Building a sustainable model of integrated stem education: investigating secondary school STEM classes after an integrated STEM project.

This study investigated the sustainability of an integrated STEM education program. Two US high school science teachers and an engineering technology teacher sustained implementation of an integrated STEM curriculum after the conclusion of the funded program, TRAILS. Class observations were conducted to examine how the teachers implemented the integrated STEM curriculum and how they maintained integrated STEM teaching in a science and engineering technology education (ETE) teacher pair using science and engineering technology shared practices. After the integrated STEM lesson, their students' academic achievements were compared to those of the students who previously participated in the project. The results reveal that the students showed no difference from the previous TRAILS students in terms of academic achievements as measured by STEM knowledge test scores, which may indicate that the teachers successfully maintained consistency and effectiveness of the implementation. Additionally, a twenty-first century skills survey was newly conducted to examine students' growth in confidence in twenty-first century skills after they were taught the integrated STEM lesson. The students showed increased confidence in critical thinking, which indicates that the students benefitted from the teachers' instructions even after the conclusion of the funded program and the absence of support. Based on the findings from the teachers' experiences of multiple years of integrated STEM teaching, the study discusses how to better support teachers for the successful implementation of an integrated STEM curriculum as a sustainable education program in secondary schools.

Deciding where to attend: Large-scale network mechanisms underlying attention and intention revealed by graph-theoretic analysis.

The neural mechanisms by which intentions are transformed into actions remain poorly understood. We investigated the network mechanisms underlying spontaneous voluntary decisions about where to focus visual-spatial attention (willed attention). Graph-theoretic analysis of two independent datasets revealed that regions activated during willed attention form a set of functionally-distinct networks corresponding to the frontoparietal network, the cingulo-opercular network, and the dorsal attention network. Contrasting willed attention with instructed attention (where attention is directed by external cues), we observed that the dorsal anterior cingulate cortex was allied with the dorsal attention network in instructed attention, but shifted connectivity during willed attention to interact with the cingulo-opercular network, which then mediated communications between the frontoparietal network and the dorsal attention network. Behaviorally, greater connectivity in network hubs, including the dorsolateral prefrontal cortex, the dorsal anterior cingulate cortex, and the inferior parietal lobule, was associated with faster reaction times. These results, shown to be consistent across the two independent datasets, uncover the dynamic organization of functionally-distinct networks engaged to support intentional acts.

Concurrent detection of targeted copy number variants and mutations using a myeloid malignancy next generation sequencing panel allows comprehensive genetic analysis using a single testing strategy.

Currently, comprehensive genetic testing of myeloid malignancies requires multiple testing strategies with high costs. Somatic mutations can be detected by next generation sequencing (NGS) but copy number variants (CNVs) require cytogenetic methods including karyotyping, fluorescence in situ hybidization and microarray. Here, we evaluated a new method for CNV detection using read depth data derived from a targeted NGS mutation panel. In a cohort of 270 samples, we detected pathogenic mutations in 208 samples and targeted CNVs in 68 cases. The most frequent CNVs were 7q deletion including LUC7L2 and EZH2, TP53 deletion, ETV6 deletion, gain of RAD21 on 8q, and 5q deletion, including NSD1 and NPM1. We were also able to detect exon-level duplications, including so-called KMT2A (MLL) partial tandem duplication, in 9 cases. In the 63 cases that were negative for mutations, targeted CNVs were observed in 4 cases. Targeted CNV detection by NGS had very high concordance with single nucleotide polymorphism microarray, the current gold standard. We found that ETV6 deletion was strongly associated with TP53 alterations and 7q deletion was associated with mutations in TP53, KRAS and IDH1. This proof-of-concept study demonstrates the feasibility of using the same NGS data to simultaneously detect both somatic mutations and targeted CNVs.

Targeted next-generation sequencing identifies a subset of idiopathic hypereosinophilic syndrome with features similar to chronic eosinophilic leukemia, not otherwise specified.

The distinction between chronic eosinophilic leukemia, not otherwise specified and idiopathic hypereosinophilic syndrome largely relies on clonality assessment. Prior to the advent of next-generation sequencing, clonality was usually determined by cytogenetic analysis. We applied targeted next-generation sequencing panels designed for myeloid neoplasms to bone marrow specimens from a cohort of idiopathic hypereosinophilic syndrome patients (n=51), and assessed the significance of mutations in conjunction with clinicopathological features. The findings were further compared with those of 17 chronic eosinophilic leukemia, not otherwise specified patients defined by their abnormal cytogenetics and/or increased blasts. Mutations were detected in 14/51 idiopathic hypereosinophilic syndrome patients (idiopathic hypereosinophilic syndrome/next-generation sequencing-positive) (28%), involving single gene in 7 and ≥2 in 7 patients. The more frequently mutated genes included ASXL1 (43%), TET2 (36%), EZH2 (29%), SETBP1 (22%), CBL (14%), and NOTCH1 (14%). Idiopathic hypereosinophilic syndrome/next-generation sequencing-positive patients showed a number of clinical features and bone marrow findings resembling chronic eosinophilic leukemia, not otherwise specified. Chronic eosinophilic leukemia, not otherwise specified patients showed a disease-specific survival of 14.4 months, markedly inferior to idiopathic hypereosinophilic syndrome/next-generation sequencing-negative (P<0.001), but not significantly different from idiopathic hypereosinophilic syndrome/next-generation sequencing-positive (P=0.117). These data suggest that targeted next-generation sequencing helps to establish clonality in a subset of patients with hypereosinophilia that would otherwise be classified as idiopathic hypereosinophilic syndrome. In conjunction with other diagnostic features, mutation data can be used to establish a diagnosis of chronic eosinophilic leukemia, not otherwise specified in patients presenting with hypereosinophilia.

Detection of BCR-ABL1 mutations that confer tyrosine kinase inhibitor resistance using massively parallel, next generation sequencing.

Detection of BCR-ABL1 mutations that confer resistance to tyrosine kinase inhibitors is important for management of patients with t(9;22);BCR-ABL1-positive (Ph+) leukemias. Testing is often performed using Sanger sequencing (SS) which has relatively poor sensitivity. Given the widespread adoption of next generation sequencing (NGS), we sought to reevaluate the testing in the context of NGS methods. We developed an NGS-based BCR-ABL1 mutation test on the Ion Torrent Personal Genome Machine (PGM) to test for resistance mutations, primarily in the kinase domain in BCR-ABL1. We analyzed 508 clinical samples from patients with Ph+ leukemias. In a subset of these samples (n = 97), we conducted a comparison of the NGS results to a classical SS-based test. NGS facilitated detection of low-level mutations (<20 % allele frequency) that were not detectable by SS. In a subset of cases with multiple mutations, NGS was also able to determine if two mutations were on the same molecule (compound) or on separate molecules (polyclonal) but this was limited by the distance between mutated positions and by the effects of apparent distance-dependent PCR recombination. We found 22 compound mutations that centered on one or two key residues including two novel compound mutants: Q252H/Y253H and F311Y/F359I. The advantages of NGS make it a superior method for inventorying BCR-ABL1 resistance mutations. However, data analysis may be complicated by short read lengths and the effects of PCR recombination.

The neural correlates of volitional attention: A combined fMRI and ERP study.

Studies of visual-spatial attention typically use instructional cues to direct attention to a relevant location, but in everyday vision, attention is often focused volitionally, in the absence of external signals. Although investigations of cued attention comprise hundreds of behavioral and physiological studies, remarkably few studies of voluntary attention have addressed the challenging question of how spatial attention is initiated and controlled in the absence of external instructions, which we refer to as willed attention. To explore this question, we employed a trial-by-trial spatial attention task using electroencephalography and functional magnetic resonance imaging (fMRI). The fMRI results reveal a unique network of brain regions for willed attention that includes the anterior cingulate cortex, left middle frontal gyrus (MFG), and the left and right anterior insula (AI). We also observed two event-related potentials (ERPs) associated with willed attention; one with a frontal distribution occurring 250-350 ms postdecision cue onset (EWAC: Early Willed Attention Component), and another occurring between 400 and 800 ms postdecision-cue onset (WAC: Willed Attention Component). In addition, each ERP component uniquely correlated across subjects with different willed attention-specific sites of BOLD activation. The EWAC was correlated with the willed attention-specific left AI and left MFG activations and the later WAC was correlated only with left AI. These results offer a comprehensive and novel view of the electrophysiological and anatomical profile of willed attention and further illustrate the relationship between scalp-recorded ERPs and the BOLD response.

BCR-ABL1 compound mutations in tyrosine kinase inhibitor-resistant CML: frequency and clonal relationships.

BCR-ABL1 compound mutations can confer high-level resistance to imatinib and other ABL1 tyrosine kinase inhibitors (TKIs). The third-generation ABL1 TKI ponatinib is effective against BCR-ABL1 point mutants individually, but remains vulnerable to certain BCR-ABL1 compound mutants. To determine the frequency of compound mutations among chronic myeloid leukemia patients on ABL1 TKI therapy, in the present study, we examined a collection of patient samples (N = 47) with clear evidence of 2 BCR-ABL1 kinase domain mutations by direct sequencing. Using a cloning and sequencing method, we found that 70% (33/47) of double mutations detected by direct sequencing were compound mutations. Sequential, branching, and parallel routes to compound mutations were common. In addition, our approach revealed individual and compound mutations not detectable by direct sequencing. The frequency of clones harboring compound mutations with more than 2 missense mutations was low (10%), whereas the likelihood of silent mutations increased disproportionately with the total number of mutations per clone, suggesting a limited tolerance for BCR-ABL1 kinase domain missense mutations. We conclude that compound mutations are common in patients with sequencing evidence for 2 BCR-ABL1 mutations and frequently reflect a highly complex clonal network, the evolution of which may be limited by the negative impact of missense mutations on kinase function.

A novel approach to quantitating leukemia fusion transcripts by qRT-PCR without the need for standard curves.

Acute myeloid leukemia patients with recurrent cytogenetic abnormalities including inv(16);CBFB-MYH11 and t(15;17);PML-RARA may be assessed by monitoring the levels of the corresponding abnormal fusion transcripts by quantitative reverse transcription-PCR (qRT-PCR). Such testing is important for evaluating the response to therapy and for the detection of early relapse. Existing qRT-PCR methods are well established and in widespread use in clinical laboratories but they are laborious and require the generation of standard curves. Here, we describe a new method to quantitate fusion transcripts in acute myeloid leukemia by qRT-PCR without the need for standard curves. Our approach uses a plasmid calibrator containing both a fusion transcript sequence and a reference gene sequence, representing a perfect normalized copy number (fusion transcript copy number/reference gene transcript copy number; NCN) of 1.0. The NCN of patient specimens can be calculated relative to that of the single plasmid calibrator using experimentally derived PCR efficiency values. We compared the data obtained using the plasmid calibrator method to commercially available assays using standard curves and found that the results obtained by both methods are comparable over a broad range of values with similar sensitivities. Our method has the advantage of simplicity and is therefore lower in cost and may be less subject to errors that may be introduced during the generation of standard curves.

T-cell clonality assessment by next-generation sequencing improves detection sensitivity in mycosis fungoides.

BACKGROUND: T-cell receptor (TCR) clonality assessment is a principal diagnostic test in the management of mycosis fungoides (MF). However, current polymerase chain reaction-based methods may produce ambiguous results, often because of low abundance of clonal T lymphocytes, resulting in weak clonal peaks that cannot be size-resolved by contemporary capillary electrophoresis (CE). OBJECTIVE: We sought to determine if next-generation sequencing (NGS)-based detection has increased sensitivity for T-cell clonality over CE-based detection in MF. METHODS: Clonality was determined by an NGS-based method in which the TCR-γ variable region was polymerase chain reaction amplified and the products sequenced to establish the identity of rearranged variable and joining regions. RESULTS: Of the 35 MF cases tested, 29 (85%) showed a clonal T-cell rearrangement by NGS, compared with 15 (44%) by standard CE detection. Three patients with MF had follow-up testing that showed identical, clonal TCR sequences in subsequent skin biopsy specimens. LIMITATIONS: Clonal T-cell populations have been described in benign conditions; evidence of clonality alone, by any method, is not sufficient for diagnosis. CONCLUSION: TCR clonality assessment by NGS has superior sensitivity compared with CE-based detection. Further, NGS enables tracking of specific clones across multiple time points for more accurate identification of recurrent MF.

Spontaneous neural fluctuations predict decisions to attend.

Ongoing variability in neural signaling is an intrinsic property of the brain. Often this variability is considered to be noise and ignored. However, an alternative view is that this variability is fundamental to perception and cognition and may be particularly important in decision-making. Here, we show that a momentary measure of occipital alpha-band power (8-13 Hz) predicts choices about where human participants will focus spatial attention on a trial-by-trial basis. This finding provides evidence for a mechanistic account of decision-making by demonstrating that ongoing neural activity biases voluntary decisions about where to attend within a given moment.

Use of tranexamic acid results in decreased blood loss and decreased transfusions in patients undergoing staged bilateral total knee arthroplasty.

BACKGROUND: Tranexamic acid (TXA) is an antifibrinolytic that reduces blood loss and transfusion rates in total joint arthroplasty. Blood loss and allogenic transfusion rates have not been well studied in patients receiving TXA and undergoing bilateral staged total knee arthroplasty (TKA). The purpose was to evaluate the effect of TXA on blood loss, hemoglobin (Hb) changes, and transfusion in patients undergoing staged bilateral TKA. STUDY DESIGN AND METHODS: The authors compared 51 patients undergoing staged bilateral TKA who received TXA (2 g; subjects) with 70 who did not (controls). There were no significant differences between the groups in terms of demographics or preoperative Hb. For each TKA, 1 g of TXA was administered intravenously 15 minutes before incision and 1 g was administered intravenously at tourniquet release. Blood loss, Hb levels, and transfusions were recorded. Statistical analyses were performed using computer software. Significance was set at 0.05. RESULTS: Subjects had a significantly lower (p < 0.001) mean (±SD) blood loss (373.8 ± 264.6 mL vs. 871.6 ± 457.7 mL), significantly higher (p < 0.005) Hb levels on Postoperative Days 1 and 2, and a significantly lower (p < 0.001) mean (±SD) number of transfused allogenic blood units (0.60 ± 0.84 units vs. 1.53 ± 1.30 units). CONCLUSIONS: TXA reduces blood loss, improves postoperative Hb, and decreases the allogenic blood transfusion requirements for patients undergoing bilateral staged TKA. TXA is an option for patients choosing bilateral staged TKA to decrease the risks associated with blood transfusion or when autologous blood is not available.

The impact of distractor congruency on stimulus processing in retinotopic visual cortex.

The brain is frequently confronted with sensory information that elicits conflicting response choices. While much research has addressed the top down control mechanisms associated with detection and resolution of response competition, the effects of response competition on sensory processing in the primary visual cortex remain unclear. To address this question we modified a typical 'flanker task' (Eriksen and Eriksen, 1974) so that the effects of response competition on human early retinotopic visual cortex could be assessed. Healthy human participants were scanned using fMRI while making a speeded choice response that classified a target object image into one of two categories (e.g. fruits, animals). An irrelevant distractor image that was either congruent (same image as target), incongruent (image from opposite category as target), or neutral (image from task-irrelevant category, e.g. household items) was also present on each trial, but in a different quadrant of the visual field relative to the target. Retinotopic V1 areas responding to the target stimuli showed increased response to targets in the presence of response-incongruent (compared to response-neutral) distractors. A negative correlation with behavioral response competition effects indicated that an increased primary visual cortical response to targets in the incongruent (vs. neutral) trials is associated with a reduced response competition effect on behavior. These results suggest a novel conflict resolution mechanism in the primary visual cortex.

Efficacy of multimodal perioperative analgesia protocol with periarticular medication injection in total knee arthroplasty: a randomized, double-blinded study.

Pain control is necessary for successful rehabilitation and outcome after total knee arthroplasty. Our goal was to compare the clinical efficacy of periarticular injections consisting of a long-acting local anesthetic (ropivacaine) and epinephrine with and without combinations of an α2-adrenergic agonist (clonidine) and/or a nonsteroidal anti-inflammatory agent (ketorolac). In a double-blinded controlled study, we randomized 160 patients undergoing total knee arthroplasty to receive 1 of 4 intraoperative periarticular injections: Group A, ropivacaine, epinephrine, ketorolac, and clonidine; Group B, ropivacaine, epinephrine, and ketorolac; Group C, ropivacaine, epinephrine, and clonidine; Group D (control), ropivacaine and epinephrine. Compared with Group D, Group A and B patients had significantly lower postoperative visual analog pain scores and nurse pain assessment and Group C patients had a significantly greater reduction in physical therapist pain assessment. We found no differences in other parameters analyzed.

Medial UKA: favorable mid-term results in middle-aged patients.

In middle-aged patients with knee arthritis, surgical treatment options include arthroscopic procedures, osteotomies, and unicompartmental and total knee arthroplasty. Unicompartmental knee osteoarthrosis is particularly challenging and controversial in such patients. From December 2001 through October 2005, we treated 32 consecutive middle-aged patients (46 to 59 years old) with 40 medial unicompartmental knee arthroplasties. Three patients were lost to follow-up, leaving 29 for our study. There were two reoperations: one for loosening at 3 years, and one for disease progression at 5 years. Implant survival was modeled using Kaplan-Meier survival function with observations censored if lost to follow-up. At the 6-year survivorship in this group, the overall implant survival rate was 94.1% (95% CI, 78.3-98.5). Knee Society scores had improved significantly (p < 0.001). There were no other instances of osteolysis or radiographic failure. We concluded that, at mid-term follow-up, unicompartmental knee arthroplasty provided satisfactory results for this challenging population.

Targeted next generation sequencing of clinically significant gene mutations and translocations in leukemia.

Leukemias are currently subclassified based on the presence of recurrent cytogenetic abnormalities and gene mutations. These molecular findings are the basis for risk-adapted therapy; however, such data are generally obtained by disparate methods in the clinical laboratory, and often rely on low-resolution techniques such as fluorescent in situ hybridization. Using targeted next generation sequencing, we demonstrate that the full spectrum of prognostically significant gene mutations including translocations, single nucleotide variants (SNVs), and insertions/deletions (indels) can be identified simultaneously in multiplexed sequence data. As proof of concept, we performed hybrid capture using a panel of 20 genes implicated in leukemia prognosis (covering a total of 1 Mbp) from five leukemia cell lines including K562, NB4, OCI-AML3, kasumi-1, and MV4-11. Captured DNA was then sequenced in multiplex on an Illumina HiSeq. Using an analysis pipeline based on freely available software we correctly identified DNA-level translocations in three of the three cell lines where translocations were covered by our capture probes. Furthermore, we found all published gene mutations in commonly tested genes including NPM1, FLT3, and KIT. The same methodology was applied to DNA extracted from the bone marrow of a patient with acute myeloid leukemia, and identified a t(9;11) translocation with single base accuracy as well other gene mutations. These results indicate that targeted next generation sequencing can be successfully applied in the clinical laboratory to identify a full spectrum of DNA mutations ranging from SNVs and indels to translocations. Such methods have the potential to both greatly streamline and improve the accuracy of DNA-based diagnostics.

Working memory load modulates distractor competition in primary visual cortex.

When stimuli compete for sensory processing and response selection, coherent goal-guided behavior requires cognitive control so that task-relevant "targets" rather than irrelevant distractors are selected. It has been shown that reduced cognitive control under high working memory load increases distractor competition for selection. It remains unknown, though, whether cognitive control by working memory has an effect on the earliest levels of sensory processing in primary visual cortex. The present study addressed this question by having subjects perform a selective attention task involving classification of meaningful target objects while also ignoring congruent and incongruent distractor images. The level of cognitive control over distractor competition was varied through a concurrent working memory task of either low (1 digit) or high (6 digits) load. Functional magnetic resonance imaging revealed greater distractor competition effects not only on behavior but also on the sensory correlates in primary visual cortex (areas V1-V2) in conditions of high (vs. low) working memory load. In addition, high working memory load resulted in increased congruency-related functional connectivity between anterior cingulate cortex and V1. These results are the first to establish the neural correlates of distractor competition effects in primary visual cortex and the critical role of working memory in their cognitive control.

All-polyethylene tibial components in obese patients are associated with low failure at midterm followup.

BACKGROUND: In the United States, the obese population has increased markedly over the last four decades, and this trend continues. High patient weight places additional stress on TKA components, which may lead to increased polyethylene wear, osteolysis, radiolucencies, and clinical failure. Metal-backed tibial components and all-polyethylene tibial components in the general population have comparable osteolysis and failure, but it is unclear whether these components yield similar osteolysis and failure in obese patients. QUESTIONS/PURPOSES: We therefore determined the (1) function, (2) occurrence of osteolysis, and (3) complications in a cohort of obese patients receiving all-polyethylene tibial components. PATIENTS AND METHODS: Between September 17, 1996, and December 19, 2002, we implanted all-polyethylene tibial components in 90 obese patients (125 knees); 24 patients (33 knees) died and 13 patients (17 knees) were lost to followup, leaving 53 patients (59%) with 75 knees. All surgeries were cruciate-retaining, tricompartmental TKAs. We evaluated patients with Knee Society Scores and serial radiographs. Minimum followup was 7 years (mean, 10.4 years; range, 7-14 years). RESULTS: At latest followup, mean Knee Society Score was 92 points. There were five tibial radiolucencies, all less than 1 mm and characterized as nonprogressive. We observed minimal, nonprogressive osteolysis in one knee. One patient required reoperation after a traumatic event. There were no implant-related failures and no implants at risk of failure. CONCLUSIONS: At an average 10-year followup, all-polyethylene tibial components were functioning well in this obese group. These findings confirm the effectiveness of all-polyethylene tibial components in obese patients.

Modern proximally tapered uncemented stems can be safely used in Dorr type C femoral bone.

Cementless femoral fixation has become widely accepted in modern total hip arthroplasty. Treating patients who have a stovepipe-shaped femur (Dorr type C) with cementless implants has traditionally been challenging. We treated 53 consecutive patients (60 hips) who had type C bone with identical tapered, proximally coated implants and postoperative weight bearing as tolerated. At 6 weeks, all 60 hips had radiographically documented bony integration, and at 1 year, there was no evidence of fracture, subsidence, thigh pain, stress shielding, loose stems, or risk of failure. Of those patients, 40 (43 hips) had midterm follow-up (average, 6 years; range, 4-9 years); the findings were the same. We conclude that modern proximally tapered stems can be used with early weight bearing in patients with type C bone.

Enhanced visual perception with occipital transcranial magnetic stimulation.

Transcranial magnetic stimulation (TMS) over the occipital pole can produce an illusory percept of a light flash (or 'phosphene'), suggesting an excitatory effect. Whereas previous reported effects produced by single-pulse occipital pole TMS are typically disruptive, here we report the first demonstration of a location-specific facilitatory effect on visual perception in humans. Observers performed a spatial cueing orientation discrimination task. An orientation target was presented in one of two peripheral placeholders. A single pulse below the phosphene threshold applied to the occipital pole 150 or 200 ms before stimulus onset was found to facilitate target discrimination in the contralateral compared with the ipsilateral visual field. At the 150-ms time window contralateral TMS also amplified cueing effects, increasing both facilitation effects for valid cues and interference effects for invalid cues. These results are the first to show location-specific enhanced visual perception with single-pulse occipital pole stimulation prior to stimulus presentation, suggesting that occipital stimulation can enhance the excitability of visual cortex to subsequent perception.