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BACKGROUND Endoscopic assessment of disease severity is a key component of ulcerative colitis (UC) clinical studies. However, interobserver variability may contribute to bias in selecting study candidates and evaluating response rates. Scoring by blinded central readers has become the recommended standard method of assessment to minimize bias and interobserver variance. We present concordance rates between central and local readings from a Phase 2b study of investigational microbiome therapeutic SER-287 in patients with mild-to-moderate UC. METHODS This double-blind, placebo-controlled study screened 494 and randomized 203 adults (51.7% male; mean age 45.6 years; 83.7% white) 1:1:1 to 3 arms (Figure 1). Active UC was defined as ≥15 cm disease from the anal verge, a Three-Component Modified Mayo Score of 3-7, endoscopic subscore (ES) ≥1, and stool frequency subscore ≥1. Endoscopy was performed at screening, after induction, after exploratory maintenance or open-label, and for flare assessments. Endoscopic videos were scored by each site’s gastroenterologist (local ES), and by any 1 of 4 blinded central readers (Alimentiv; Ontario), with adjudication by a 2nd blinded central reader (Alimentiv), if necessary. If all 3 scores were discordant, the median score was designated as the final ES. Concordance between local and central readers were analyzed by timepoint and in total and quantified using weighted kappa statistics. RESULTS Concordance analyses were conducted for 608 ES comparisons between local and the 1st central reader (Local vs Central1 [LvC1]), and 337 comparisons where adjudication was necessary (Local vs Central2 [LvC2]; Central1 vs Central2 [C1vC2]). Observed concordance rates were: -LvC1: 44% (271/608, kappa = 0.61, Figure 2); -LvC2: 26% (86/337, kappa = 0.46); -C1vC2: 64% (216/337, kappa = 0.74). Concordance rates were comparable across study periods: -Screening: -LvC1: 42% (131/310, kappa = 0.59) -LvC2: 27% (48/179, kappa = 0.48) -C1vC2: 66% (118/179, kappa = 0.76) -Week 11: -LvC1: 46% (79/170, kappa = 0.61) -LvC2: 25% (22/91, kappa = 0.46) -C1vC2: 62% (56/91, kappa = 0.75) Local readers scored lower than C1 readers in 274/337 (81%) discordant cases (Figure 2), and lower than C2 in 221/251 (88%) discordant cases. The most frequent sources of discordance were L=ES2 vs C1=ES3 (153 videos) and L=ES1 vs C1=ES2 (77 videos). CONCLUSIONS In this Phase 2b study, low ES concordance highlights marked interobserver variability between local and blinded central readers. Local scores were consistently lower than central scores across all study time points. Despite low concordance rates, efficacy conclusions remain unchanged if only local ES scores were used for endpoint assessments. Objective, standardized, and consistent methods for evaluating ES are needed to better quantify disease severity and the impact of UC therapies.

Radiology education is essential in medical school; however, developing an integrated and comprehensive curriculum remains a challenge. Many novel methods have been implemented with varying outcomes. In this study, the authors sought to examine published pedagogical methods of radiology instruction and query US academic faculty members on their current use within radiology education.A literature search for current and novel pedagogical methods of radiology instruction was performed and studies were assessed for positive educational outcomes. Educational approaches were grouped according to encountered themes. A survey was distributed to faculty members of the Alliance of Medical Student Educators in Radiology to ascertain the prevalence of these pedagogical methods in the radiology education of medical students.The following themes were encountered: supplemental instruction of anatomy and pathology; radiology-clinical correlation electives; flipped classrooms; hands-on and simulation training; peer-to-peer learning; e-learning; adaptive tutorials; and asynchronous learning. Of the survey respondents, 90% reported that their institution offers a formal radiology clerkship. The majority of respondents reported the use of flipped classrooms (70%) and e-learning (78%); however, few reported offering hands-on clinical experiences (31%) and simulation-based training (36%). Only 5% reported use of adaptive tutorials.In the review of the literature, a combination of hands-on, case-based, team-based, and didactic training, in addition to other forms of active learning within an integrated curriculum, was found to be highly effective and preferred by students and faculty. Virtual and in-person learning incorporating modern technology was found to either increase knowledge and skills or yield similar outcomes as traditional in-person instruction. These methods are currently heterogeneously used across the US medical schools represented by survey respondents, with utilization ranging from 5% to 78%.

In a post-quantum world, where attackers may have access to full-scale quantum computers, all classical password-based authentication schemes will be compromised. Quantum copy-protection prevents adversaries from making copies of existing quantum software; we suggest this as a possible approach for designing post-quantum-secure password authentication systems. In this paper, we show an implementation of quantum copy-protection for password verification on IBM quantum computers. We also share our quantum computation results and analyses, as well as lessons learned., Comment: 7 pages, 8 figures, 4 tables

Adaptive reward-related decision making often requires accurate and detailed representation of potential available rewards. Environmental reward-predictive stimuli can facilitate these representations, allowing one to infer which specific rewards might be available and choose accordingly. This process relies on encoded relationships between the cues and the sensory-specific details of the rewards they predict. Here, we interrogated the function of the basolateral amygdala (BLA) and its interaction with the lateral orbitofrontal cortex (lOFC) in the ability to learn such stimulus-outcome associations and use these memories to guide decision making. Using optical recording and inhibition approaches, Pavlovian cue-reward conditioning, and the outcome-selective Pavlovian-to-instrumental transfer (PIT) test in male rats, we found that the BLA is robustly activated at the time of stimulus-outcome learning and that this activity is necessary for sensory-specific stimulus-outcome memories to be encoded, so they can subsequently influence reward choices. Direct input from the lOFC was found to support the BLA in this function. Based on prior work, activity in BLA projections back to the lOFC was known to support the use of stimulus-outcome memories to influence decision making. By multiplexing optogenetic and chemogenetic inhibition we performed a serial circuit disconnection and found that the lOFC→BLA and BLA→lOFC pathways form a functional circuit regulating the encoding (lOFC→BLA) and subsequent use (BLA→lOFC) of the stimulus-dependent, sensory-specific reward memories that are critical for adaptive, appetitive decision making.

INTRODUCTION Cataract formation is a tissue reaction effected by radiation exposure. The purpose of this study was to evaluate the occupational exposure to the lens of the eye of interventional radiologists (IR's) and interventional radiology staff, with and without lead glasses. METHODS Ethical approval was provided by the hospital research and ethics committee. A prospective cohort study was performed over 1 year, doses recorded, lifetime dose (estimated at working 5 days in angiography, for 30 years) was estimated and dose compared to current guidelines. Thermoluminescent dosimeters (TLDs; Landauer, Glenwood, USA) Hp(3) were placed on both the exterior and interior side of the personal lead glasses worn by three interventional radiologists and two radiographers. They were monitored during all procedures performed within 1 year. Lead glasses (AttenuTech® Microlite® , Florida, USA) with specifications were 0.75 mm lead equivalent front shield, and Side shield 0.3 mm Pb equivalent. A control TLD was placed in the storage location of the lead glasses when not in use. Yearly dose was measured and lifetime dose was calculated from the data obtained. Calculation of dose received per day(s) spent performing procedures for both annual and lifetime exposure was performed. In addition a record of occurrence of splashes on glasses was made after each case. RESULTS Eye doses without protection were double the recommended limits for both annual and lifetime dose. For interventional radiologists working between 3 and 4 or more days in the lab per week, annual dose thresholds would be exceeded (20 mSv/year averaged over 5 years, no more than 50 mSv in 1 year). If interventional radiologists worked between 3 and 4 or more days in the lab, lifetime dose thresholds would be exceeded (500 mSv lifetime dose). Lead glasses reduced radiation exposure by an average of 79%. If lead glasses were worn no interventional radiologists would exceed annual or lifetime dose thresholds to the eyes even if working 5 days per week as the primary operator. Radiographers would not exceed annual or lifetime dose thresholds even without lead glasses. Splash incidents occurred for all interventional radiologists and one radiographer. CONCLUSION The use of lead glasses even in this small study resulted in a decreased dose of radiation to the lens of the eye. Regular use of radiation protection eyewear will reduce eye dose for primary proceduralists to well below yearly and lifetime thresholds.

Adaptive reward-related decision making often requires accurate and detailed representation of potential available rewards. Environmental reward-predictive stimuli can facilitate these representations, allowing one to infer which specific rewards might be available and choose accordingly. This process relies on encoded relationships between the cues and the sensory-specific details of the reward they predict. Here we interrogated the function of the basolateral amygdala (BLA) and its interaction with the lateral orbitofrontal cortex (lOFC) in the ability to learn such stimulus-outcome associations and use these memories to guide decision making. Using optical recording and inhibition approaches, Pavlovian cue-reward conditioning, and an outcome-selective Pavlovian-to-instrumental transfer (PIT) test in male rats, we found that the BLA is robustly activated at the time of stimulus-outcome learning and that this activity is necessary for sensory-specific stimulus-outcome memories to be encoded, so that they can subsequently influence reward choices. Direct input from the lOFC was found to support the BLA in this function. Based on prior work, activity in BLA projections back to the lOFC was known to support the use of stimulus-outcome memories to influence decision making. By multiplexing optogenetic and chemogenetic inhibition to perform a serial circuit disconnection, we found that activity in lOFC[->]BLA projections regulates the encoding of the same components of the stimulus-outcome memory that are later used to allow cues to guide choice via activity in BLA[->]lOFC projections. Thus, the lOFC[->]BLA[->]lOFC circuit regulates the encoding (lOFC[->]BLA) and subsequent use (BLA[->]lOFC) of the stimulus-dependent, sensory-specific reward memories that are critical for adaptive, appetitive decision making.

Background & Aims Firmicutes bacteria produce metabolites that maintain the intestinal barrier and mucosal immunity. Firmicutes are reduced in the intestinal microbiota of patients with ulcerative colitis (UC). In a phase 1b trial of patients with UC, we evaluated the safety and efficacy of SER-287, an oral formulation of Firmicutes spores, and the effects of vancomycin preconditioning on expansion (engraftment) of SER-287 species in the colon. Methods We conducted a double-blind trial of SER-287 in 58 adults with active mild-to-moderate UC (modified Mayo scores 4–10, endoscopic subscores ≥1). Participants received 6 days of preconditioning with oral vancomycin (125 mg, 4 times daily) or placebo followed by 8 weeks of oral SER-287 or placebo. Patients were randomly assigned (2:3:3:3) to groups that received placebo followed by either placebo or SER-287 once weekly, or vancomycin followed by SER-287 once weekly or SER-287 once daily. Clinical endpoints included safety and clinical remission (modified Mayo score ≤ 2; endoscopic subscores 0 or 1). Microbiome endpoints included SER-287 engraftment (dose species detected in stool after, but not before, SER-287 administration). Engraftment of SER-287 and changes in microbiome composition and associated metabolites were measured by analyses of stool specimens collected at baseline, after preconditioning, and during and 4 weeks after administration of SER-287 or placebo. Results Proportions of patients with adverse events did not differ significantly among groups. A higher proportion of patients in the vancomycin/SER-287 daily group (40%) achieved clinical remission at week 8 than patients in the placebo/placebo group (0), placebo/SER-287 weekly group (13.3%), or vancomycin/SER-287 weekly group (17.7%) (P=.024 for vancomycin/SER-287 daily vs placebo/placebo). By day 7, higher numbers of SER-287 dose species were detected in stool samples from all SER-287 groups compared with the placebo group (P

Introduction: In recent years, the ability to detect several proteins on a single slide in Multiplex Immunofluorescence (mIF) experiments has become common place. This is possible thanks to factors such as development of antibodies of high quality, improvements in fluorescent technology, and automation of IHC staining. One of the advantages of multiplexing protein detection is to gain maximal data per tissue section, which is critical when samples are limited. Understanding co-expression and spatial organization of multiple targets within preserved tissue architecture is also important, especially in tumor microenvironment (TME) analysis. mIF plays a key role in research fields like tumor immunology where it is needed to catalog subsets of immune and cancer cells within the TME. Ultimately, this will enable the development of personalized, combinatorial therapeutic interventions. Materials and Methods: Selected antibody panels: Tonsil Panel 1: p40, CD3, CD8, CD20 & CD68 Tonsil Panel 2: LAG3 [BC40], LAG3 [CAL26], CD57, CD19, FOXP3, T-bet, PD-L1, PD-1 & GATA-3 Colon Cancer Panel: Granzyme B, CD8, CDX2, CD3 & Pan Cytokeratin Plus Melanoma Panel: Melan A, CD3, SOX10, CD8 & Granzyme B Detection was performed with Biocare MACH 2 Universal HRP Polymer. Fluorochromes: Opal 480, Opal 520, Opal 570, Opal 620, Opal 650 VALENT (Biocare Medical) fully automated staining platform was utilized to stain slides. Image acquisition and analysis was performed on an Olympus BX61 microscope coupled with the ASI FISH imaging system. In the workflow, a proprietary room temperature (RT) stripping reagent was utilized instead of the heat stripping step commonly used between antibody detection incubations. Results and Discussion: The antibodies used have been previously optimized for standard IHC protocols on VALENT using DAB. However, the sensitivity of tyramide detection is very high and for most of the antibodies, the incubation times had to be drastically reduced to minimize non-specific staining and the potential bleed-through to other filters for neighboring fluors. Additionally, Opal fluorochromes were diluted between 1:200 and 1:300. The RT stripping reagent helps in the preservation of tissue integrity even after the sequential detection of 5 antibodies. Additionally, by using this reagent, a rigorous order of primary antibody application is no longer needed. Finally, this reagent dimmed some autofluorescence coming from red blood cells. Following some brief optimization steps, we obtained strong and specific staining patterns similar to ones obtained with individual IHC experiments visualized with DAB. Conclusion: We have demonstrated the use on an automated platform of a RT stripping solution. Additionally, this reagent can be used with different antibodies in order to effectively characterize the TME in many cancers. Citation Format: Julio S. Masabanda, Sherry Wang, Joseph Vargas, Jason Ramos. Development of automated multiplex immunofluorescence protocols for tumor microenvironment evaluation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2780.

The interfollicular epidermis regenerates from heterogeneous basal skin cell populations that divide at different rates. It has previously been presumed that infrequently dividing basal cells known as label-retaining cells (LRCs) are stem cells, whereas non-LRCs are short-lived progenitors. Here we employ the H2B-GFP pulse-chase system in adult mouse skin and find that epidermal LRCs and non-LRCs are molecularly distinct and can be differentiated by Dlx1(CreER) and Slc1a3(CreER) genetic marking, respectively. Long-term lineage tracing and mathematical modelling of H2B-GFP dilution data show that LRCs and non-LRCs constitute two distinct stem cell populations with different patterns of proliferation, differentiation and upward cellular transport. During homeostasis, these populations are enriched in spatially distinct skin territories and can preferentially produce unique differentiated lineages. On wounding or selective killing, they can temporarily replenish each other's territory. These two discrete interfollicular stem cell populations are functionally interchangeable and intrinsically well adapted to thrive in distinct skin environments.

The prevalence of bulimia nervosa (BN) and binge eating disorder (BED) among Latinas is comparable to those of the general population; however, few interventions and treatment trial research have focused on this group. Cognitive-behavioral therapy (CBT) is the treatment of choice for binge eating related disorders. CBT-based guided self-help (CBTgsh)-a low-cost minimal intervention-has also been shown effective in improving binge eating related symptom, but the effectiveness of the CBTgsh among ethnic minority women is not well understood. Cultural adaptation of evidence-based treatments can be an important step for promoting treatment accessibility and engagement among underserved groups. This qualitative study was part of a larger investigation that examined the feasibility and efficacy of a culturally adapted CBTgsh program among Mexican American women with binge eating disorders. Posttreatment focus groups were conducted with 12 Mexican American women with BN or BED who participated in the intervention. Data were analyzed with the grounded theory methodology (Corbin & Strauss, 2008). Three themes emerged from the data: (a) eating behavior and body ideals are socially and culturally constructed, (b) multifaceted support system is crucial to Mexican American women's treatment engagement and success, and (c) the culturally adapted CBTgsh program is feasible and relevant to Mexican American women's experience, but it can be strengthened with increased family and peer involvement. The findings provide suggestions for further adaptation and refinement of the CBTgsh, and implications for future research as well as early intervention for disordered eating in organized care settings.

BACKGROUND Using smartphones to enroll, obtain consent, and gather self-reported data from patients has the potential to enhance our understanding of disease burden and quantify physiological impact in the real world. It may also be possible to harness integral smartphone sensors to facilitate remote collection of clinically relevant data. OBJECTIVE We conducted the Patient Rheumatoid Arthritis Data From the Real World (PARADE) observational study using a customized ResearchKit app with a bring-your-own-device approach. Our objective was to assess the feasibility of using an entirely digital approach (social media and smartphone app) to conduct a real-world observational study of patients with rheumatoid arthritis. METHODS We conducted this observational study using a customized ResearchKit app with a bring-your-own-device approach. To recruit patients, the PARADE app, designed to guide patients through a series of tasks, was publicized via social media platforms and made available for patients in the United States to download from the Apple App Store. We collected patient-reported data, such as medical history, rheumatoid arthritis-related medications (past and present), and a range of patient-reported outcome measures. We included in the assessment a joint-pain map and a novel objective assessment of wrist range of movement, measured by the smartphone-embedded gyroscope and accelerometer. RESULTS Within 1 month of recruitment via social media campaigns, 399 participants self-enrolled, self-consented, and provided complete demographic data. Joint pain was the most frequently reported rheumatoid arthritis symptom to bother study participants (344/393, 87.5%). Severe patient-reported wrist pain appeared to be inversely linked with the range of wrist movement measured objectively by the app. At study entry, 292 of 399 participants (73.2%) indicated a preference for participating in a mobile app–based study. The number of participants in the study declined to 45 of 399 (11.3%) at week 12. CONCLUSIONS Despite the declining number of participants over time, the combination of social media and smartphone app with sensor integration was a feasible and cost-effective approach for the collection of patient-reported data in rheumatoid arthritis. Integral sensors within smartphones can be harnessed to provide novel end points, and the novel wrist range of movement test warrants further clinical validation.

This study presents a roadmap for converting California's all-purpose (electricity, transportation, heating/ cooling, and industry) energy infrastructure to one derived entirely from wind, water, and sunlight (WWS) generating electricity and electrolytic hydrogen. California's available WWS resources are first evaluated. A mix of WWS generators is then proposed to match projected 2050 electric power demand after all sectors have been electrified. The plan contemplates all new energy from WWS by 2020, 80e85% of existing energy converted by 2030, and 100% by 2050. Electrification plus modest efficiency measures may reduce California's end-use power demand ~44% and stabilize energy prices since WWS fuel costs are zero. Several methods discussed should help generation to match demand. A complete conversion in California by 2050 is estimated to create ~220,000 more 40-year jobs than lost, eliminate ~12,500 (3800 e23,200) state air-pollution premature mortalities/yr, avoid $103 (31e232) billion/yr in health costs, representing 4.9 (1.5e11.2)% of California's 2012 gross domestic product, and reduce California's 2050 global climate cost contribution by $48 billion/yr. The California air-pollution health plus global climate cost benefits from eliminating California emissions could equal the $1.1 trillion installation cost of 603 GW of new power needed for a 100% all-purpose WWS system within ~7 (4e14) years.

The social constructionist perspective has become a useful framework for understanding women’s psychological experiences. However, tools for examining how women construct meanings of their experiences as female in relevant social situations remain limited. Over two studies, our article describes the development and psychometric properties of the Subjective Femininity Stress Scale (SFSS), which assesses women’s stress associated with their subjective experiences of being female. Participants ( N = 468) described their personal experiences of what it means to be a woman by completing the sentence “As a woman…” 10 times; participants then rated how often each of the femininity experiences was stressful for them. Participants’ qualitative responses were coded based on 20 dimensions of femininity experiences, and their relationships with SFSS scores were examined. Study 1 provided evidence for test–retest reliability, convergent, and discriminant validity of the SFSS. Study 2 demonstrated concurrent and incremental validity of the SFSS. Several dimensions of femininity experiences were significantly correlated with women’s stress appraisal. The open-ended nature of the SFSS is easy to administer in clinical and organizational settings. The dual qualitative/quantitative aspects of the SFSS enable clients to create personally relevant meanings for their femininity experiences while also providing a stress index for exploring links with health outcomes.

Maintenance of hair follicle (HF) stem cell quiescence and self-renewal are key functions of a specific cellular niche represented by the HF bulge and adjacent cell populations. The unique context of this niche is crucial for normal HF functioning, but mechanisms implicated in its maintenance are still not quite clear. The Letter to the Editor by Sada et al. introduces a novel Slc1a3-CreER genetic mouse model which, in contrast to previously reported marking tools, selectively and highly efficiently demarcates the telogen bulge inner layer, one of the critical structural components of the bulge stem cell niche. The successful use of this tool identified the inner root sheath as a cellular source of telogen bulge inner layer and suggested its potential for further endeavors into fundamental and applied aspects of HF stem cell biology and club hair formation.

Electrospinning collagen with a synthetic polymer has been frequently investigated as a method of fabricating composite micro and nanofibers for tissue engineering applications. Using trifluoroacetic acid as a solvent for collagen and poly(L-lactic acid) (PLLA), electrospun fibers were well formed, and the filament surface was relatively smooth, with diameters averaging around 350 nm. Thermal analysis showed evidence of a PLLA phase with a well-developed enthalpy recovery peak along with a lower temperature for cold crystallization as compared to the unprocessed PLLA pellets. These results can be interpreted in terms of the non-equilibrium configuration of the solid state in the nanofilament as a consequence of the high elongational flow and rapid vitrification that occurs during electrospinning resulting in extended polymer chains for PLLA. In the solid state, this extended conformation allows PLLA to rapidly collapse to a more dense state. This conformation can also facilitate the formation of crystals with minimum segmental diffusion. Transmission electron microscope and solid extraction experiments suggest that phase separation has occurred, with an extended interphase, sheath-core morphology where PLLA forms the sheath, while collagen is in the core. This morphology can account for previous observations that collagen-polymer blends are not soluble in physiological fluids.

Existing research points toward dissonance as the primary catalyst in self-authorship development. This study investigated the cultural relevance of current conceptions of dissonance's role in self-authorship development. A total of 166 participants of color were recruited from three large public research universities from different regions in the United States. The results of our qualitative study suggest two kinds of dissonance lead to self-authorship development: Identity dissonance and relationship dissonance, and that type of dissonance experienced impacts developmental pathways. The implications of our results suggest that there is a need to further specify the current understandings of the catalysts and processes involved in self-authorship development.

Objectives: Endothelin-1 is a 21-amino acid peptide that together with specific receptors, A (ETrA) and B (ETrB) is induced following traumatic brain injury (TBI) and has been closely linked to regulation of cerebral vasospasm, oxidative stress, and hypoperfusion. Specific endothelin receptor antagonists have been shown to ameliorate early evidence of neuronal cell injury, activation of microglial cells, and hypoperfusion following TBI. The exact mechanism involved in TBI-induced hypoperfusion is still unclear; however, it is thought that endothelin-1 engagement of ETrA is primarily responsible for changes in blood flow. In this study we question the role of the microvascular pericyte in endothelin-1-mediated pathophysiology in TBI. Methods: Pericyte expression of endothelin-1, ETrA, and ETrB was examined in primary culture and in sham and impacted rat brain. Adult male rats were also given intracerebroventricular injections of ETrA (BQ-123) before being subjected to TBI using a closed head acceleration impact model. Results: Primary pericytes express both endothelin-1 and its receptors ETrA and ETrB. Following TBI, the number of alpha-smooth muscle actin (SMA) positive pericytes located in microvessels is significantly increased by 4 hours post-traumatic impact. Increases in pericyte expression of alpha-SMA correlated with evidence of a reduction in both arteriolar and capillary diameter. Capillary endothelin-1, ETrA, and ETrB transcript and protein was also increased. Increased endothelin-1 expression was seen by 2–4 hours postimpact. Upregulation of receptors was observed by 4–8 hours and maximum by 24 hours. ETrA antagonists decreased the number of alpha-SMA z pericytes as well as changes in microvascular diameter. Conclusion: These results suggest that decreased vasoconstriction following TBI may be due to an endothelin-1-induced pericyte-mediated regulation of microvessel blood flow following TBI. Furthermore, results suggest that ETrA antagonists ameliorate trauma induced hypoperfusion, in part, by inhibiting endothelin-1-mediated upregulation of alpha-SMA in pericytes.

Objective Ramelteon, an MT 1 /MT 2 melatonin receptor agonist, was evaluated for its ability to reduce sleep-onset difficulties associated with eastward jet travel. Methods Healthy adults ( n =110) with a history of jet lag sleep disturbances were flown eastward across five time zones from Hawaii to the east coast of the US. Ramelteon 1, 4, or 8mg or placebo was administered 5min before bedtime (local time) for four nights. Sleep parameters were measured using polysomnography (PSG) on Nights 2, 3, and 4. Next-day residual effects were assessed using psychomotor and memory function tests. Results Compared to placebo, there was a significant decrease in mean latency to persistent sleep (LPS) on Nights 2–4 with ramelteon 1mg (−10.64min, P =0.030). No consistent significant differences were observed with ramelteon vs. placebo on measures of next-day residual effects except on Day 4 where participants in all ramelteon groups performed significantly worse on the immediate memory recall test compared with placebo ( P ⩽0.05). The incidence of adverse events was similar for ramelteon and placebo. Conclusion After a 5-h phase advance due to eastward jet travel, ramelteon 1mg taken before bedtime for four nights reduced mean LPS relative to placebo in healthy adults.

A sensitive, selective, and quantitative method for the simultaneous determination of gemcitabine and 2,2-difluoro-2-deoxyuridine (dFdU) has been developed and validated in human plasma in the presence of tetrahydrouridine, a cytidine deaminase inhibitor. The method employs derivatization of gemcitabine and dFdU with dansyl chloride to improve the chromatographic retention and separation. The derivatization was performed in plasma without prior sample clean-up, followed by extraction of the dansyl-derivatives using methyl tertiary-butyl ether (MTBE). Ultra performance liquid chromatography (UPLC) technology on a BEH C18 stationary phase column with 1.7 microm particle size was used for chromatographic separation coupled to tandem mass spectrometry. The method was validated over the concentration ranges of 20-5000 and 100-25,000 ng/mL for gemcitabine and dFdU, respectively. The results from assay validation show that the method is rugged, precise, accurate, and well-suited to support pharmacokinetic studies. In addition, the relatively small sample volume (50 microL) and a run time of 1.5 min facilitate automation and allow for high-throughput analysis.

Ramelteon is an MT(1)/MT(2) melatonin receptor agonist indicated for the treatment of insomnia characterized by difficulty with sleep onset. In previous clinical studies, ramelteon reduced latency to persistent sleep (LPS) in subjects with chronic insomnia. The goal of the current analysis was to determine the average reduction in LPS and overall adverse event profile for subjects taking ramelteon 8 mg.This pooled analysis examined four randomized, double-blind, placebo-controlled clinical trials of ramelteon in subjects with chronic insomnia. The analysis included adults (age 18-83 years) with chronic insomnia who took ramelteon 8 mg or placebo. The primary endpoint of each trial was mean LPS, measured by polysomnography (PSG) on nights 1 and 2. Adverse events were collected for all subjects for the duration of each trial.Efficacy data were available for 566 subjects who took ramelteon 8 mg (mean age 46.7 years) and 556 subjects who took placebo (mean age 47.8 years). Mean LPS at baseline was 66.6 min for the placebo group and 66.9 min for the ramelteon group. At nights 1 and 2, mean LPS for the ramelteon 8 mg group (30.2 min) was significantly less than the mean LPS for the placebo group (43.3 min). The least squares mean difference from placebo was -13.1 min (p0.001). Headache (8.9% ramelteon 8 mg, 8.8% placebo) and somnolence (3.5% ramelteon 8 mg, 0.7% placebo) were the most common adverse events.Ramelteon 8 mg, on average, reduced LPS by approximately 13 min more than placebo on nights 1 and 2 of treatment in adults with chronic insomnia. Ramelteon was well tolerated with a low incidence of adverse events. This mean reduction in LPS versus placebo is similar to what has been reported for other classes of insomnia medications. However, these results reflect nights 1 and 2 of treatment and may not be representative of longer treatments.

To evaluate the effect of ramelteon on middle-of-the-night balance, mobility, and memory in older insomniacs.Thirty-three older adults (ageor = 65 years) with insomnia were enrolled in a single-dose, 3-way crossover study of balance after bedtime administration of ramelteon, 8 mg; zolpidem, 10 mg (positive control); or placebo. Subjects were administered study medication 30 minutes before bedtime and were awakened 2 hours after dosing to evaluate balance (Sensory Organization Test), turning speed and stability, memory (immediate and delayed word recall), and adverse events. There was a 4- to 10-day washout between treatments.Ramelteon or zolpidem (positive control) was compared with placebo. There were no differences between placebo and ramelteon on the Sensory Organization Test (p = 0.837), turn time (p = 0.776), or turn sway (p = 0.982). The positive control (zolpidem) did reveal significant impairments on the Sensory Organization Test, turn time, and turn sway (p0.001, all). Immediate and delayed memory recall were not significantly different with ramelteon (p = 0.683 and p = 0.650, respectively). Immediate recall declined significantly with zolpidem (p = 0.002). Adverse events were infrequent (ramelteon, n = 7; placebo, n = 7; zolpidem, n = 13); none were serious.In older adults, ramelteon did not impair middle-of-the night balance, mobility, or memory relative to placebo.

To assess the ability of repeated daily oral ramelteon to facilitate re-entrainment of human circadian rhythms after an imposed phase advance of the sleep-wake cycle.A total of 75 healthy adult volunteers aged 18-45 years remained in a sleep laboratory for 6 days and 5 nights; a 5-h phase advance in their sleep-wake cycle was imposed under dim-light conditions. Oral ramelteon (1,2, 4, or 8 mg once daily for 4 days) or placebo was administered 30 min before bedtime. The primary endpoint was the phase of the circadian rhythm as assessed by the time at which salivary melatonin concentrations declined below 3 pg/mL after morning awakening (dim-light melatonin offset [DLMoff]).After 4 days of once-daily treatment, participants receiving 1, 2, or 4 mg ramelteon exhibited statistically significant phase shifts in DLMoff of -88.0 (16.6), -80.5 (14.8), and -90.5 (15.2) minutes respectively, versus -7.1 (18.6) minutes for placebo (least-squares mean(SEM), p = 0.002, p = 0.003, p = 0.001, respectively). Change in DLMoff for the 8 mg dose of ramelteon, -27.9 (16.4) minutes, was not significantly different than that for placebo (p = 0.392).Ramelteon (1, 2, or 4 mg per day) administered before bedtime significantly advanced the phase of the circadian rhythm after a 5-h phase advance in the sleep-wake cycle. These findings suggest that ramelteon has potential as a specific therapy for circadian rhythm sleep disorders.

A highly sensitive and selective quantitative method to accurately determine testosterone (Te) and 5alpha-dihydrotestosterone (DHT) in human serum is crucial to the success of Te replacement therapy for hypogonadism. To this end we have developed and validated a semi-automated and relatively high-throughput method in a 96-well plate format using ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC/MS/MS) for the simultaneous determination of Te and DHT in human serum. Te and DHT along with the internal standards [(2)H(3)]-Te and [(2)H(3)]-DHT were extracted from 300 microL of human serum by liquid-liquid extraction using methyl tertiary-butyl ether (MTBE), followed by derivatization with 2,3-pyridinedicarboxylic anhydride and solid-phase extraction for sample clean up. A novel chemical derivatization approach using 2,3-pyridinedicarboxylic anhydride was employed to achieve the MS sensitivity and selectivity required for DHT. Baseline separation of Te and DHT derivatives from endogenous steroid derivatives was achieved using UPLC technology on a C18 stationary-phase column with 1.7 microm particle size. The validity of using double charcoal-stripped female human serum as surrogate matrix for preparation of calibration standards was demonstrated through standard addition experiments. The method was validated over the concentration ranges of 0.2-40 ng/mL for Te and 0.01-2 ng/mL for DHT. The validation and study sample analysis results show that the method is rugged, precise, accurate, and well suited to support pharmacokinetic studies where approximately 300 samples can be extracted and analyzed in 1 day.

This study examines the effect of a strike on the wealth of shareholders, and whether there is a difference in the effects during various phases of a stock market cycle. Although a significant loss of shareholders' value is observed at the beginning of a strike, most of the losses are recovered upon the settlement of the strike. During a bear market, the negative impact of a strike on a firm's value is more profound, and the firm does not fully recover its value when the strike ends. On the other hand, in a bull market, struck firms not only suffer a less severe loss but also experience a quicker rebound in their firms' value. Since the duration of a strike during a bear and bull market period is not significantly different, we conclude that the observed difference in the impact of a strike during different phases of a stock market cycle cannot be attributed to the difference in the duration of a strike.

Ramelteon, a selective MT(1)/MT(2) melatonin receptor agonist, was evaluated in subjects with mild to moderate chronic obstructive pulmonary disease (COPD) to determine whether it would have a negative effect on measures of safety and respiration. This randomized, double-blind, crossover study in 26 subjects with mild to moderate COPD compared the effects of a single bedtime dose of ramelteon 16 mg and placebo on sleep, oxygenation, and sleep-related abnormal breathing events. Compared with placebo, ramelteon had no statistically significant effect on mean arterial oxygen percent saturation (SaO(2)) for the entire night (92.9 vs 92.9%; 95% confidence interval [CI], -0.6 to 0.6; P = 0.972), for each of the 8 h of the night, for each sleep stage (awake, rapid eye movement, nonrapid eye movement) or for the percentage of the night that SaO(2) was less than 85 and 90%. The mean apnea-hypopnea index was similar between ramelteon and placebo groups (9.0 vs 8.3; 95% CI, -1.5 to 3.0; P = 0.515). Polysomnography documented a significant increase in total sleep time (380.6 vs 353.6, P = 0.015), sleep efficiency (79.3 vs 73.7, P = 0.017), and number of awakenings (11.1 vs 9.5, P = 0.036) with ramelteon vs placebo. Other polysomnography and subject-reported sleep measures were comparable between groups. Only one adverse event was reported; it was not considered treatment related. No clinically meaningful changes in laboratory test results, vital signs, electrocardiogram, and physical examination were observed. In this study, ramelteon 16 mg (two times the recommended therapeutic dose) showed no clinically meaningful or statistically significant effects on oxygenation or abnormal breathing events, was well tolerated, and improved sleep duration and efficiency in subjects with mild to moderate COPD.

Background: Ramelteon is a selective MT1/MT2 melatonin receptor agonist indicated for the treatment of insomnia characterized by difficulty with sleep onset. Objective: The current analysis was conducted to determine the effectiveness of ramelteon 8 mg in reducing the time to fall asleep in older adults with severe baseline sleep-onset difficulties. Methods: Patients with severe sleep-onset difficulty (defined as subjective sleep latency [sSL] ≥60 minutes) who had received ramelteon 8 mg or placebo were selected from a previously published multicenter outpatient trial of 829 older adults (aged ≥65 years) with primary, chronic insomnia (according to Diagnostic and Statistical Manual of Mental Disorders [Fourth Edition, Text Revision] criteria). Patients received single-blind placebo for 7 days (baseline) before receiving double-blind ramelteon 8 mg or placebo nightly for 5 weeks (35 nights). A 7-day, single-blind, placebo washout period followed. The primary end point was mean sSL for nights 1 through 7 (week 1). The mean changes in sSL from baseline at weeks 3 and 5 were evaluated to assess sustained efficacy. Adverse events (AEs) were collected in this analysis for both the ramelteon 8-mg and placebo groups. Results: A total of 157 patients from the rameltcon 8-mg group (mean age, 72.7 years; 87 women, 70 men) and 170 patients from the placebo group (mean age, 72.3 years; 111 women, 59 men) met the entry criteria for this post hoc analysis. Ramelteon 8 mg significantly reduced sSL at week 1 compared with placebo (change from baseline, -23.2 vs -7.5 minutes; P = 0.002). This statistically significant improvement was sustained at week 3 (-33.7 vs -19.8 minutes; P = 0.005) and week 5 (-37.4 vs -17.1 minutes; P < 0.001). The incidence of AEs was low. The most commonly reported treatment-emergent AEs were dizziness (ramclteon, 8.9%; placebo, 7.1%), dysgeusia (ramelteon, 7.0%; placebo, 2.9%), myalgia (ramelteon, 6.4%; placebo, 3.5%), and headache (ramelteon, 5.1%; placebo, 5.9%). Conclusions: In this subset analysis of older adults with severe baseline sleep-onset difficulties, ramelteon 8 mg significantly and persistently reduced subjective reports of time to sleep onset during 5 weeks of nightly treatment. Ramelteon appeared to be an effective and well-tolerated treatment for these older adults with primary, chronic insomnia.

Two semi-automated, relatively high throughput methods using ultra performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) were developed for the simultaneous determination of ethinyl estradiol (EE) in combination with either 19-norethindrone (NE) or levonorgestrel (LN) in human plasma. Using 300 microL plasma, the methods were validated over the concentration ranges of 0.01-2 ng/mL and 0.1-20 ng/mL for EE and NE (or LN), respectively. The existing methods for the determination of the oral contraceptives in human plasma require large volumes of plasma (or =500 microL), and sample extraction is labor-intensive. The LC run time is at least 6 min, enabling analysis of only about 100 samples a day. In the present work the throughput was greatly improved by employing a semi-automated sample preparation process involving liquid-liquid extraction and derivatization with dansyl chloride followed by UPLC separation on a small particle size column achieving a run time of 2.7 min. The validation and actual sample analysis results show that both methods are rugged, precise, accurate, and well suitable to support pharmacokinetic studies where approximately 300 samples can be extracted and analyzed in a day.

Ramelteon is a selective MT(1)/MT(2)-receptor agonist indicated for insomnia treatment. Because it has no depressant effects on the nervous system, it is not expected to affect the control of breathing. The potential effects of ramelteon on apneic and hypopneic events and arterial oxygen saturation (SaO(2)) in individuals with obstructive sleep apnea were assessed. In this double-blind, randomized, crossover study, 26 adults with mild to moderate obstructive sleep apnea received ramelteon 16 mg and placebo for one night each, with a 5- to 12-day washout period between treatments. Treatments were administered 30 min before habitual bedtime. Respiratory effort was monitored using respiratory inductance plethysmography, SaO(2) was measured by pulse oximetry, and sleep onset and duration were measured by polysomnography and post-sleep questionnaire. Post-sleep questionnaire also measured next-day residual effects. The primary measure was apnea-hypopnea index. Apnea-hypopnea index was similar in ramelteon and placebo groups (11.4 vs 11.1, respectively; CI = -2.1, 2.6, P = 0.812). Ramelteon had no effect on the number of central, obstructive, or mixed apnea episodes. No significant differences were observed in SaO(2) for the entire night between ramelteon and placebo (95.1 vs 94.7%; P = 0.070). Ramelteon did not meaningfully affect sleep when evaluated by polysomnography and post-sleep questionnaire. Compared with placebo, ramelteon had no significant effect on next-day residual effects. Adverse events were reported by three subjects in the ramelteon group: headache (n = 2) and urinary tract infection (n = 1). No adverse events were reported with placebo. Ramelteon was well-tolerated and, as expected, did not worsen sleep apnea when administered to subjects with mild to moderate obstructive sleep apnea.

A stable isotopically labeled (SIL) analogue is believed to be the most appropriate internal standard in a quantitative bioanalytical liquid chromatography/tandem mass spectrometry (LC/MS/MS) assay. It is assumed that a SIL internal standard always compensates for variability in chemical derivatization, sample extraction and LC/MS/MS analysis due to its nearly identical chemical and physical properties to the unlabeled analyte. Hence, the analyte to internal standard peak area ratio should be constant despite any variations in sample processing or analysis. However, in our laboratories, a deuterium labeled internal standard of carvedilol demonstrated an unexpected behavior-the analyte to internal standard peak area ratio changed with two specific lots of commercially supplied human plasma. Several experiments, including dilution of the extract with LC mobile phase and post-column infusion of the carvedilol solution followed by the injection of extracted blank plasma, have indicated that a high level of matrix suppression affected the ionization of the carvedilol-S enantiomer and its deuterated internal standard differently in these two lots of plasma. For the first time, it was clearly demonstrated that a slight difference in retention time between the analyte and the SIL internal standard, caused by deuterium isotope effect, has resulted in a different degree of ion suppression between these two analogues. This difference was significant enough to change the analyte to internal standard peak area ratio and affect the accuracy of the method.