (55.6%) vs. 28/247 (11.3%), p = 0.002] and was not observed in any of 7 BRCA1 carriers receiving taxane-containing regimens. CHEK2 heterozygotes did not experience pathological complete response and showed lower frequency of objective clinical responses as compared to mutation non-carriers [4/8 (50%) vs. 333/388 (85.5%), p = 0.020]; the efficacy of neoadjuvant therapy was particularly poor in CHEK2 carriers receiving anthracyclines without taxanes. This study provides evidence for distinct sensitivity of BRCA1 and CHEK2 mutation-driven breast carcinomas to standard chemotherapeutic schemes.

Background: This multicentric randomized phase II study investigated the feasibility and toxicity of temozolomide (TMZ) added to whole brain radiotherapy (WBRT) followed by adjuvant TMZ in patients with multiple brain metastases of non-small-cell lung cancer (NSCLC)., Methods: Patients with multiple brain metastases from NSCLC aged ≥ 18 years, classified according to recursive partitioning analysis class I or II and with adequate organ functions were eligible. Treatment consisted of WBRT + TMZ 75 mg/m² for 2 weeks followed at day 28 by TMZ 100 mg/m²/day 2 weeks on/2 weeks off for up to 6 months (radiochemotherapy, RCT) or WBRT alone (radiotherapy, RT)., Results: The study enrolled only 35 patients (22 patients in RCT and 13 in RT) and had to be closed prematurely due to poor accrual. The toxicity was mainly due to TMZ with WHO grade 3 and 4 thrombocytopenia in 3/22 versus 0/13, leucocytopenia in 1/22 versus 0/13 and lymphocytopenia in 7/22 versus 12/13 patients in RCT and RT respectively. Thirteen patients in RCT and six in RT progressed systemically and dropped out before first restaging of the response in brain. Median time to progression (TTP) was 2.4 months (95 % CI: 2-2.6 months) and 2.0 months (95 % CI: 0.5-3.5 months), median overall survival (OAS) was 3 months (95% CI: 1.7-3.1 months) and 6.3.months (95 % CI: 0.2-7.6 months) in RCT and RT, respectively., Conclusions: Like other studies before on patients with brain metastases, insufficient number of recruited patients does not allow conclusions on efficacy and toxicity as the study closed prematurely.

Purpose: To investigate the incidence and characteristics of occlusion amblyopia with prescribed full-time patching and determine its effect on long-term visual acuity outcomes., Methods: The records of patients younger than 10 years diagnosed as having amblyopia between 1970 and 2000 were retrospectively reviewed. Patients were prescribed full-time occlusion and observed until completion of therapy., Results: Of 597 patients treated for amblyopia by full-time patching, 115 were diagnosed as having occlusion amblyopia (19.3%). Seventy-five percent (86 of 115) developed occlusion amblyopia during the first episode of full-time patching. Occlusion amblyopia occurred more frequently in children prescribed full-time patching at an earlier age (P = .0002), with an odds ratio of 8.56 (95% confidence interval: 2.73, 26.84) in children younger than 36 months and 2.66 (95% confidence interval: 0.96, 7.37) in children between 36 and 59 months old. Seven of the patients with occlusion amblyopia did not reverse fixation and continued to fixate with the initially amblyopic eye after treatment. Final visual acuity in these eyes with occlusion amblyopia was 20/30 or better. After cessation of treatment, the final interocular difference in visual acuity was less in patients with a history of occlusion amblyopia (P = .003)., Conclusion: Occlusion amblyopia occurred at all ages, but the incidence decreased with increasing age. Patients who developed occlusion amblyopia with prescribed full-time occlusion had less interocular visual acuity difference than patients who did not, suggesting that development of occlusion amblyopia can indicate the potential for the development of better vision in the originally amblyopic eye., (Copyright 2013, SLACK Incorporated.)

Objective: Recent US Preventive Services Task Force recommendations on vision screening reported insufficient data to recommend vision screening in children <3 years of age. The Iowa photoscreening program, KidSight, has screened children from 6 months of age and older since 2000. We report our experience with vision screening in these children and compare the results of the photoscreens in children younger than 3 years with those of children of preschool age and older., Methods: A retrospective review of results from the Iowa KidSight database using the MTI PhotoScreener containing results of children screened between May 1, 2000, and April 30, 2011., Results: During the 11 years of the study, 210 695 photoscreens on children were performed at 13 750 sites. In the <3-year age group, the unreadable rate was 13.0%, the referral rate was 3.3%, and the overall positive-predictive value was 86.6%. In the 3- to 6-year-old children, the unreadable rate was 4.1%, the referral rate was 4.7%, and the overall positive-predictive value was 89.4%., Conclusions: No statistically significant difference was found in screening children from 1 to 3 years old compared with screening children >3 years old. These results confirm that early screening, before amblyopia is more pronounced, can reliably detect amblyogenic risk factors in children younger than 3 years of age, and we recommend initiation of photoscreening in children aged 1 year and older.

The BLM gene belongs to the RecQ helicase family and has been implicated in the maintenance of genomic stability. Its homozygous germline inactivation causes Bloom syndrome, a severe genetic disorder characterized by growth retardation, impaired fertility and highly elevated cancer risk. We hypothesized that BLM is a candidate gene for breast cancer (BC) predisposition. Sequencing of its entire coding region in 95 genetically enriched Russian BC patients identified two heterozygous carriers of the c.1642 C>T (Q548X) mutation. The extended study revealed this allele in 17/1,498 (1.1%) BC cases vs. 2/1,093 (0.2%) healthy women (p = 0.004). There was a suggestion that BLM mutations were more common in patients reporting first-degree family history of BC (6/251 (2.4%) vs. 11/1,247 (0.9%), p = 0.05), early-onset cases (12/762 (1.6%) vs. 5/736 (0.7%), p = 0.14) and women with bilateral appearance of the disease (2/122 (1.6%) vs. 15/1376 (1.1%), p = 0.64). None of the BLM-associated BC exhibited somatic loss of heterozygosity at the BLM gene locus. This study demonstrates that BLM Q548X allele is recurrent in Slavic subjects and may be associated with BC risk., (Copyright © 2011 UICC.)

Introduction: This randomized phase II study investigated pemetrexed in combination with the epidermal growth factor receptor (EGFR)-targeting monoclonal antibody matuzumab compared with pemetrexed alone as second-line therapy for patients with advanced non-small cell lung cancer., Methods: Patients received pemetrexed 500 mg/m every 3 weeks either alone (n = 50) or in combination with matuzumab at either 800 mg weekly (n = 51) or 1600 mg every 3 weeks (n = 47). The primary end point was objective response, as assessed by an independent review committee., Results: Tumor EGFR expression was detected in 87% of randomized patients. The objective response rate for the pooled matuzumab-treated arms was 11% compared with 5% for pemetrexed alone (p = 0.332). Apart from one patient in the pemetrexed alone group, all responses occurred in patients whose tumors expressed EGFR. The objective response rate for patients receiving weekly matuzumab was 16% compared with 2% for those receiving matuzumab every 3 weeks. There was also a trend for improved overall survival in patients receiving matuzumab weekly versus every 3 weeks (12.4 months versus 5.9 months, respectively, versus 7.9 months for pemetrexed alone). The combination of pemetrexed and matuzumab demonstrated an acceptable safety profile, with the most common grade 3/4 adverse event being neutropenia., Conclusion: Although the analysis on the pooled matuzumab-treated arms did not demonstrate a statistically significant improvement in objective response for the addition of matuzumab to pemetrexed compared with pemetrexed alone, the trends for improvement in objective response and overall survival for pemetrexed plus weekly matuzumab compared with pemetrexed alone warrant confirmation in additional clinical trials.

The purpose of this study was to determine if patients and their families in a pediatric ophthalmology and adult strabismus clinic have a preference regarding physician and staffattire. Patients and/or parents were invited to complete a three-question survey. Respondents were queried based on three types of attire preference: white coat, professional clothing without white coat, or casual attire without white coat. Two hundred twenty-seven patients participated. Of the patients queried, no preference for any one style of physician attire was found. These results do not support previous studies indicating significant preference for white coats.

Purpose: To present the largest cohort of preschool children screened by the MTI PhotoScreener over a 9-year period from a single, statewide vision screening effort., Design: Cross-sectional study., Participants: We included 147,809 children screened between May 1, 2000, and April 30, 2009 by a photoscreening program., Methods: Retrospective review of results from the Iowa photoscreening program using the MTI PhotoScreener. The photographs were taken by volunteers from local Lions clubs and sent to the University of Iowa for interpretation. Children who failed the photoscreening were referred to local eye care professionals, who preformed a comprehensive eye evaluation and forwarded the results to the Iowa KidSight program., Main Outcome Measures: Number of screenings, referral rate, positive predictive value (PPV), follow-up rate, and associated costs per year are described., Results: Over the 9 years of the continuously operating program, 147,809 children underwent photoscreens to detect amblyopic risk factors at 9746 sites. Because of abnormal photoscreen results, 6247 children (4.2%) were referred. Of the children, 24.3% were evaluated by local ophthalmologists and 76.7% were seen by local optometrists. Between 2000 and 2009, the follow-up rate ranged from a low of 36.1% to a high of 89.5%, with an overall program follow-up rate after the addition of the follow-up coordinator of 81.3%. The overall PPV of the MTI PhotoScreener was 94.2%. Taking into account overall operating budget including salaries and associated costs, the cost of screening 1 child has been reduced to $US9 per child., Conclusions: The addition of a part-time follow-up coordinator to the photoscreening program produced 89.5% follow-up rate when screening 147,809 children for amblyopia risk factors over a 9-year period., (Copyright © 2010 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Background: The Lung Cancer Cetuximab Study is an open-label, randomized phase II pilot study of cisplatin and vinorelbine combined with the epidermal growth factor receptor (EGFR)-targeted monoclonal antibody cetuximab versus cisplatin and vinorelbine alone, in patients with advanced EGFR-expressing, non-small-cell lung cancer (NSCLC). End points of the study are activity, safety and pharmacokinetics., Patients and Methods: Following randomization, for a maximum of eight cycles, patients received three-weekly cycles of cisplatin (80 mg/m(2), day 1) and vinorelbine (25 mg/m(2) on days 1 and 8) alone or following cetuximab treatment (initial dose 400 mg/m(2), followed by 250 mg/m(2) weekly thereafter)., Results: Eighty-six patients were randomly allocated to the study (43 per arm). Confirmed response rates were 28% in the cisplatin/vinorelbine arm (A) and 35% in the cetuximab plus cisplatin/vinorelbine arm (B). Median progression-free survival (PFS) was 4.6 months in arm A and 5.0 months in arm B, with PFS rates at 12 months of 0% and 15%, respectively. Median survival was 7.3 months in arm A and 8.3 months in arm B. The 24-month survival rates were 0% and 16%, respectively. The cetuximab combination was well tolerated., Conclusion: In the first-line treatment of advanced NSCLC, the combination of cetuximab plus cisplatin/vinorelbine demonstrated an acceptable safety profile and the potential to improve activity over cisplatin/vinorelbine alone.

Objective: The aim of this investigation was to study the effect of vitamin E treatment in oxidative stress of red and white cells of beta-thalassaemia intermedia patients., Methods: Nine patients undergoing occasional transfusions (5 females/4 males), median age 39 years (range 15-74), were recruited for oral daily administration of 400 IU vitamin E for 3 months. Twenty-seven milliliters of peripheral blood was obtained before and after 3 months of treatment, and 3 months after treatment completion. In the case of transfused patients (n = 4), blood was obtained at least 30 days after transfusion. Reactive oxygen species (ROS) was measured by flow cytometry; red blood cell (RBC) reduced glutathione (GSH) was measured by dinitrothiocyanobenzene reduction, serum malondialdehyde was measured in terms of thiobarbituric acid-reactive substances (TBARS), and alpha-haemoglobin-stabilizing protein (AHSP) mRNA expression was measured by real-time polymerase chain reaction of reticulocyte RNA extracts., Results: beta-Thalassaemia patients presented basal levels of RBC ROS, GSH and serum TBARS statistically different compared with healthy controls. However, after vitamin E administration, patients presented a significant reduction in erythrocyte RBC ROS and serum TBARS levels. In parallel, red cell GSH was significantly increased after treatment. Peripheral mononuclear cells and T lymphocytes also demonstrated a reduction in ROS production. Therefore, after treatment, no significant differences were detected comparing patients and normal controls. Three months after treatment completion, all measurements showed a tendency of returning to basal values. A significant reduction in reticulocyte number was observed after vitamin E treatment. Vitamin E treatment did not modify levels of haemoglobin or AHSP mRNA expression., Conclusion: Although vitamin E is not capable of reducing anaemia in these patients, it could be useful for reducing oxidative damage in other target organs of beta-thalassaemic patients. Finally, this is the first study to analyse the effects of vitamin E on ROS production in red and white blood cells and AHSP mRNA expression., (Copyright 2009 S. Karger AG, Basel.)

Purpose: To determine the stability of visual acuity (VA) after a standardized occlusion regimen in children with strabismic and/or anisometropic amblyopia., Design: Retrospective, population-based, consecutive observational case series., Participants: Four hundred forty-nine patients younger than 10 years who underwent an occlusion trial for amblyopia and were observed until there was a recurrence of amblyopia or for a maximum of 1 year after decrease or cessation of occlusion therapy., Methods: We performed a retrospective chart review of all patients treated by occlusion therapy for strabismic and/or anisometropic amblyopia at our institution over a 34-year period. Of the 1621 patients identified in our database, 449 met the eligibility criteria and were included in this study. Patients having at least a 2 logarithm of the minimum angle of resolution (logMAR)-level improvement in VA by optotypes or a change from unmaintained to maintained fixation preference during the course of occlusion therapy were included. A recurrence of amblyopia was defined as > or =2 logMAR levels of VA reduction or reversal of fixation preference within 1 year after a decrease or cessation of occlusion therapy., Main Outcome Measure: Recurrence of amblyopia after a decrease or cessation of occlusion therapy and its relationship with patient age and VA of the amblyopic eye at the time of decrease or cessation of occlusion therapy., Results: Of 653 occlusion trials, 179 (27%) resulted in recurrence of amblyopia. The recurrence was found to be inversely correlated with patient age. There was no statistically significant association between the recurrence of amblyopia and VA of the amblyopic eye at the end of maximal occlusion therapy., Conclusions: There is a clinically important risk of amblyopia recurrence when occlusion therapy is decreased before the age of 10 years. The risk of recurrence is inversely correlated with age (P<0.0001).

Timely screening for the early detection of eye and vision problems in children is vital to avoid lifelong visual impairment. Early detection provides the best opportunity for effective treatment. Organizations devoted to the health and well-being of children have issued recommendations or policy statements regarding the timing of eye screenings and examinations. The American Academy of Pediatrics (AAP), American Association of Pediatric Ophthalmology and Strabismus (AAPOS), American Academy of Ophthalmology (AAO), American Association of Certified Orthoptists (AACO), and the American Academy of Family Physicians (AAFP) recommend screening at birth and all well-child visits. The American Optometric Association (AOA) recommends that vision examinations begin at 6 months of age. All children found to have an ocular abnormality or who fail vision assessment should be referred to an eye care specialist trained to treat pediatric patients.

Background: Earlier detection of childhood vision disorders is a prominent goal of vision screening. The Medical Technology and Innovation (MTI) PhotoScreener addresses this objective. Use of this camera does not require verbal feedback and may be administered early in a child's development. Decreasing the variability in photograph grading results will boost the utility of any photoscreening system. This report aims to understand and to decrease intra- and inter-observer variability in grading photoscreening photographs., Methods: We dissected the photograph grading process and quantified the intra- and inter-observer agreement using intraclass plot correlation coefficients. We evaluated the outcome of a two grader verification system vs. adjudicated measurements with Receiver Operator Characteristic (ROC) curves., Participants: Data on 955 children under 5 years of age, normal except for refractive error, each with complete photoscreening and eye examination data, culled from two previous studies., Main Outcome Measures: Intra- and inter-observer agreement in measuring bright crescent dimensions and pupillary diameters. Sensitivity and specificity of detection of hyperopia., Results: Measurements of bright crescents are associated with greater variability than are measurements of pupillary diameters. Recognition and omission of light "rim" measurements from photograph grading will result in superior inter-observer agreement. Photograph independent errors increase variability and may be corrected by remeasurement. A verification system in which the most discrepant 5% of measurements are redone results in ROC curves similar to adjudicated dimension., Conclusions: We conclude: 1) two novices grading photographs can do as well as one expert in most cases; 2) the proposed grading methodology has undergone statistical validation and can be used in other areas of ophthalmology and medicine; and 3) inter-observer variability, one of the limitations of photoscreening photograph grading, can be reduced. For 95% of the photographs, two novices achieve similar true positive and true negative values with or without adjudication.

Background: CD30-positive cutaneous lymphoproliferative disorders (LPD) represent a spectrum of diseases ranging from low-grade (lymphomatoid papulosis; LyP) to high-grade (pleomorphic and anaplastic large-cell lymphoma; PTL, ALCL) with overlapping morphologic and immunophenotypic features. The common phenotypic hallmark is the expression of CD30-antigen by the tumor cells which morphologically resemble Reed-Sternberg cells. Although LyP is a non-fatal recurring disorder, it is associated with systemic lymphomas including Hodgkin's lymphoma (HL), mycosis fungoides (MF) and ALCL in 5-20% of the cases. Currently there is no marker to predict the development of systemic lymphomas in patients with LyP. Fascin, an actin bundling protein, has recently been shown to be a unique marker found in almost 100% of classical HL., Methods: Because of the association of LyP with HL, fascin expression was analyzed by immunohistochemistry in LyP (n = 45), cutaneous CD30+ ALCL (n = 17) and pleomorphic T-cell lymphoma (n = 9) (PTL) and LyP associated with systemic lymphomas (7 HL, 2 ALCL, 1 MF), with the intent to determine if fascin expression can predict disease progression., Results: Fascin was expressed by tumor cells in 11/45 (24%) cases of LyP, 11/17 (64%) cases of ALCL, 7/9 (77%) cases of PTL and 6/10 (60%) cases of LyP associated with systemic lymphomas. Fascin expression in LyP was significantly less frequent than in ALCL (p < 0.001) and also than in LyP associated with lymphomas (p < 0.05). There was no significant difference of fascin expression within the histological subtypes of LyP. We found no evidence of ALK expression nor of Epstein-Barr virus expression in any case either by in situ hybridization or immunohistochemistry in the LyP cases associated with HL., Conclusions: This is the first study to demonstrate that fascin is expressed in cutaneous CD30+ LPD and that it is a candidate marker of disease progression in LyP.

A pupillary examination takes very little time and does not require expensive equipment. Unfortunately, many ophthalmic personnel do not take the time to thoroughly evaluate pupillary function. Pupillary examinations can be easy if divided into specific elements for assessment. These elements should include color, shape, size, equality, and reaction to stimulus. This article will briefly review some of the more common pupillary abnormalities and review the steps of the pupillary examination.

Reflecting the stepwise process of oncogenesis, lymphomas may cumulatively develop a more aggressive phenotype during the course of disease, a process referred to as lymphoma progression. Although morphological, clinical and biological aspects of lymphoma progression do not always overlap, changes in lymphoma morphology frequently indicate alterations in the clinical and biological behaviour of the disease. Indolent and aggressive lymphomas in disease progression can either be clonally related or represent clonally unrelated neoplasms. We propose to use the term 'lymphoma progression' in a biological sense denoting only clonal development of and within a lymphoma entity. The term 'composite lymphoma' should be used as a merely descriptive morphological designation for different lymphoma entities in one individual irrespective of clonal relationship. Many types of aggressive B-cell non-Hodgkin's lymphomas and Hodgkin's lymphomas are reported to secondarily develop in lymphoma progression. Genetic changes associated with lymphoma progression frequently abrogate the differentiating effects of alterations occurring in indolent lymphomas, leading to increased cell proliferation. Within different lymphoma entities, high-risk disease variants mimicking lymphoma progression exist.

The Bundeskonferenz für Erziehungsberatung (bke) is the federation for child guidance and family-counseling in Germany. Members are the working groups for child guidance and family-counseling of the Länder, where the employees of the child guidance are organized. The federation was founded in 1962. It is sponsored by the Federal Ministry for Family, Seniors, Women and Youth. Further education for professional youth workers is central mission of bke. The constitution of the federation lais down these contents. Every autumn the bke publishes the program for further education for the following year in a brochure called Zentrale Weiterbildung--the program for child guidance, family counseling and youth counseling. A special Kommission Zentrale Weiterbildung acquires the contents of the program for further education und selects the referents. Since 1968 the Zentrale Weiterbildung of Bundeskonferenz für Erziehungsberatung offers events for professional youth workers in child guidance, family counseling. The program for further education contributes the quality of the institution of child guidance. After a time with above average use of the program for further education with therapeutical focal point, the orientation of the program changes to child and youth services. A constant number of counselors use the program. Beside the change of contents there was a change to effectiveness of contents and a reduction of participants.

Lymphomatoid papulosis is part of a spectrum of CD30+ cutaneous lymphoproliferative disorders characterized by spontaneous tumor regression. The mechanism(s) of regression is unknown. In a recent study, a selective increase in CD30 ligand expression in regressing lesions of lymphomatoid papulosis and cutaneous CD30+ anaplastic large cell lymphoma was shown, suggesting that activation of the CD30 signaling pathway may be responsible for tumor regression, whereas no difference in Fas/Fas ligand expression was found between regressing and nonregressing lesions. Therefore we tested the effects of CD30 and Fas activation on three CD30+ cutaneous lymphoma cell lines (Mac-1, Mac-2 A, JK) derived from nonregressing tumors of two patients who had progressed from lymphomatoid papulosis to systemic anaplastic large cell lymphoma. To evaluate the effects of CD30 signaling, the cell lines were incubated with a CD30 agonistic antibody, HeFi-1. Proliferative responses, mitogen-activated protein kinase, and nuclear factor kappa B activities were determined with and without CD30 activation. Mac-1 and Mac-2 A showed increased proliferative responses to incubation with CD30 activating antibody, HeFi-1. Inhibition of the mitogen-activated protein kinase activity caused growth inhibition of the Mac-1, Mac-2 A, and JK cell lines. Activation of the Fas pathway induced apoptosis in all three cell lines. Taken together, these findings suggest that resistance to CD30-mediated growth inhibition provides a possible mechanism for escape of cutaneous anaplastic large cell lymphoma from tumor regression. Mitogen-activated protein kinase inhibitors are potential therapeutic agents for the treatment of advanced cutaneous anaplastic large cell lymphoma. J Invest Dermatol 115:1034-1040, 2000

Spontaneous regression of skin lesions is characteristic of lymphomatoid papulosis (LyP), a clonal cutaneous lymphoproliferative disorder. A minority of LyP patients progress to anaplastic large cell lymphoma (ALCL) in which skin lesions no longer regress and extracutaneous dissemination often occurs. In 1 such case, we developed a tumor cell line, JK cells, and show that these cells are resistant to the growth inhibitory effects of transforming growth factor beta (TGF-beta) due to the loss of cell surface expression of the TGF-beta type I receptor (TbetaR-I). Reverse transcriptase-polymerase chain reaction (RT-PCR) and sequencing of JK cell TbetaR-I cDNA clones identified a deletion that spanned the last 178 bp of exon 1, including the initiating methionine. Hybridization of a radiolabeled fragment internal to the deletion was detected in the genomes of TGF-beta-responsive cells, but not in JK cells, indicating that they contain no wild-type TbetaR-I gene. PCR primers that flanked the deleted TbetaR-I region amplified a single band from JK cell genomic DNA that lacked the last 178 bp of exon 1 and all of the approximately 5 kb of intron 1. This JK cell-specific genomic TbetaR-I PCR product was distinct from products amplified from TGF-beta-responsive cells and was also readily detected in tumor biopsies obtained before the establishment of the JK cell line. Our results identify the first inactivating mutation in TbetaR-I gene in a human lymphoma that renders it insensitive to growth inhibition by TGF-beta.

To develop a model for the biology and treatment of CD30+ anaplastic large cell lymphoma (ALCL), we transplanted leukemic tumor cells from a 22-month-old girl with multiple relapsed ALCL. Tumor cells were inoculated intraperitoneally into a 4-week-old SCID/bg mouse and produced a disseminated tumor within 8 weeks; this tumor was serially transplanted by subcutaneous injections to other mice. Morphology, immunohistochemistry, and molecular genetics which demonstrated the NPM-ALK fusion protein, resulting from the t(2;5)(p23;q35), confirmed the identity of the xenograft with the original tumor. The tumor produced transcripts for interleukin-1alpha, tumor necrosis factor-alpha, and interferon-gamma which could explain the patient's B-symptoms. Treatment of mice with monoclonal antibody (HeFi-1) which activates CD30 antigen administered on day 1 after tumor transplantation prevented tumor growth. Treatment with HeFi-1 after tumors had reached a 0.2 cm(3) volume caused tumor growth arrest and prevention of tumor dissemination. We conclude that transplantation of CD30+ ALCL to SCID/bg mice may provide a valuable model for the study of the biology and design of treatment modalities for CD30+ ALCL.

Purpose: To evaluate the feasibility of interferon gamma (IFNgamma) as an adjunct to chemotherapy in advanced non-small-cell lung cancer (NSCLC)., Methods: A total of 32 patients were recruited and received 25 mg/m2 cisplatin and 100 mg/m2 etoposide on days 8, 10 and 12 every 3 weeks for a total of three cycles. A dose of 100 microg IFNgamma was given subcutaneously three times weekly from days 1 to 8 and between days 15 and 29. After induction, all patients except those with progressive disease were offered IFNgamma maintenance therapy: 100 microg three times weekly., Results: The following responses were obtained: partial response, 5 (16%); minor response, 12 (37%); stable disease, 4 (13%); progressive disease, 11 (34%). The survival rates after 1 and 2 years were 47% and 25% respectively. Patients receiving maintenance IFNgamma had a 2-year survival rate of 58%. Toxic side-effects were rare and included grade III/IV fever (7%/1%) and grade III/IV leucopenia (4%/1%)., Conclusions: In patients with advanced NSCLC, an adjunctive dose of 100 microg IFNgamma, given three times weekly in the induction and maintenance phase, is feasible. Survival data seem favourable so this regimen may warrant further investigation in a phase III study.