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To present a study that investigated sacrococcygeal skeletal structure as a possible nonmodifiable intrinsic risk factor for pressure injury and identify possible issues caused by its morphology.This continuing education activity is intended for physicians, physician assistants, nurse practitioners, and nurses with an interest in skin and wound care.After participating in this educational activity, the participant will:1. Recognize the background information the authors considered when planning and conducting their study of sacrococcygeal skeletal structure as a possible pressure injury risk factor.2. Identify the characteristics of the two groups of study participants.3. Choose the results of the study clinicians may consider when implementing evidence-based practice.To determine if sacrococcygeal skeletal morphology and morphometry characteristics were possible pressure injury (PI) intrinsic risk factors; determine the exact location of these PIs; and generate hypotheses and determine methodological considerations required for future larger studies. This case-control pilot study compared 30 patients who had an MRI scan—15 patients had a PI and 15 patients did not. Key sacrococcygeal morphology and morphometry parameters were assessed. On average, patients with PIs had less of a lumbosacral and sacrococcygeal angle and a greater sacral curvature and intercoccygeal angle than did patients without a PI. Patients with PIs had more variable coccyx types. Tissue and bone destruction precluded several measurements in some patients. The most common area of destruction was located distally. Sacrococcygeal measurements differed in patients with PIs, and PIs were predominately located distally. Authors recommend replicating this study on a larger scale because certain key attributes warrant further investigation to determine their influence on sacrococcygeal PIs. Sacrococcygeal morphology and morphometry parameters have not been previously studied as possible intrinsic risk factors for PIs; yet, this is the most common location for their occurrence. Knowledge regarding possible injury mechanisms due to the forces from overlying skeletal structures with respective tissue loading over the sacrococcygeal area has the potential to inform practice; preventive strategies; and equipment, products, and technology developed.

Purpose To report device-related adverse events 6 months after placement or conversion of the VenaTech convertible vena cava filter (VTCF). Materials and Methods A review of 6-month follow-up data of an investigational device exemption multicenter, prospective, single-arm study was performed. The VTCF was implanted in 149 patients. Conversion was attempted in 64.4% of those patients (n = 96) and successfully in 96.9% of the patients (n = 93). A total of 76 patients completed imaging evaluation at 6 months after filter conversion. Patients who required continued venous thromboembolism prophylaxis at 6 months did not undergo a conversion attempt and were designated as nonconverted filter subjects. A total of 28 nonconverted filter subjects completed imaging evaluation at 6 months after implantation. Results Evaluation of patients at 6 months after conversion demonstrated 1 of 76 (1.3%) inferior vena cava (IVC) perforations with a filter strut greater than 3 mm outside of the caval lumen. No cases of recurrent PE, clinically significant filter migration, filter fracture, or IVC thrombosis were reported in the converted subjects. In the nonconverted filter subjects, there was a 14.3% (4 of 28) complete or nearly complete rate of IVC thromboses. There were no cases of recurrent pulmonary embolism, penetration, fracture, or spontaneous conversion in the nonconverted filter subjects. There was a significant reduction in the rate of IVC thrombosis and migration in the converted cohort compared to that in the nonconverted cohort. Conclusions At 6 months, the VTCF demonstrated low adverse event rates in the converted configuration, whereas a minority of patients with the nonconverted configuration demonstrated a high risk of IVC thrombosis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is responsible for coronavirus disease 2019 (COVID-19), is known to cause severe respiratory infections with occasional accompanying pleural effusion (PE), pericardial effusion (PCE), or peritoneal effusion (PTE). The effect of COVID-19 on effusion cytology is not yet known. This study aimed to examine the cytomorphologic features and workup of effusion fluids in patients with active COVID-19 infection versus those in recovery.PE (n = 15), PCE (n = 1), and PTE samples (n = 20) from hospitalized patients with a SARS-CoV-2 infection (from June 1, 2020, to December 30, 2020) were reviewed. Effusion fluids with metastatic carcinoma were excluded. Differential cell counts, cytomorphology, and relevant immunostains for effusion fluids were retrospectively evaluated and compared between patients with active infection (positive on a SARS-CoV-2 nucleic acid amplification test [NAAT] within 2 months; n = 23) and those in the recovery phase from COVID-19 (negative on a SARS-CoV-2 NAAT for2 months; n = 13).The cytology diagnoses were negative for malignancy (n = 31), atypical (n = 4), and suspicious for malignancy (n = 1). Active infection cases showed more atypical mesothelial cells than recovery cases (P.05); some had enlarged nuclei, prominent nucleoli, occasional multinucleation, and bizarre nuclei. Immunostains were performed more often in active infection cases than recovery cases (47.8% vs 7.7%; P.05). Differential cell counts (available for 28 cases) showed no significant differences between the active infection and recovery groups.This study found atypical and bizarre mesothelial cells more often in effusions of cases with active COVID-19 infection in comparison with patients in recovery. It is important for cytopathologists to become familiar with the cytomorphologic effects of SARS-CoV-2 on effusion cytology so that these cases can be properly triaged.

Background: COVID-19 is a global pandemic with patients at increased risk for all-cause mortality. Virus-platelet interactions are linked to viral pathogenesis and increased risk of adverse events. Aim: To investigate the relationship between in vivo platelet activity markers and all-cause mortality in hospitalized patients with COVID-19. Method: Plasma samples were collected from 100 hospitalized patients on the day of PCR-confirmed COVID-19 diagnosis. Thromboxane B 2 (TxB 2 ), P-selectin, and soluble CD40 ligand (sCD40L) were measured in plasma, and mean platelet volume (MPV) assessed. Subjects were followed until discharge or death. Results: Among 100 patients, the median age was 65 years (IQR: 55, 75), 39% were female, and 32 died or experienced a thrombotic event. The baseline platelet activation markers, P-selectin (p=0.02), sCD40L (p=0.03) and MPV (p=0.005) were higher in patients who died. After adjustment for age, sex, race/ethnicity, platelet count, antiplatelet therapy, and chronic obstructive pulmonary disease, TxB 2 (p=0.036), P-selectin (p=0.007), sCD40L (p=0.02) and MPV (p=0.01) were each independently associated with death after multivariable adjustment ( Table 1 ). Conclusions: Biomarkers of platelet activation are significantly associated with death or thrombosis in patients hospitalized with COVID-19. Our findings suggest multiple platelet activation mechanisms may contribute to adverse events. Further investigation into the mechanistic role of platelets in COVID-19 pathogenesis and the potential role of antiplatelet therapy is warranted. Table 1. Multivariable regression models of all-cause mortality. Odds ratios (OR) from logistic regression analysis per SD increase for biomarker levels adjusted for age, sex, race, antiplatelet therapy, platelet count, and chronic obstructive pulmonary disease (COPD).

Requiring regional or in-country confirmatory clinical trials before approval of drugs already approved elsewhere delays access to medicines in low- and middle-income countries and raises drug costs. Here, we discuss the scientific and technological advances that may reduce the need for in-country or in-region clinical trials for drugs approved in other countries and limitations of these advances that could necessitate in-region clinical studies.

Human organic anion transporters (OATPs) are vital for the uptake and efflux of drugs and endogenous compounds. Current identification of inhibitors of these transporters is based on experimental screening. Virtual screening remains a challenge due to a lack of experimental three-dimensional protein structures. Here, we describe a workflow to identify inhibitors of the OATP2B1 transporter in the DrugBank library of over 5,000 drugs and druglike molecules. OATP member 2B1 transporter is highly expressed in the intestine, where it participates in oral absorption of drugs. Predictions from a Random forest classifier, prioritized by docking against multiple comparative protein structure models of OATP2B1, indicated that 33 of the 5,000 compounds were putative inhibitors of OATP2B1. Ten predicted inhibitors that are prescription drugs were tested experimentally in cells overexpressing the OATP2B1 transporter. Three of these ten were validated as potent inhibitors of estrone-3-sulfate uptake (defined as more than 50% inhibition at 20 μM) and tested in multiple concentrations to determine exact IC

After centuries of physicians believing that they know what is best for patients, this paternalistic approach to patient care is being challenged. The explosion in readily accessible information has resulted in patients being empowered to take on a greater role in decision making, from clinical care to regulatory decision making. The Centers of Excellence in Regulatory Science and Innovation (CERSIs) are collaborating with the US Food and Drug Administration (FDA) to promote patient engagement and advance patient preference research.

For deep sub-wavelength lithography, loss of wafer fidelity, such as line end shortening and corner rounding, is a known patterning phenomenon due to the diffraction limitation of the optical systems and some other processing effects. Without properly correcting these effects, particularly for gate corner rounding at the active area, sometime device performance might be limited or wafer yield will be impacted. It is addressable to improve the feature fidelity by OPC (Optical proximity correction) methodology, which is intentionally to offset the light distortion while in mask synthesis process. However, it is most likely becoming better, but not be completely solved at all. In this paper, first, the acceptable gate corner rounding criteria is examined. From the design rule constraint for gate region and test key electrical performance result, primary geometrical specification is determined. At the same time, considering inline process variation, such as ADI CD/overlay variation and loading effect of etching process, then the OPC corner rounding target specification comes accordingly. Second, OPC countermeasure for gate corner rounding improvement is studied. Usually gate CD uniformity is increased near the corner region, that is, gate poly CD is larger than expectation at the beginning of the active area near the L-shape or U-shape poly pattern. Since the transistor performance will be degraded, the improvement for corner rounding is important for OPC development. Aggressive OPC recipe to manipulate the polygon is required. We make use of localized fragmentation rule as well as specific retargeting to eliminate the impact of corner rounding effect in optical system. This methodology for corner fidelity improvement was proved by the wafer result.

Total deviation index (TDI) captures a prespecified quantile of the absolute deviation of paired observations from raters, observers, methods, assays, instruments, etc. We compare the performance of TDI using nonparametric quantile regression to the TDI assuming normality (Lin, 2000). This simulation study considers three distributions: normal, Poisson, and uniform at quantile levels of 0.8 and 0.9 for cases with and without contamination. Study endpoints include the bias of TDI estimates (compared with their respective theoretical values), standard error of TDI estimates (compared with their true simulated standard errors), and test size (compared with 0.05), and power. Nonparametric TDI using quantile regression, although it slightly underestimates and delivers slightly less power for data without contamination, works satisfactorily under all simulated cases even for moderate (say, ≥40) sample sizes. The performance of the TDI based on a quantile of 0.8 is in general superior to that of 0.9. The performances of nonparametric and parametric TDI methods are compared with a real data example. Nonparametric TDI can be very useful when the underlying distribution on the difference is not normal, especially when it has a heavy tail.

The normal wound healing process involves a well-organized cascade of biological pathways and any failure in this process leads to wounds becoming chronic. Non-healing wounds are a burden on healthcare systems and set to increase with aging population and growing incidences of obesity and diabetes. Stem cell-based therapies have the potential to heal chronic wounds but have so far seen little success in the clinic. Current research has been focused on using polymeric biomaterial systems that can act as a niche for these stem cells to improve their survival and paracrine activity that would eventually promote wound healing. Furthermore, different modification strategies have been developed to improve stem cell survival and differentiation, ultimately promoting regenerative wound healing. This review focuses on advanced polymeric scaffolds that have been used to deliver stem cells and have been tested for their efficiency in preclinical animal models of wounds.

A majority of patients with orofacial cleft deformity requires cleft repair through a bone graft. However, elevated amount of bone resorption and subsequent bone graft failure remains a significant clinical challenge. Bisphosphonates (BPs), a class of anti-resorptive drugs, may offer great promise in enhancing the clinical success of bone grafting. In this study, we compared the effects of systemic and local delivery of BPs in an intraoral bone graft model in rats. We randomly divided 34 female 20-week-old Fischer F344 Inbred rats into four groups to repair an intraoral critical-sized defect (CSD): (1) Control: CSD without graft (n = 4); (2) Graft/Saline: bone graft with systemic administration of saline 1 week post-operatively (n = 10); (3) Graft/Systemic: bone graft with systemic administration of zoledronic acid 1 week post-operatively (n = 10); and (4) Graft/Local: bone graft pre-treated with zoledronic acid (n = 10). At 6-weeks post-operatively, microCT volumetric analysis showed a significant increase in bone fraction volume (BV/TV) in the Graft/Systemic (62.99 ±14.31%) and Graft/Local (69.35 ±13.18%) groups compared to the Graft/Saline (39.18±10.18%). Similarly, histological analysis demonstrated a significant increase in bone volume in the Graft/Systemic (78.76 ±18.00%) and Graft/Local (89.95 ±4.93%) groups compared to the Graft/Saline (19.74±18.89%). The local delivery approach resulted in the clinical success of bone grafts, with reduced graft resorption and enhanced osteogenesis and bony integration with defect margins while avoiding the effects of BPs on peripheral osteoclastic function. In addition, local delivery of BPs may be superior to systemic delivery with its ease of procedure as it involves simple soaking of bone graft materials in BP solution prior to graft placement into the defect. This new approach may provide convenient and promising clinical applications towards effectively managing cleft patients.

© 2015 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved. Solute carrier (SLC) transporters — a family of more than 300 membrane-bound proteins that facilitate the transport of a wide array of substrates across biological membranes — have important roles in physiological processes ranging from the cellular uptake of nutrients to the absorption of drugs and other xenobiotics. Several classes of marketed drugs target well-known SLC transporters, such as neurotransmitter transporters, and human genetic studies have provided powerful insight into the roles of more-recently characterized SLC transporters in both rare and common diseases, indicating a wealth of new therapeutic opportunities. This Review summarizes knowledge on the roles of SLC transporters in human disease, describes strategies to target such transporters, and highlights current and investigational drugs that modulate SLC transporters, as well as promising drug targets.

BackgroundMicroporous Polysaccharide Hemospheres (MPH) are a new plant-derived polysaccharide powder hemostat. Previous studies investigated MPH as a replacement to nonflowable hemostatic agents of different application techniques (e.g., oxidized cellulose, collagen); therefore, the purpose of this study was to determine if MPH is a surrogate for flowable hemostatic agents of similar handling and application techniques, specifically a flowable thrombin-gelatin hemostatic matrix.MethodsHemostatic efficacy was compared using a heparinized porcine abrasion model mimicking a capsular tear of a parenchymal organ. MPH (ARISTA, 1 g) and hemostatic matrix (Floseal, 1 mL) were applied, according to a randomized scheme, to paired hepatic abrasions (40 lesions per group). Hemostatic success, control of bleeding, and blood loss were assessed 2, 5, and 10 min after treatment. Hemostatic success and control of bleeding were analyzed using odds ratios and blood loss using mean differences.ResultsHemostatic matrix provided superior hemostatic success relative to MPH at 5 (odds ratio: 0.035, 95% confidence interval: 0.004–0.278) and 10 min (0.032, 0.007–0.150), provided superior control of bleeding at 5 (0.006

Patients with cleft lip and/or palate (CLP), who undergo numerous medical interventions from infancy, can suffer from lifelong debilitation caused by underdeveloped maxillae. Conventional treatment approaches use maxillary expansion techniques to develop normal speech, achieve functional occlusion for nutrition intake, and improve esthetics. However, as patients with CLP congenitally lack bone in the cleft site with diminished capacity for bone formation in the expanded palate, more than 80% of the patient population experiences significant postexpansion relapse. While such relapse has been a long-standing battle in craniofacial care of patients, currently there are no available strategies to address this pervasive problem. Estrogen, 17β-estradiol (E2), is a powerful therapeutic agent that plays a critical role in bone homeostasis. However, E2's clinical application is less appreciated due to several limitations, including its pleiotropic effects and short half-life. Here, we developed a treatment strategy using an injectable system with photo-cross-linkable hydrogel (G) and nanodiamond (ND) technology to facilitate the targeted and sustained delivery of E2 to promote bone formation. In a preclinical expansion/relapse model, this functionalized E2/ND/G complex substantially reduced postexpansion relapse by nearly threefold through enhancements in sutural remodeling compared with unmodified E2 administration. The E2/ND/G group demonstrated greater bone volume by twofold and higher osteoblast number by threefold, compared with the control group. The E2/ND/G platform maximized the beneficial effects of E2 through its extended release with superior efficacy and safety at the local level. This broadly applicable E2 delivery platform shows promise as an adjuvant therapy in craniofacial care of patients.

The goal of pharmacogenomics research is to discover genetic polymorphisms that underlie variation in drug response. Increasingly, pharmacogenomics research involves large numbers of patients and the application of new technologies and methodologies to enable discovery. The Pharmacogenomics Research Network (PGRN) has become a community-driven network of investigators spanning scientific and clinical disciplines. Here, we highlight the activities and types of resources that enable PGRN members to enhance and drive basic and translational research in pharmacogenomics.

Beyond 40 nm technology node, the pattern weak points and hotspot types increase dramatically. The typical patterns for lithography verification suffers huge turn-around-time (TAT) to handle the design complexity. Therefore, in order to speed up process development and increase pattern variety, accurate design guideline and realistic design combinations are required. This paper presented a flow for creating a cell-based layout, a lite realistic design, to early identify problematic patterns which will negatively affect the yield. A new random layout generating method, Design Technology Co-Optimization Pattern Generator (DTCO-PG), is reported in this paper to create cell-based design. DTCO-PG also includes how to characterize the randomness and fuzziness, so that it is able to build up the machine learning scheme which model could be trained by previous results, and then it generates patterns never seen in a lite design. This methodology not only increases pattern diversity but also finds out potential hotspot preliminarily. This paper also demonstrates an integrated flow from DTCO pattern generation to layout modification. Optical Proximity Correction, OPC and lithographic simulation is then applied to DTCO-PG design database to detect hotspots and then hotspots or weak points can be automatically fixed through the procedure or handled manually. This flow benefits the process evolution to have a faster development cycle time, more complexity pattern design, higher probability to find out potential hotspots in early stage, and a more holistic yield ramping operation.

Made available in DSpace on 2018-11-26T17:12:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-12-01 Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) Donald P. Goldstein MD Trophoblastic Tumor Registry Endowment Dyett Family Trophoblastic Disease Research and Registry Endowment Objective. To compare the outcomes of Brazilian patients with molar pregnancy who continue human chorionic gonadotropin (hCG) surveillance with those treated with chemotherapy when hCG was still positive, but falling at 6 months after uterine evacuation. Methods. Retrospective chart review of 12,526 patients with hydatidiform mole treated at one of nine Brazilian reference centers from January 1990 to May 2016. Results. At 6 months from uterine evacuation', 96 (0.8%) patients had hCG levels raised but falling. In 15/96 (15.6%) patients, chemotherapy was initiated immediately per FIGO 2000 criteria, while 81/96 (84.4%) patients were managed expectantly. Among the latter, 65/81 (80.2%) achieved spontaneous remission and 16 (19.8%) developed postmolar gestational trophoblastic neoplasia (GTN). Patients who received chemotherapy following expectant management required more time for remission (11 versus 8 months; p = 0.001), had a greater interval between uterine evacuation and initiating chemotherapy (8 versus 6 months; p < 0.001), and presented with a median WHO/FIGO risk score higher than women treated according to FIGO 2000 criteria (4 versus 2, p = 0.04), but there were no significant differences in the need for multiagent treatment regimens (1/15 versus 3/16 patients, p = 0.60). None of the women relapsed, and no deaths occurred in either group. Conclusion. In order to avoid unnecessary exposure of women to chemotherapy, we no longer follow the FIGO 2000 recommendation to treat all patients with molar pregnancy and hCG raised but falling at 6 months after evacuation. Instead, we pursue close hormonal and radiological surveillance as the best strategy for these patients. (C) 2016 Elsevier Inc. All rights reserved. Univ Fed Fluminense, Rio de Janeiro Trophoblast Dis Ctr, Antonio Pedro Univ Hosp, Matern Sch,Matern Ward Santa Casa Misericordia Ri, Rio De Janeiro, RJ, Brazil Univ Fed Fluminense, Postgrad Program Med Sci, Niteroi, RJ, Brazil Univ Fed Rio de Janeiro, Matern Sch, Postgrad Program Perinatal Hlth, Fac Med, Rio De Janeiro, RJ, Brazil Sao Paulo State Univ, Trophoblast Dis Ctr, Clin Hosp, Botucatu Med Sch,Dept Gynecol & Obstet, Botucatu, SP, Brazil Univ Fed Sao Paulo, Paulista Sch Med, Sao Paulo Hosp, Trophoblast Dis Ctr, Sao Paulo, SP, Brazil Univ Sao Paulo, Sao Paulo Clin Hosp, Trophoblast Dis Ctr, Sao Paulo, SP, Brazil Caxias Do Sul Univ, Caxias Do Sul Trophoblast Dis Ctr, Gen Hosp Caxias Do Sul, Sch Med,Ctr Biol & Hlth Sci, Caxias Do Sul, MS USA Irmandade Santa Casa Misericordia Hosp, Porto Alegre Trophoblast Dis Ctr, Mario Totta Matern Ward, Porto Alegre, RS, Brazil Goias Fed Univ, Goias Trophoblast Dis Ctr, Clin Hosp Goias, Goiania, Go, Brazil Harvard Med Sch, Brigham & Womens Hosp, New England Trophoblast Dis Ctr, Div Gynecol Oncol,Dept Obstet & Gynecol & Reprod, Boston, MA USA Sao Paulo State Univ, Trophoblast Dis Ctr, Clin Hosp, Botucatu Med Sch,Dept Gynecol & Obstet, Botucatu, SP, Brazil

The purpose of this work was to prepare a stable paclitaxel nanosuspension and test it for potential use as a targeted chemotherapeutic. Different particle coatings were employed to assess their impact on cellular uptake in vitro. In vivo work was then performed to demonstrate efficacy in tumor-bearing mouse models.Paclitaxel nanosuspensions were prepared using a homogenization process and coated with excipients. Surface charge was measured by zeta potential, potency by high-performance liquid chromatography, and solubility using an in-line UV probe. Cellular uptake studies were performed via flow cytometry. In vivo experiments were performed to determine residence time, maximum tolerated dose, and the efficacy of paclitaxel nanosuspensions (Paclitaxel-NS).A stable paclitaxel nanosuspension was prepared and coated with various excipients. Studies in mice showed that the nanosuspension was well-tolerated and at least as effective as the IV Taxol control in prolonging mouse survival in a head and neck cancer model as well as an ovarian cancer model with a lower overall drug dose than the traditional IV administration route.The paclitaxel nanosuspension is suitable for cellular uptake. The nanosuspension was effective in prolonging life in two separate xenograft orthotopic murine cancer models through two separate routes of administration.

Background: Surgical hemostasis is achieved using adjunctive hemostats when conventional methods fail. Objective: This study compares the effectiveness of two adjunctive gelatin-thrombin hemostats. Hypothesis: To determine effectiveness, hemostats were compared in vivo, in vitro, and using scanning electron microscopy (SEM). Methods: In vivo, a heparinized porcine liver abrasion model was used to compare hemostatic success, degree of bleeding, and blood loss at 2, 5, and 10 minutes post-treatment. In vitro, thrombin in the supernatant of each hemostat and Red Blood Cells (RBC'S) in the supernatant of clots formed by each was compared. Results: Ultrastructure of one gelatin was smooth and the other stellate. In vivo, smooth gelatin provided superior hemostatic success at 5 (85% vs. 60%; OR: 5.3; 95% CI: 1.66 to 17.9) and 10 mins (72.5% vs. 47.5%; OR: 5.0; 95% CI: 1.55 to 16.1). Smooth gelatin had a statistically different degree of bleeding at 5 (0.58 ± 0.87 [Mean ± SD] vs. 1.03 ± 1.12; OR: 3.36; 95% CI: 1.34 to 8.41) and 10 mins (1.13 ± 1.14 vs. 1.65 ± 1.05; OR: 3.87; 95% CI: 1.62 to 9.21). Mean blood loss was less with smooth gelatin at 2 (0.07 ± 0.19 vs. 0.13 ± 0.63 ml/min), 5 (0.04 ± 0.13 vs. 0.23 ± 0.45 ml/min), and 10 mins (0.09 ± 0.24 vs. 0.21 ± 0.32 ml/min). In vitro, supernatant of smooth gelatin had significantly less thrombin (6.81 vs. 10.9 IU/ml, p = .001), and significantly less RBC's than stellate gelatin (0.07 vs. 0.09 × 106/ul, p = .0085). Conclusion: Smooth gelatin has an increased ability to retain thrombin and RBC's in vitro which may explain why it provides superior hemostatic effectiveness, superior control of bleeding, and greater reduced blood loss in vivo.

Metformin is used as a first-line therapy for type 2 diabetes (T2D) and prescribed for numerous other diseases. However, its mechanism of action in the liver has yet to be characterized in a systematic manner. To comprehensively identify genes and regulatory elements associated with metformin treatment, we carried out RNA-seq and ChIP-seq (H3K27ac, H3K27me3) on primary human hepatocytes from the same donor treated with vehicle control, metformin or metformin and compound C, an AMP-activated protein kinase (AMPK) inhibitor (allowing to identify AMPK-independent pathways). We identified thousands of metformin responsive AMPK-dependent and AMPK-independent differentially expressed genes and regulatory elements. We functionally validated several elements for metformin-induced promoter and enhancer activity. These include an enhancer in an ataxia telangiectasia mutated (ATM) intron that has SNPs in linkage disequilibrium with a metformin treatment response GWAS lead SNP (rs11212617) that showed increased enhancer activity for the associated haplotype. Expression quantitative trait locus (eQTL) liver analysis and CRISPR activation suggest that this enhancer could be regulating ATM, which has a known role in AMPK activation, and potentially also EXPH5 and DDX10, its neighboring genes. Using ChIP-seq and siRNA knockdown, we further show that activating transcription factor 3 (ATF3), our top metformin upregulated AMPK-dependent gene, could have an important role in gluconeogenesis repression. Our findings provide a genome-wide representation of metformin hepatic response, highlight important sequences that could be associated with interindividual variability in glycemic response to metformin and identify novel T2D treatment candidates., Author Summary Metformin is among the most widely prescribed drugs. It is used as a first line therapy for type 2 diabetes (T2D), and for additional diseases including cancer. The variability in response to metformin is substantial and can be caused by genetic factors. However, the molecular mechanisms of metformin action are not fully known. Here, we used various genomic assays to analyze human liver cells treated with or without metformin and identified in a genome-wide manner thousands of differentially expressed genes and gene regulatory elements affected by metformin. Follow up functional assays identified several novel genes and regulatory elements to be associated with metformin response. These include ATF3, a gene that showed gluconeogenesis repression upon metformin response and a potential regulatory element of the ATM gene that is associated with metformin treatment differences through genome-wide association studies. Combined, this work identifies several novel genes and gene regulatory elements that can be activated due to metformin treatment and thus provides candidate sequences in the human genome where nucleotide variation can lead to differences in metformin response. It also enables the identification and prioritization of novel candidates for T2D treatment.

© 2016 Elsevier Ltd Large neutral amino acid transporter 1 (LAT1) is a solute carrier protein located primarily in the blood–brain barrier (BBB) that offers the potential to deliver drugs to the brain. It is also up-regulated in cancer cells, as part of a tumor's increased metabolic demands. Previously, amino acid prodrugs have been shown to be transported by LAT1. Carboxylic acid bioisosteres may afford prodrugs with an altered physicochemical and pharmacokinetic profile than those derived from natural amino acids, allowing for higher brain or tumor levels of drug and/or lower toxicity. The effect of replacing phenylalanine's carboxylic acid with a tetrazole, acylsulfonamide and hydroxamic acid (HA) bioisostere was examined. Compounds were tested for their ability to be LAT1 substrates using both cis-inhibition and trans-stimulation cell assays. As HA-Phe demonstrated weak substrate activity, its structure–activity relationship (SAR) was further explored by synthesis and testing of HA derivatives of other LAT1 amino acid substrates (i.e., Tyr, Leu, Ile, and Met). The potential for a false positive in the trans-stimulation assay caused by parent amino acid was evaluated by conducting compound stability experiments for both HA-Leu and the corresponding methyl ester derivative. We concluded that HA's are transported by LAT1. In addition, our results lend support to a recent account that amino acid esters are LAT1 substrates, and that hydrogen bonding may be as important as charge for interaction with the transporter binding site.

© 2016 Elsevier Ltd. All rights reserved. The transporter protein Large-neutral Amino Acid Transporter 1 (LAT-1, SLC7A5) is responsible for transporting amino acids such as tyrosine and phenylalanine as well as thyroid hormones, and it has been exploited as a drug delivery mechanism. Recently its role in cancer has become increasingly appreciated, as it has been found to be up-regulated in many different tumor types, and its expression levels have been correlated with prognosis. Substitution at the meta position of aromatic amino acids has been reported to increase affinity for LAT-1; however, the SAR for this position has not previously been explored. Guided by newly refined computational models of the binding site, we hypothesized that groups capable of filling a hydrophobic pocket would increase binding to LAT-1, resulting in improved substrates relative to parent amino acid. Tyrosine and phenylalanine analogs substituted at the meta position with halogens, alkyl and aryl groups were synthesized and tested in cis-inhibition and trans-stimulation cell assays to determine activity. Contrary to our initial hypothesis we found that lipophilicity was correlated with diminished substrate activity and increased inhibition of the transporter. The synthesis and SAR of meta-substituted phenylalanine and tyrosine analogs is described.

Free radicals or reactive oxygen species (ROS) are relatively short-lived and are difficult to measure directly; so indirect methods have been explored for measuring these transient species. One technique that has been developed using Escherichia coli and Saccharomyces cerevisiae systems, relies on a connection between elevated superoxide levels and the build-up of a high-spin form of iron (Fe(III)) that is detectable by electron paramagnetic resonance (EPR) spectroscopy at g = 4.3. This form of iron is referred to as "free" iron. EPR signals at g = 4.3 are commonly encountered in biological samples owing to mononuclear high-spin (S = 5/2) Fe(III) ions in sites of low symmetry. Unincorporated iron in this study refers to this high-spin Fe(III) that is captured by desferrioxamine which is detected by EPR at g value of 4.3. Previously, we published an adaptation of Fe(III) EPR methodology that was developed for Caenorhabditis elegans, a multi-cellular organism. In the current study, we have systematically characterized various factors that modulate this unincorporated iron pool. Our results demonstrate that the unincorporated iron as monitored by Fe(III) EPR at g = 4.3 increased under conditions that were known to elevate steady-state ROS levels in vivo, including: paraquat treatment, hydrogen peroxide exposure, heat shock treatment, or exposure to higher growth temperature. Besides the exogenous inducers of oxidative stress, physiological aging, which is associated with elevated ROS and ROS-mediated macromolecular damage, also caused a build-up of this iron. In addition, increased iron availability increased the unincorporated iron pool as well as generalized oxidative stress. Overall, unincorporated iron increased under conditions of oxidative stress with no change in total iron levels. However, when total iron levels increased in vivo, an increase in both the pool of unincorporated iron and oxidative stress was observed suggesting that the status of the unincorporated iron pool is linked to oxidative stress and iron levels.

This paper is an overview that summarizes recently developed tools in assessing agreement for methods comparison and instrument/assay validation in medical devices. This paper emphasizes concept, sample sizes, and examples more than analytical formulas. We have considered a unified approach of evaluating agreement among multiple instruments (k), each with multiple replicates (m) for both continuous and categorical data. We start with the basic scenario of two instruments (k = 2), each with only one measurement (m = 1). In this basic scenario for continuous data, we also consider if the target values are considered random (values of a gold standard instrument) or fixed (known values). In the more general case, we will not consider when the target values are fixed. We discuss the simplified sample size calculations. When there is a disagreement between methods, one needs to know if the source of the disagreement was due to a systematic shift (bias) or random error. The coefficients of accuracy and precision will be discussed to characterize these sources. This is important because a systematic shift usually can be easily fixed through calibration, while a random error usually is a more cumbersome variation reduction exercise. For categorical variables, we consider scaled agreement statistics. For continuous variables, we use scaled or unscaled agreement statistics. For variables with proportional error, we can simply apply a log transformation to the data. Finally, three examples are given: one for assay validation, one for a lab proficiency assessment, and one for a lab comparison on categorical assay.

This paper proposes several Concordance Correlation Coefficient (CCC) indices to measure the agreement among k raters, with each rater having multiple (m) readings from each of the n subjects for continuous and categorical data. In addition, for normal data, this paper also proposes the coverage probability (CP) and total deviation index (TDI). Those indices are used to measure intra, inter and total agreement among all raters. Intra-rater indices are used to measure the agreement among the multiple readings from the same rater. Inter-rater indices are used to measure the agreement among different raters based on the average of multiple readings. Total-rater indices are used to measure the agreement among different raters based on individual readings. In addition to the agreement, the paper also assess intra, inter, and total precision and accuracy. Through a two-way mixed model, all CCC, precision and accuracy, TDI, and CP indices are expressed as functions of variance components, and GEE method is used to obtain the estimates and perform inferences for all the functions of variance components. Each of previous proposed approaches for assessing agreement becomes one of the special case of the proposed approach. For continuous data, when m approaches infinity, the proposed estimates reduce to the agreement indices proposed by Barnhart et al. (2005). When m = 1, the proposed estimate reduces to the ICC proposed by Carrasco and Jover (2003). When m = 1, the proposed estimate also reduces to the OCCC proposed by Lin (1989), King and Chinchilli (2001a) and Barnhart et al. (2002). When m = 1 and k = 2, the proposed estimate reduces to the original CCC proposed by Lin (1989). For categorical data, when k = 2 and m = 1, the proposed estimate and its associated inference reduce to the kappa for binary data and weighted kappa with squared weight for ordinal data.

PurposeVesicular monoamine transporters 1 and 2 (VMAT1 and VMAT2) are thought to mediate MIBG uptake in adult neuroendocrine tumors. In neuroblastoma, the norepinephrine transporter (NET) has been investigated as the principal MIBG uptake protein, though some tumors without NET expression concentrate MIBG. We investigated VMAT expression in neuroblastoma and correlated expression with MIBG uptake and clinical features.MethodsWe evaluated VMAT1 and VMAT2 expression by immunohistochemistry (IHC) in neuroblastoma tumors from 76 patients with high-risk metastatic disease treated in a uniform cooperative group trial (COG A3973). All patients had baseline MIBG diagnostic scans centrally reviewed. IHC results were scored as the product of intensity grading (0 - 3+) and percent of tumor cells expressing the protein of interest. The association between VMAT1 and VMAT2 scores and clinical and biological features was tested using Wilcoxon rank-sum tests.ResultsPatient characteristics were typical of high-risk neuroblastoma, though the cohort was intentionally enriched in patients with MIBG-nonavid tumors (n = 20). VMAT1 and VMAT2 were expressed in 62% and 75% of neuroblastoma tumors, respectively. VMAT1 and VMAT2 scores were both significantly lower in MYCN amplified tumors and in tumors with high mitotic karyorrhectic index. MIBG-avid tumors had significantly higher VMAT2 scores than MIBG-nonavid tumors (median 216 vs. 45; p = 0.04). VMAT1 expression did not correlate with MIBG avidity.ConclusionVMAT1 and VMAT2 are expressed in the majority of neuroblastomas. Expression correlates with other biological features. The expression level of VMAT2 but not that of VMAT1 correlates with avidity for MIBG.

← BackgroundHyaluronan (HA) is a glycosaminoglycan found in connective tissues and tissue spaces, including the peritoneal cavity. In vivo studies in a rat model of peritoneal dialysis (PD) have shown that addition of HA to PD solution during an intraperitoneal dwell can alter peritoneal fluid transport and protect the peritoneal membrane from the effects of inflammation and repeated infusions of dialysis solution. The current study sought to evaluate the safety of intraperitoneal HA and its effect on peritoneal fluid and solute transport when administered during a dialysis dwell in humans.← Methods13 PD patients were enrolled in a prospective, randomized crossover study involving three dialysis treatments using the following PD solutions: ( 1 ) a commercially available PD solution (Dianeal PD-4, 1.36% glucose; Baxter Healthcare Corporation, Alliston, Ontario, Canada); ( 2 ) Dianeal PD-4 containing 0.1 g/L HA, and ( 3 ) Dianeal PD-4 containing 0.5 g/LHA. Each 6-hour dialysis exchange was separated from the other exchanges by a 2-week washout period. Radioiodinated human serum albumin (RISA) was administered with the dialysis solution to evaluate intraperitoneal volume, net ultrafiltration (UF), and fluid reabsorption. Peritoneal clearances, dialysate/plasma ratios (D/P), and mass transfer area coefficients (MTACs) were determined for sodium, urea, creatinine, albumin, and glucose. Safety was evaluated by monitoring adverse events and changes in serum chemistries. Ten patients completed all three dialysis exchanges and two additional patients completed at least one treatment exchange.← ResultsThere were no reported adverse events related to HA administration and no significant changes in serum chemistries. There were no significant differences in net UF or peritoneal volume profiles among the three treatments. Mean net UF calculated using residual volumes, estimated by RISA dilution, tended to be slightly higher during treatment with solution containing 0.1 g/L HA and 0.5 g/L HA [74 ± 86 (SE) and 41 ± 99 mL, respectively] compared to control treatment (–58 ± 129 mL). Although not statistically significant, there was a trend toward decreased fluid reabsorption during treatment with HA. Solute clearances, D/P ratios, and MTACs were similar for the three treatments. Serum levels of HA were also unaffected by the two treatment solutions.← ConclusionsThese data support the acute safety of HA when administered intraperitoneally with the dialysis solution to PD patients. Due to the small sample size and variability in net UF and fluid reabsorption, statistically significant differences were not demonstrated for these parameters. However, a trend toward decreased fluid reabsorption was observed, suggesting that HA may act by a mechanism similar to that observed in animal studies. Further studies are necessary to evaluate whether the beneficial effects of HA observed in animal studies can be shown in humans.

Measurements of agreement are needed to assess the acceptability of a new or generic process, methodology, and formulation in areas of laboratory performance, instrument or assay validation, method comparisons, statistical process control, goodness of fit, and individual bioequivalence. In all of these areas, one needs measurements that capture a large proportion of data that are within a meaningful boundary from target values. Target values can be considered random (measured with error) or fixed (known), depending on the situation. Various meaningful measures to cope with such diverse and complex situations have become available only in the last decade. These measures often assume that the target values are random. This article reviews the literature and presents methodologies in terms of “coverage probability.” In addition, analytical expressions are introduced for all of the aforementioned measurements when the target values are fixed and when the error structure is homogenous or heterogeneous (proport...

Despite having high-resolution structures for eukaryotic large ribosomal subunits, it remained unclear how these ribonucleoprotein complexes are constructed in living cells. Nevertheless, knowing where ribosomal proteins interact with ribosomal RNA (rRNA) provides a strategic platform to investigate the connection between spatial and temporal aspects of 60S subunit biogenesis. We previously found that the function of individual yeast large subunit ribosomal proteins (RPLs) in precursor rRNA (pre-rRNA) processing correlates with their location in the structure of mature 60S subunits. This observation suggested that there is an order by which 60S subunits are formed. To test this model, we used proteomic approaches to assay changes in the levels of ribosomal proteins and assembly factors in preribosomes when RPLs functioning in early, middle, and late steps of pre-60S assembly are depleted. Our results demonstrate that structural domains of eukaryotic 60S ribosomal subunits are formed in a hierarchical fashion. Assembly begins at the convex solvent side, followed by the polypeptide exit tunnel, the intersubunit side, and finally the central protuberance. This model provides an initial paradigm for the sequential assembly of eukaryotic 60S subunits. Our results reveal striking differences and similarities between assembly of bacterial and eukaryotic large ribosomal subunits, providing insights into how these RNA–protein particles evolved.

BackgroundThe primary rescue medication to treat acute asthma exacerbation is the short-acting β₂-adrenergic receptor agonist; however, there is variation in how well a patient responds to treatment. Although these differences might be due to environmental factors, there is mounting evidence for a genetic contribution to variability in bronchodilator response (BDR).ObjectiveTo identify genetic variation associated with bronchodilator drug response in Latino children with asthma.MethodsWe performed a genome-wide association study (GWAS) for BDR in 1782 Latino children with asthma using standard linear regression, adjusting for genetic ancestry and ethnicity, and performed replication studies in an additional 531 Latinos. We also performed admixture mapping across the genome by testing for an association between local European, African, and Native American ancestry and BDR, adjusting for genomic ancestry and ethnicity.ResultsWe identified 7 genetic variants associated with BDR at a genome-wide significant threshold (P < 5 × 10(-8)), all of which had frequencies of less than 5%. Furthermore, we observed an excess of small P values driven by rare variants (frequency

This article proposes a general comparison model for assessing individual agreement of k ≥ 2 raters evaluating n subjects with m ≥ 2 replicated readings. Users can explore total-rater agreement relative to intrarater agreement where any subset of the k raters can be selected in the numerator and denominator. Users are also allowed to compare intrarater agreement among selected raters. Based on the ratio of mean squared deviations (MSDs), two comparative agreement indices, total-intra ratio (TIR) and intra-intra ratio (IIR), are proposed. The TIR is a noninferiority assessment such that the differences of individual readings from different raters cannot be inferior by a prespecified margin to the differences of the replicated readings within raters. TIR can be used whether a reference exists or not. The method used by the Food and Drug Administration (FDA) for evaluating individual bioequivalence under relative scale becomes the special case of our approach. The IIR is a classical assessment such that the precision of selected raters can be better than; equal to; or worse than that of other raters. The estimation and statistical inference of TIR and IIR are obtained through GEE methodology.

Rice Helping Hands (RHH) is tasked with designing and developing a wrist rehabilitation device for pediatric patients with spastic cerebral palsy (CP).Copyright © 2012 by ASME

In this chapter, in addition to the formal method of computing the sample size and power, we will discuss ways to compute sample size and power based on normal approximation through a simplified and conservative approach. As pointed out previously in assessing agreement, the traditional null and alternative hypotheses should be reversed. The conventional rejection region is now the region of declaring agreement, and is usually one-sided. Asymptotic power and sample size calculations will proceed by this principle.