Retinoblastoma has properties of a cone precursor tumor and depends upon cone-specific MDM2 signaling.

Retinoblastomas result from the inactivation of the RB1 gene and the loss of Rb protein, yet the cell type in which Rb suppresses retinoblastoma and the circuitry that underlies the need for Rb are undefined. Here, we show that retinoblastoma cells express markers of postmitotic cone precursors but not markers of other retinal cell types. We also demonstrate that human cone precursors prominently express MDM2 and N-Myc, that retinoblastoma cells require both of these proteins for proliferation and survival, and that MDM2 is needed to suppress ARF-induced apoptosis in cultured retinoblastoma cells. Interestingly, retinoblastoma cell MDM2 expression was regulated by the cone-specific RXRgamma transcription factor and a human-specific RXRgamma consensus binding site, and proliferation required RXRgamma, as well as the cone-specific thyroid hormone receptor-beta2. These findings provide support for a cone precursor origin of retinoblastoma and suggest that human cone-specific signaling circuitry sensitizes to the oncogenic effects of RB1 mutations.

Non-synonymous, synonymous, and non-coding nucleotide variants contribute to recurrently altered biological processes during retinoblastoma progression.

Retinoblastomas form in response to biallelic RB1 mutations or MYCN amplification and progress to more aggressive and therapy-resistant phenotypes through accumulation of secondary genomic changes. Progression-related changes include recurrent somatic copy number alterations and typically non-recurrent nucleotide variants, including synonymous and non-coding variants, whose significance has been unclear. To determine if nucleotide variants recurrently affect specific biological processes, we identified altered genes and over-represented variant gene ontologies in 168 exome or whole-genome-sequenced retinoblastomas and 12 tumor-matched cell lines. In addition to RB1 mutations, MYCN amplification, and established retinoblastoma somatic copy number alterations, the analyses revealed enrichment of variant genes related to diverse biological processes including histone monoubiquitination, mRNA processing (P) body assembly, and mitotic sister chromatid segregation and cytokinesis. Importantly, non-coding and synonymous variants increased the enrichment significance of each over-represented biological process term. To assess the effects of such mutations, we examined the consequences of a 3' UTR variant of PCGF3 (a BCOR-binding component of Polycomb repressive complex I), dual 3' UTR variants of CDC14B (a regulator of sister chromatid segregation), and a synonymous variant of DYNC1H1 (a regulator of P-body assembly). One PCGF3 and one of two CDC14B 3' UTR variants impaired gene expression whereas a base-edited DYNC1H1 synonymous variant altered protease sensitivity and stability. Retinoblastoma cell lines retained only ~50% of variants detected in tumors and enriched for new variants affecting p53 signaling. These findings reveal potentially important differences in retinoblastoma cell lines and tumors and implicate synonymous and non-coding variants, along with non-synonymous variants, in retinoblastoma oncogenesis.

Full-field Scotopic Threshold Improvement after Voretigene Neparvovec-rzyl Treatment Correlates with Chorioretinal Atrophy.

PURPOSE: To analyze demographic and ophthalmic data in patients with and without chorioretinal atrophy after voretigene neparvovec-rzyl (VN) to identify possible causes for this phenomenon. DESIGN: Retrospective cohort study with longitudinal follow-up. PARTICIPANTS: A total of 71 eyes of 38 patients aged 2 to 44 years with RPE65-mediated retinal dystrophy treated with VN across 2 large gene therapy centers in the United States and Germany. METHODS: Patients treated with VN who developed atrophy were compared with those who did not. MAIN OUTCOME MEASURES: Gender, age, surgical center, spherical equivalent refraction, best-corrected visual acuity (BCVA), baseline full-field scotopic threshold testing (FST), and posttreatment change in FST. RESULTS: A total of 20 eyes of 12 patients developed atrophy after treatment with VN (28% of all eyes). There was no significant difference in gender, age, surgical center, or spherical equivalent refraction between the atrophy group and the no atrophy group. However, patients between school age and young adulthood were predominantly affected, whereas the youngest and the oldest patients did not develop atrophy. Baseline BCVA was better in patients who developed atrophy than those who did not (P = 0.006). The postoperative improvement in FST at 1 month was significantly higher in the atrophy group than in the no atrophy group (P = 0.0005), and this difference remained statistically significant at 1 year (P = 0.0001). There was no correlation to baseline FST, to inflammation, or to which eye was treated first. CONCLUSIONS: The degree of FST improvement after VN appears to be strongly correlated with the development of VN-related chorioretinal atrophy. This finding raises the possibility that atrophy may develop as a toxic or metabolic sequela of vector-mediated RPE65 expression. In light of the expanding number of retinal gene therapy clinical trials, this complication warrants further study because it may not be limited to VN. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Neutralisation of SARS-CoV-2 by anatomical embalming solutions.

Teaching and learning anatomy by using human cadaveric specimens has been a foundation of medical and biomedical teaching for hundreds of years. Therefore, the majority of institutions that teach topographical anatomy rely on body donation programmes to provide specimens for both undergraduate and postgraduate teaching of gross anatomy. The COVID-19 pandemic has posed an unprecedented challenge to anatomy teaching because of the suspension of donor acceptance at most institutions. This was largely due to concerns about the potential transmissibility of the SARS-CoV-2 virus and the absence of data about the ability of embalming solutions to neutralise the virus. Twenty embalming solutions commonly used in institutions in the United Kingdom and Ireland were tested for their ability to neutralise SARS-CoV-2, using an established cytotoxicity assay. All embalming solutions tested neutralised SARS-CoV-2, with the majority of solutions being effective at high-working dilutions. These results suggest that successful embalming with the tested solutions can neutralise the SARS-CoV-2 virus, thereby facilitating the safe resumption of body donation programmes and cadaveric anatomy teaching.

Development and Evaluation of a Tele-Education Program for Neonatal ICU Nurses in Armenia.

PURPOSE: Neonatal mortality currently accounts for more than 60% of all infant mortality in Armenia. The majority of health professionals in Neonatal Intensive Care Units (NICU) however, have had no specialized training in neonatology. Local and global agencies have emphasized the need for improvement in the education and skills of NICU health professionals. The objective of this study was to design, implement, and evaluate an innovative tele-education program for NICU nurses in Armenia. DESIGN AND METHODS: Seven online tele-education courses were designed on various neonatal nursing topics using Final Cut Pro and translated professionally to Armenian. The videos were uploaded to YouTube. All NICU nurses employed in two hospitals (n = 35) in Yerevan, Armenia, completed a diagnostic pretest, course viewing, a posttest, and a satisfaction survey for each topic. A difference in knowledge was defined as the number of correct test answers obtained before and after the course. Participant satisfaction was measured using a Likert scale. RESULTS: The combined average for completed pre-tests for all courses was 45% and 71% for post-tests, which was statistically significant for each course (p < 0.05). A majority of the nurses either agreed or strongly agreed with all of the satisfaction parameters of the course. CONCLUSIONS: These results support a tele-education model for effectively providing continuing education to NICU nurses in Armenia. A similar platform could be used to establish nationwide certification programs for neonatal nurses. PRACTICE IMPLICATIONS: Tele-education technology can be used effectively by nursing educators working in global health as part of international learning collaboratives.

VISUAL AND ANATOMIC OUTCOMES OF PEDIATRIC ENDOSCOPIC VITRECTOMY IN 326 CASES.

PURPOSE: To report on the indications, outcomes, and complications of endoscopic vitrectomy in a large cohort of pediatric vitreoretinal patients. METHODS: This is a retrospective interventional case series consisting of 244 eyes of 211 patients aged 18 years or younger undergoing a total of 326 endoscopic vitrectomies from 2008 to 2017. A 23-gauge vitrectomy was performed with use of a 19-gauge endoscope. RESULTS: Two hundred and eleven patients with a mean age of 7.5 years (range: 0-18 years) and median follow-up since last surgery of 28 months (range: 3 months-8.7 years) were included. The most common indication for endoscopic vitrectomy was retinal detachment (234/326; 72%) with proliferative vitreoretinopathy (162/234; 69%). Other diagnoses included trauma (25%), retinopathy of prematurity (15%), and glaucoma (9%). Twenty-five percent of surgeries (80/326) were performed on eyes with significant corneal opacities. Retinal reattachment was achieved in 67% of eyes with retinal detachment (119/178). Visual acuity improved in 26% of retinal detachment eyes versus 53% of nonretinal detachment eyes (P = 0.005). Surgical complications included band keratopathy (15%), hypotony (8%), cataract (7%), and elevated intraocular pressure (3%). CONCLUSION: In this large series of pediatric endoscopic vitreoretinal surgeries, anatomic outcomes and complication rates were comparable with previous studies.

MRI-Guided Cryoablation of Epidural Malignancies in the Spinal Canal Resulting in Neural Decompression and Regrowth of Bone.

OBJECTIVE: The purpose of this article is to describe the use of MRI to safely monitor cryoablation for the treatment of spinal epidural malignancies. CONCLUSION: Use of MRI guidance to monitor percutaneous cryoablation allows ablation margins more distinct than those allowed by heat-based ablation modalities. MRI-guided cryoablation is a feasible option for treating epidural tumors involving the spinal canal, resulting in successful decompression of the tumor away from the spinal cord with regrowth of previously eroded bone around the spinal canal.

Effect of common processing of soybeans on the enzymatic activity and detectability of the protein, Dicamba Mono-Oxygenase (DMO), introduced into dicamba-tolerant MON 87708.

Assessing the safety of genetically engineered crops includes evaluating the risk (hazard and exposure) of consuming their newly expressed proteins. The dicamba monooxygenase (DMO) protein, introduced into soybeans to confer tolerance (DT) to dicamba herbicide, was previously characterized and identified to pose no food or feed safety hazards. Most agricultural commodities (e.g., soybeans, maize) enter the food supply after processing methods that can include exposure to high temperatures, harsh solvents or pH extremes that can adversely impact the structure and function of proteins. To understand the likelihood of exposure to DMO in foods from DT soy, enzymatically active and/or immunodetectable forms of DMO were measured in pilot-scale productions of two soy foods (soymilk and tofu), and eight processed fractions (full fat flour, inactivated full fat flour, defatted flour, toasted meal, protein isolate, protein concentrate, crude lecithin, and refined, bleached and deodorized oil). Western blot analysis detected DMO in tofu and in five of the eight processed fractions. DMO activity was not detected in either soymilk or tofu, nor in six of the eight processed fractions. Therefore, many commercial soy processing methods can denature and/or degrade introduced proteins, like DMO. Although the DMO protein has shown no evidence of hazard, this study demonstrates that processing further reduces any food or feed risk by limiting dietary exposure to intact DMO protein.

Safety of the Bacillus thuringiensis-derived Cry1A.105 protein: Evidence that domain exchange preserves mode of action and safety.

The lepidopteran-active Cry1A.105 protein is a chimeric three-domain insecticidal toxin with distinct structural domains derived from the naturally occurring Cry1Ab, Cry1Ac and Cry1F proteins from the soil bacterium Bacillus thuringiensis (Bt). The X-ray crystal structure of the Cry1A.105 tryptic core at 3.0 Šresolution demonstrates its high structural similarity to the tryptic core of Cry1Ac. Bioinformatics analyses demonstrate that Cry1A.105 has no significant amino acid sequence similarity to known allergens or mammalian toxins, which is the same conclusion reached for its component domains. Like its intact donor proteins, Cry1A.105 was heat labile at temperatures ≥75 °C and degraded upon exposure to gastrointestinal proteases. No adverse effects were observed in mice when Cry1A.105 was dosed orally at 2451 mg/kg body weight. Therefore, the weight of evidence supports that Cry1A.105 is safe for human and animal consumption. These results support the conclusion that the safety of a chimeric protein for human or animal consumption can be evaluated in the context of the safety of its donor proteins.

Three-dimensional printing of MRI-visible phantoms and MR image-guided therapy simulation.

PURPOSE: To demonstrate the use of anatomic MRI-visible three-dimensional (3D)-printed phantoms and to assess process accuracy and material MR signal properties. METHODS: A cervical spine model was generated from computed tomography (CT) data and 3D-printed using an MR signal-generating material. Printed phantom accuracy and signal characteristics were assessed using 120 kVp CT and 3 Tesla (T) MR imaging. The MR relaxation rates and diffusion coefficient of the fabricated phantom were measured and 1 H spectra were acquired to provide insight into the nature of the proton signal. Finally, T2 -weighted imaging was performed during cryoablation of the model. RESULTS: The printed model produced a CT signal of 102 ± 8 Hounsfield unit, and an MR signal roughly 1/3rd that of saline in short echo time/short repetition time GRE MRI (456 ± 36 versus 1526 ± 121 arbitrary signal units). Compared with the model designed from the in vivo CT scan, the printed model differed by 0.13 ± 0.11 mm in CT, and 0.62 ± 0.28 mm in MR. The printed material had T2 ∼32 ms, T2*∼7 ms, T1 ∼193 ms, and a very small diffusion coefficient less than olive oil. MRI monitoring of the cryoablation demonstrated iceball formation similar to an in vivo procedure. CONCLUSION: Current 3D printing technology can be used to print anatomically accurate phantoms that can be imaged by both CT and MRI. Such models can be used to simulate MRI-guided interventions such as cryosurgeries. Future development of the proposed technique can potentially lead to printed models that depict different tissues and anatomical structures with different MR signal characteristics. Magn Reson Med 77:613-622, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Percutaneous Image-Guided Cryoablation of Head and Neck Tumors for Local Control, Preservation of Functional Status, and Pain Relief.

OBJECTIVE: We report nine consecutive percutaneous image-guided cryoablation procedures of head and neck tumors in seven patients (four men and three women; mean age, 68 years; age range, 50-78 years). Ablation of the entire tumor for local control or ablation of a region of tumor for pain relief or preservation of function was achieved in eight of nine procedures. One patient experienced intraprocedural bradycardia, and another developed a neopharyngeal abscess. There were no deaths, permanent neurologic or functional deficits, vascular complications, or adverse cosmetic sequelae due to the procedures. CONCLUSION: Percutaneous image-guided cryoablation offers a potentially less morbid minimally invasive treatment option than salvage head and neck surgery. The complications that we encountered may be avoidable with increased experience. Further work is needed to continue improving the safety and efficacy of cryoablation of head and neck tumors and to continue expanding the use of cryoablation in patients with head and neck tumors that cannot be treated surgically.

Interventional MR Imaging for Deep-Brain Stimulation Electrode Placement.

Purpose To investigate the safety and targeting errors of deep-brain stimulation (DBS) electrodes placed under interventional magnetic resonance (MR) imaging, which allows near real-time anatomic placement without physiologic mapping. Materials and Methods Retrospectively evaluated were 10 consecutive patients (five women, five men) with a mean age of 59.9 years (age range, 17-79 years). These patients underwent interventional MR imaging-guided DBS placement for movement disorders from September 2013 to August 2014 for placement of 19 DBS electrodes in cases where traditional frame-based surgery may be challenging because of the following: dystonia resulting in difficulty in placing the patients in frame, patient's inability or unwillingness to tolerate awake surgery, or anatomic anomaly or variant that could increase the risk of bleeding from microelectrode mapping. Outcomes measured included perioperative hemorrhage, death, and stroke, and electrode functionality assessed at 2 weeks after the operation. In addition, the mean radial error and mean trajectory error were calculated. Results No intraoperative neurologic complications (n = 10 [95% confidence interval: 0%, 31%]) were observed. One patient developed aspiration pneumonia in the postoperative period. Mean radial error was 0.7 mm ± 0.4 (standard deviation) and mean trajectory error was 0.5 mm ± 0.4. All leads delivered clinically effective stimulation. Conclusion Interventional MR imaging-guided DBS electrode placement may be a safe and effective alternative to conventional frame-based surgery in well-selected patients. © RSNA, 2016.

Computer-Based Vertebral Tumor Cryoablation Planning and Procedure Simulation Involving Two Cases Using MRI-Visible 3D Printing and Advanced Visualization.

OBJECTIVE: We report the development and use of MRI-compatible and MRI-visible 3D printed models in conjunction with advanced visualization software models to plan and simulate safe access routes to achieve a theoretic zone of cryoablation for percutaneous image-guided treatment of a C7 pedicle osteoid osteoma and an L1 lamina osteoblastoma. Both models altered procedural planning and patient care. CONCLUSION: Patient-specific MRI-visible models can be helpful in planning complex percutaneous image-guided cryoablation procedures.

Endoscopic vitrectomy.

PURPOSE OF REVIEW: To update on the recent developments and surgical applications of intraocular endoscopy, and highlight its role in the modern era of microincision vitreoretinal surgery. RECENT FINDINGS: Recent progress in our understanding of the unique intraocular illumination properties of endoscopy, specifically the use of reflected (coaxial) versus conventional transmitted (dissociated) light, is redefining its role in vitreoretinal surgery. Indications for endoscope-enabled intraoperative viewing during pars plana vitrectomy include posterior segment disease with significant anterior segment opacity, difficult-to-access retroirideal diseases involving the sclerotomy, pars plana, pars plicata, ciliary sulcus, ciliary body, or peripheral lens, and complex anterior retinal detachments, particularly in diseases in children. The recent introduction of 23-gauge endoscope that works with standard microcannulas increases its utility. SUMMARY: Endoscopic vitrectomy, particularly with the recent advent of 23-gauge technology, expands our surgical armamentarium, making it a useful complement to conventional viewing systems.

Characteristics and safety assessment of intractable proteins in genetically modified crops.

Genetically modified (GM) crops may contain newly expressed proteins that are described as "intractable". Safety assessment of these proteins may require some adaptations to the current assessment procedures. Intractable proteins are defined here as those proteins with properties that make it extremely difficult or impossible with current methods to express in heterologous systems; isolate, purify, or concentrate; quantify (due to low levels); demonstrate biological activity; or prove equivalency with plant proteins. Five classes of intractable proteins are discussed here: (1) membrane proteins, (2) signaling proteins, (3) transcription factors, (4) N-glycosylated proteins, and (5) resistance proteins (R-proteins, plant pathogen recognition proteins that activate innate immune responses). While the basic tiered weight-of-evidence approach for assessing the safety of GM crops proposed by the International Life Sciences Institute (ILSI) in 2008 is applicable to intractable proteins, new or modified methods may be required. For example, the first two steps in Tier I (hazard identification) analysis, gathering of applicable history of safe use (HOSU) information and bioinformatics analysis, do not require protein isolation. The extremely low level of expression of most intractable proteins should be taken into account while assessing safety of the intractable protein in GM crops. If Tier II (hazard characterization) analyses requiring animal feeding are judged to be necessary, alternatives to feeding high doses of pure protein may be needed. These alternatives are discussed here.

Initiation of retinopathy of prematurity screening examinations in extremely premature infants.

PURPOSE: To determine whether extremely premature infants require screening for retinopathy of prematurity (ROP) if <31 weeks' postmenstrual age (PMA). METHODS: The medical records of infants born in community hospital settings at <31 weeks' gestational age (GA) were reviewed retrospectively. Prevalence and progression of ROP in infants born at <24 weeks' GA were compared with infants born at 24-30 weeks' GA. RESULTS: A total of 2,061 records were reviewed: 1,969 infants were born at 24-30 weeks' GA; 92, at <24 weeks. Infants born <24 weeks' GA were more likely to develop pre-plus and plus disease or require treatment than infants born 24-30 weeks' GA (P < 0.0001) and did so earlier (P = 0.0001). Eight infants developed pre-plus or greater ROP <31 weeks' PMA; 6 were born <24 weeks' GA. Three infants developed plus disease or required treatment <31 weeks' PMA, the earliest at 27 and 3/7 weeks. CONCLUSIONS: Clinicians should consider initiating ROP screening examinations before 31 weeks' PMA, particularly for infants born <24 weeks' GA and those with lower birth weights.

Purification of phosphinothricin acetyltransferase using Reactive brown 10 affinity in a single chromatography step.

The expression of phosphinothricin N-acetyltransferase (PAT) protein in transgenic plants confers tolerance to the herbicide glufosinate. To enable the characterization of PAT protein expressed in plants, it is necessary to obtain high purity PAT protein from the transgenic grain. Because transgenically expressed proteins are typical present at very low levels (i.e. 0.1-50 µg protein/g grain), a highly specific and efficient purification protocol is required to purify them. Based on the physicochemical properties of PAT, we developed a novel purification method that is simple, time-saving, inexpensive and reproducible. The novel method employs a single chromatography step using a reactive dye resin, Reactive brown 10-agarose. Reactive brown 10 preferentially binds the PAT protein, which can then be specifically released by one of its substrates, acetyl-CoA. Using Reactive brown 10-agarose, PAT protein was purified to homogeneity from cottonseed with high recovery efficiency. As expected, the Reactive brown 10-produced PAT was enzymatically active. Other applications of the method on protein expression and purification, and development of PAT enzymatic inhibitors were also discussed.

Long-term outcomes of Group D eyes in bilateral retinoblastoma patients treated with chemoreduction and low-dose IMRT salvage.

BACKGROUND: To evaluate outcomes of Group D eyes of bilateral retinoblastoma patients treated with primary chemoreduction and external beam radiation as salvage. PROCEDURE: Retrospective chart review of patients diagnosed with bilateral retinoblastoma and designated Group D in at least one eye from January 1, 2000 to December 31, 2009. Overall, 62 Group D eyes of 49 patients were included; 13 had bilateral Group D disease. Primary chemoreduction with vincristine, etoposide, and carboplatin with local consolidation was administered, followed by external beam radiation in the form of intensity-modulated radiation therapy (IMRT) as salvage for recurrent tumor. Primary outcome measure was globe salvage. RESULTS: Of 62 Group D eyes, 7 were enucleated primarily; 55 were treated with systemic chemoreduction, and local therapy. Chemoreduction cured 26 of 55 eyes (47%). Recurrences were found in 29 eyes; 5 underwent enucleation and 24 were treated with IMRT at a dose of 24 Gy (2 eyes) or 36 Gy (22 eyes). Of the 24 irradiated eyes, 19 (79%) were salvaged and 5 required enucleation. Final visual acuity ranged from 20/20 to light perception with 10 eyes having 20/80 or better visual acuity. Average follow-up was 54.2 months. CONCLUSION: Kaplan-Meier estimates of eye survival of Group D eyes in bilateral patients at 12 months is 82% (95% confidence interval [CI] 70.1-89.7%); at 60 months eye survival is estimated to be 68% (95% confidence interval [CI] 55.4-82.8%). Systemic treatment for retinoblastoma demonstrated a high rate of globe preservation with acceptable complications and many eyes retaining functional vision.