Translation is the fundamental biological process converting mRNA information into proteins. Single-molecule imaging in live cells has illuminated the dynamics of RNA transcription; however, it is ...not yet applicable to translation. Here, we report single-molecule imaging of nascent peptides (SINAPS) to assess translation in live cells. The approach provides direct readout of initiation, elongation, and location of translation. We show that mRNAs coding for endoplasmic reticulum (ER) proteins are translated when they encounter the ER membrane. Single-molecule fluorescence recovery after photobleaching provides direct measurement of elongation speed (5 amino acids per second). In primary neurons, mRNAs are translated in proximal dendrites but repressed in distal dendrites and display "bursting" translation. This technology provides a tool with which to address the spatiotemporal translation mechanism of single mRNAs in living cells.
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BFBNIB, NMLJ, NUK, ODKLJ, PNG, SAZU, UL, UM, UPUK
A rational approach is needed to design hydrogénation catalysts that make use of Earth-abundant elements to replace the rare elements such as ruthenium, rhodium, and palladium that are traditionally ...used. Here, we validate a prior mechanistic hypothesis that partially saturated amine(imine) diphosphine ligands (P-NH-N-P) activate iron to catalyze the asymmetric reduction of the polar bonds of ketones and imines to valuable enantiopure alcohols and amines, with isopropanol as the hydrogen donor, at turnover frequencies as high as 200 per second at 28°C. We present a direct synthetic approach to enantiopure ligands of this type that takes advantage of the iron(ll) ion as a template. The catalytic mechanism is elucidated by the spectroscopic detection of iron hydride and amide intermediates.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Sex cord-stromal tumours of the ovary include many of the most morphologically intriguing ovarian neoplasms and albeit many of them are rare, they factor into the differential diagnosis more often ...than their frequency might suggest. The most common malignant form, the adult granulosa cell tumour, may grossly simulate various surface epithelial neoplasms. Microscopically, confusion with endometrioid carcinoma may occur because the cords and microfollicles of the granulosa cell tumour may be mimicked by endometrioid carcinoma and the latter may have pale nuclei with nuclear grooves. Thorough sampling generally resolves this differential and if not immunohistochemistry aids. Although the adult granulosa cell tumour typically has cells with scant cytoplasm in some cases the tumour cells are luteinised and others have cells with abundant pale cytoplasm. A reticulum stain may be of great aid in indicating whether cells of the type just noted are of granulosa or theca nature. Variations in the morphology of the juvenile variant of granulosa cell tumour that can be diagnostically challenging include those that have a macronodular pattern with scant follicular differentiation, those with marked sclerosis, and those that are unusually pleomorphic. The uncommon but histologically varied Sertoli–Leydig cell tumour is considered, emphasis being placed on the most recently described variant, the retiform pattern, with its potential to mimic surface epithelial neoplasms and even mixed mesodermal tumours. Considering the usual young age of the patient may be paramount in making this tumour come to the mind of the pathologist. The rare pure Sertoli cell tumour is briefly noted as is the sex cord tumour with annular tubules, well known because of its association in some cases with Peutz–Jeghers syndrome. Most do not have that association, however, but have their own interesting features including a greater than average risk, among sex cord stromal tumours, of nodal metastasis and progesterone production, and an occasional development from them of an otherwise typical Sertoli cell tumour. The stromal family includes the common fibroma which is challenging when it is cellular with some mitotic activity and the approach to such neoplasms is reviewed. Emphasis in the consideration of thecoma is placed on its typical cytological features and the overlap with what may be seen in some adult granulosa cell tumours. The review concludes with three fascinating pure stromal tumours all described within the last several decades: the sclerosing stromal tumour, the unusual luteinised thecoma associated with sclerosing peritonitis and the microcystic stromal tumour. The first is sometimes misdiagnosed when pure stromal neoplasms of other types are vascular and may have pseudolobules and it is essential that the pseudolobules of the sclerosing stromal tumour contain a haphazard admixture of fibroblasts and weakly luteinised cells. The remarkable tumours associated with peritonitis exhibit brisk mitotic activity but appear not to have a metastatic potential; they can cause significant problems because of the sclerosing peritonitis. The microcystic stromal tumour may mimic a steroid cell tumour or thecoma but unlike them is inhibin and calretinin negative, and stains for CD10 and β-catenin. It often shows bizarre nuclei atypia but limited mitotic activity and appears to be clinically benign on the basis of still limited experience.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Chimeric antigen receptors (CAR) are synthetic molecules that provide new specificities to T cells. Although successful in treatment of hematologic malignancies, CAR T cells are ineffective for solid ...tumors to date. We found that the cell-surface molecule c-Met was expressed in ∼50% of breast tumors, prompting the construction of a CAR T cell specific for c-Met, which halted tumor growth in immune-incompetent mice with tumor xenografts. We then evaluated the safety and feasibility of treating metastatic breast cancer with intratumoral administration of mRNA-transfected c-Met-CAR T cells in a phase 0 clinical trial (NCT01837602). Introducing the CAR construct via mRNA ensured safety by limiting the nontumor cell effects (on-target/off-tumor) of targeting c-Met. Patients with metastatic breast cancer with accessible cutaneous or lymph node metastases received a single intratumoral injection of 3 × 10
or 3 × 10
cells. CAR T mRNA was detectable in peripheral blood and in the injected tumor tissues after intratumoral injection in 2 and 4 patients, respectively. mRNA c-Met-CAR T cell injections were well tolerated, as none of the patients had study drug-related adverse effects greater than grade 1. Tumors treated with intratumoral injected mRNA c-Met-CAR T cells were excised and analyzed by immunohistochemistry, revealing extensive tumor necrosis at the injection site, cellular debris, loss of c-Met immunoreactivity, all surrounded by macrophages at the leading edges and within necrotic zones. We conclude that intratumoral injections of mRNA c-Met-CAR T cells are well tolerated and evoke an inflammatory response within tumors.
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Interception of potential invasive species at ports-of-entry is essential for effective biosecurity and biosurveillance programs. However, taxonomic assessment of the immature stages of most ...arthropods is challenging; characters for identification are often dependent on adult morphology and reproductive structures. This study aims to strengthen the identification of such specimens through DNA barcoding, with a focus on microlepidoptera. A sample of 241 primarily immature microlepidoptera specimens intercepted at U.S. ports-of-entry from 2007 to 2011 were selected for analysis. From this sample, 201 COI-5P sequences were generated and analyzed for concordance between morphology-based and DNA-based identifications. The retrospective analysis of the data over 10 years (2009 to 2019) using the Barcode of Life Data (BOLD) system demonstrates the importance of establishing and growing DNA barcode reference libraries for use in specimen identification. Additionally, analysis of specimen identification using public data (43.3% specimens identified) vs. non-public data (78.6% specimens identified) highlights the need to encourage researchers to make data publicly accessible. DNA barcoding surpassed morphological identification with 42.3% (public) and 66.7% (non-public) of the sampled specimens achieving a species-level identification, compared to 38.3% species-level identification by morphology. Whilst DNA barcoding was not able to identify all specimens in our dataset, its incorporation into border security programs as an adjunct to morphological identification can provide secondary lines of evidence and lower taxonomic resolution in many cases. Furthermore, with increased globalization, database records need to be clearly annotated for suspected specimen origin versus interception location.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
IMPORTANCE: Remdesivir demonstrated clinical benefit in a placebo-controlled trial in patients with severe coronavirus disease 2019 (COVID-19), but its effect in patients with moderate disease is ...unknown. OBJECTIVE: To determine the efficacy of 5 or 10 days of remdesivir treatment compared with standard care on clinical status on day 11 after initiation of treatment. DESIGN, SETTING, AND PARTICIPANTS: Randomized, open-label trial of hospitalized patients with confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and moderate COVID-19 pneumonia (pulmonary infiltrates and room-air oxygen saturation >94%) enrolled from March 15 through April 18, 2020, at 105 hospitals in the United States, Europe, and Asia. The date of final follow-up was May 20, 2020. INTERVENTIONS: Patients were randomized in a 1:1:1 ratio to receive a 10-day course of remdesivir (n = 197), a 5-day course of remdesivir (n = 199), or standard care (n = 200). Remdesivir was dosed intravenously at 200 mg on day 1 followed by 100 mg/d. MAIN OUTCOMES AND MEASURES: The primary end point was clinical status on day 11 on a 7-point ordinal scale ranging from death (category 1) to discharged (category 7). Differences between remdesivir treatment groups and standard care were calculated using proportional odds models and expressed as odds ratios. An odds ratio greater than 1 indicates difference in clinical status distribution toward category 7 for the remdesivir group vs the standard care group. RESULTS: Among 596 patients who were randomized, 584 began the study and received remdesivir or continued standard care (median age, 57 interquartile range, 46-66 years; 227 39% women; 56% had cardiovascular disease, 42% hypertension, and 40% diabetes), and 533 (91%) completed the trial. Median length of treatment was 5 days for patients in the 5-day remdesivir group and 6 days for patients in the 10-day remdesivir group. On day 11, patients in the 5-day remdesivir group had statistically significantly higher odds of a better clinical status distribution than those receiving standard care (odds ratio, 1.65; 95% CI, 1.09-2.48; P = .02). The clinical status distribution on day 11 between the 10-day remdesivir and standard care groups was not significantly different (P = .18 by Wilcoxon rank sum test). By day 28, 9 patients had died: 2 (1%) in the 5-day remdesivir group, 3 (2%) in the 10-day remdesivir group, and 4 (2%) in the standard care group. Nausea (10% vs 3%), hypokalemia (6% vs 2%), and headache (5% vs 3%) were more frequent among remdesivir-treated patients compared with standard care. CONCLUSIONS AND RELEVANCE: Among patients with moderate COVID-19, those randomized to a 10-day course of remdesivir did not have a statistically significant difference in clinical status compared with standard care at 11 days after initiation of treatment. Patients randomized to a 5-day course of remdesivir had a statistically significant difference in clinical status compared with standard care, but the difference was of uncertain clinical importance. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT04292730
•Composition of mRNPs is dynamic across space and time.•mRNPs in dendrites and axons share common features of diffusion and directed transport.•HMM Bayes approach is a useful tool to analyze mRNA ...transport dynamics.•mRNA localization and local translation in response to synaptic activity are important for long-lasting plasticity.
Neurons are highly polarized cells that can extend processes far from the cell body. As such, transport of messenger RNAs serves as a set of blueprints for the synthesis of specific proteins at distal sites. RNA localization to dendrites and axons confers the ability to regulate translation with extraordinary precision in space and time. Although the rationale for RNA localization is quite compelling, it is unclear how a neuron orchestrates such a complex task of distributing over a thousand different mRNAs to their respective subcellular compartments. Recent single-molecule imaging studies have led to insights into the kinetics of individual mRNAs. We can now peer into the transport dynamics of mRNAs in both dendrites and axons.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The author reviews highlights of advances in knowledge concerning ovarian tumor pathology since the time of an essay in the first issue of this Journal written by Dr Robert E. Scully, who, both ...before and for several decades after the Journal was instituted, made many original contributions to the field and was the major architect of the 1973 World Health Organization classification of ovarian tumors which was much more clear and logical than prior ones. The current review considers the neoplasms in essentially the same order as was done in the first issue of this journal and presents a personal look at the highlights of new information concerning various well-known categories, surface epithelial, germ cell, sex cord-stromal, metastatic neoplasms and briefly, benign so-called tumor-like lesions. Some of the most notable developments are as follows: (1) an orderly approach to the classification of implants of serous borderline tumors into noninvasive and invasive categories; (2) recognition of distinctive micropapillary patterns seen in some borderline tumors and low-grade carcinomas; (3) a remarkable propensity for some endometrioid carcinomas to mimic sex cord- stromal tumors; (4) appreciation of the differences between primary mucinous tumors of intestinal and müllerian types; (5) the importance of distinguishing within primary mucinous carcinomas between expansile and destructive stromal invasion; (6) emphasis on the diagnosis of immature teratoma being based on the presence of primitive–embryonic–appearing tissues; (7) appreciation of variant morphology of cases of struma ovarii which may lead to significant diagnostic problems; (8) subdivision of granulosa cell tumors into adult and juvenile types because of the differing features of the two groups including in the second category the propensity for more malignant neoplasms to be mimicked; (9) recognition of a distinctive form of Sertoli-Leydig cell tumor, the retiform variant, with a propensity to occur in the young; (10) appreciation of a unique highly malignant neoplasm that typically afflicts the young and may be associated with hypercalcemia, so-called small cell carcinoma of hypercalcemic type; (11) greater awareness than was hitherto the case of the propensity for metastatic intestinal adenocarcinoma to mimic primary endometrioid carcinoma and similarly for metastatic mucinous carcinomas to simulate primary mucinous cystic tumors; (12) recognition of the distinctive features of low-grade appendiceal mucinous neoplasms that spread to the ovary and are typically associated with pseudomyxoma peritonei; and (13) appreciation that the histologic spectrum seen in cases of Krukenberg tumor is broader than often previously thought.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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