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Context In an open-label, randomized, controlled, phase 3 trial in 61 children aged 1 to 12 years with X-linked hypophosphatemia (XLH), burosumab improved rickets vs continuing conventional therapy with active vitamin D and phosphate. Objective We conducted an analysis to determine whether skeletal responses differed when switching to burosumab vs continuing higher or lower doses of conventional therapy. Methods Conventional therapy dose groups were defined as higher-dose phosphate [greater than 40 mg/kg] (HPi), lower-dose phosphate [40 mg/kg or less] (LPi), higher-dose alfacalcidol [greater than 60 ng/kg] or calcitriol [greater than 30 ng/kg] (HD), and lower-dose alfacalcidol [60 ng/kg or less] or calcitriol [30 ng/kg or less] (LD). Results At week 64, the Radiographic Global Impression of Change (RGI-C) for rickets was higher (better) in children randomly assigned to burosumab vs conventional therapy for all prebaseline dose groups: HPi (+1.72 vs +0.67), LPi (+2.14 vs +1.08), HD (+1.90 vs +0.94), LD (+2.11 vs +1.06). At week 64, the RGI-C for rickets was also higher in children randomly assigned to burosumab (+2.06) vs conventional therapy for all on-study dose groups: HPi (+1.03), LPi (+1.05), HD (+1.45), LD (+0.72). Serum alkaline phosphatase (ALP) also decreased in the burosumab-treated patients more than in the conventional therapy group, regardless of on-study phosphate and active vitamin D doses. Conclusion Prior phosphate or active vitamin D doses did not influence treatment response after switching to burosumab among children with XLH and active radiographic rickets. Switching from conventional therapy to burosumab improved rickets and serum ALP more than continuing either higher or lower doses of phosphate or active vitamin D.

SummaryDuring vertebrate embryogenesis, axial tendons develop from the paraxial mesoderm and differentiate through specific developmental stages to reach the syndetome stage. While the main roles of signaling pathways in the earlier stages of the differentiation have been well established, pathway nuances in syndetome specification from the sclerotome stage have yet to be explored. Here, we show stepwise differentiation of human iPSCs to the syndetome stage using chemically defined media and small molecules that were modified based on single cell RNA-sequencing and pathway analysis. We identified a significant population of branching off-target cells differentiating towards a neural phenotype overexpressing Wnt. Further transcriptomics post-addition of a WNT inhibitor at the somite stage and onwards revealed not only total removal of the neural off-target cells, but also increased syndetome induction efficiency. Fine-tuning tendon differentiationin vitrois essential to address the current challenges in developing a successful cell-based tendon therapy.

The type III secretion system (T3SS) is a large, transmembrane protein machinery used by various pathogenic gram‐negative bacteria to transport virulence factors into the host cell during infection. Understanding the structure of T3SSs is crucial for future developments of therapeutics that could target this system. However, much of the knowledge about the structure of T3SS is available only for Salmonella, and it is unclear how this large assembly is conserved across species. Here, we combined cryo‐electron microscopy, cross‐linking mass spectrometry, and integrative modeling to determine the structure of the T3SS needle complex from Shigella flexneri. We show that the Shigella T3SS exhibits unique features distinguishing it from other structurally characterized T3SSs. The secretin pore complex adopts a new fold of its C‐terminal S domain and the pilotin MxiM[SctG] locates around the outer surface of the pore. The export apparatus structure exhibits a conserved pseudohelical arrangement but includes the N‐terminal domain of the SpaS[SctU] subunit, which was not present in any of the previously published virulence‐related T3SS structures. Similar to other T3SSs, however, the apparatus is anchored within the needle complex by a network of flexible linkers that either adjust conformation to connect to equivalent patches on the secretin oligomer or bind distinct surface patches at the same height of the export apparatus. The conserved and unique features delineated by our analysis highlight the necessity to analyze T3SS in a species‐specific manner, in order to fully understand the underlying molecular mechanisms of these systems. The structure of the type III secretion system from Shigella flexneri delineates conserved and unique features, which could be used for the development of broad‐range therapeutics.

Introduction Lubiprostone capsules are approved for managing three different chronic constipation conditions. A “sprinkle” formulation may facilitate use in individuals with difficulty swallowing capsules. Our objective was to evaluate the bioequivalence, pharmacokinetics (PK), and bioavailability of lubiprostone sprinkles vs lubiprostone capsules, compared with placebo. Methods A 1-week randomized, placebo-controlled, double-blinded, bioequivalence study (study 302) and a single-dose PK and bioavailability crossover study (study 304) were conducted. In study 302, 522 subjects with chronic constipation were randomized to lubiprostone sprinkle 24 μg twice daily (BID), lubiprostone capsule 24 μg BID, or placebo. The primary efficacy endpoint was observed spontaneous bowel movement (SBM) counts (equivalence defined as showing the 90% confidence interval [CI] of the “between-group SBM ratio” to be contained within 0.8–1.25). Study 304 included two cohorts of healthy volunteers randomized to a single 48-μg lubiprostone dose, sprinkle, or capsule (n = 35) or to a single 48-μg sprinkle dose, in fed or fasted state (n = 14). Results Both lubiprostone formulations significantly improved SBM count (sprinkle, 4.82 ± 3.66, P = 0.002; capsule, 5.74 ± 3.79, P

Background Cranial bone loss presents a major clinical challenge and new regenerative approaches to address craniofacial reconstruction are in great demand. Induced pluripotent stem cell (iPSC) differentiation is a powerful tool to generate mesenchymal stem cells (MSCs). Prior research demonstrated the potential of bone marrow-derived MSCs (BM-MSCs) and iPSC-dervied mesenchymal progenitor cells via the neural crest (NCC-MPCs) or mesodermal lineages (iMSCs) to be a promising cell source for bone regeneration. Overexpression of human recombinant bone morphogenetic protein (BMP)6 efficiently stimulates bone formation. The goal of this study was to evaluate the potential of iPSC-derived stem cells via neural crest or mesoderm overexpressing BMP6 and embedded in 3D printable bio-ink to generate viable bone graft alternatives for cranial reconstruction. Methods Cell viability and osteogenic potential of cells and bio-ink (Ink-Bone or GelXa) combinations were investigated in vitro using bioluminescent imaging. Osteogenic potential of bio-ink-cell constructs was evaluated in osteogenic media or nucleofected with BMP6 using qRT-PCR and in vitro µCT. For in vivo testing, two 2mm circular defects were created in the frontal and parietal bones of NOD/SCID mice and treated with Ink-Bone, Ink-Bone + BM-MSC-BMP6, Ink-Bone + iMSC-BMP6 or Ink-Bone + iNCC-MPC-BMP6 or left untreated. For follow up, µCT was performed at weeks 0, 4 and 8 weeks. At time of (week 8), histological and immunofluorescent analyses were performed. Results Both bio-inks supported cell survival and promoted osteogenic differentiation of iNCC-MPCs and BM-MSCs in vitro. At 4 weeks, cell viability of both BM-MSCs and iNCC-MPCs were increased in Ink-Bone compared to GelXA. Combination of Ink-Bone with iNCC-MPC-BMP6 resulted in an increased bone volume in the frontal bone compared to the other groups at 4 weeks post-surgery. At 8 weeks, both iNCC-MPC-BMP6 and iMSC-MSC-BMP6 resulted in an increased bone volume and partial bone bridging between implant and host bone compared to the other groups. Conclusions The results of this study show the potential of NCC-MPC-incorporated bio-ink to regenerate frontal cranial defects. Therefore, this bio-ink-cell combination should be further investigated for its therapeutic potential in large animal models with larger cranial defects, allowing for 3D printing of the cell-incorporated material.

Background and objectives With multiple medications indicated for mineral metabolism, dialysis providers can apply various strategies to achieve target phosphate and parathyroid hormone (PTH) levels. We describe common prescribing patterns and practice variation in mineral metabolism treatment strategies over the last decade. Design, setting, participants, & measurements In a cohort of adults initiating hemodialysis at Dialysis Clinic, Inc. facilities, we assessed prescriptions of vitamin D sterols, phosphate binders, and cinacalcet longitudinally. To identify the influence of secular trends in clinical practice, we stratified the cohort by dialysis initiation year (2006–2008, 2009–2011, and 2012–2015). To measure practice variation, we estimated the median odds ratio for prescribing different mineral metabolism treatment strategies at 12 months post–dialysis initiation across facilities using mixed effects multinomial logistic regression. Sensitivity analyses evaluated strategies used after detection of first elevated PTH. Results Among 23,549 incident patients on hemodialysis, there was a decline in vitamin D sterol–based strategies and a corresponding increase in strategies without PTH-modifying agents (i.e., phosphate binders alone or no mineral metabolism medications) and cinacalcet-containing treatment strategies between 2006 and 2015. The proportion with active vitamin D sterol–based strategies at dialysis initiation decreased across cohorts: 15% (2006–2008) to 5% (2012–2015). The proportion with active vitamin D sterol–based strategies after 18 months of dialysis decreased across cohorts: 52% (2006–2008) to 34% (2012–2015). The odds of using individual strategies compared with reference (active vitamin D sterol with phosphate binder) varied from 1.5- to two-fold across facilities in 2006–2008 and 2009–2011 cohorts, and increased to two- to three-fold in the 2012–2015 cohort. Findings were similar in sensitivity analyses starting from first elevated PTH measurement. Conclusions Over time, mineral metabolism management involved less use of vitamin D sterol–based strategies, greater use of both more conservative and cinacalcet-containing strategies, and increased practice variation, suggesting growing equipoise.

Background and hypothesis Post-traumatic elbow contracture remains a common and challenging complication with often unsatisfactory outcomes. Although the etiology is unknown, elevated or abnormal post-fracture synovial fluid cytokine levels may result in the migration of fibroblasts to the capsule and contribute to capsular pathology. Thus, the purpose of this study was to characterize the cytokine composition in the synovial fluid fracture hematoma of patients with intra-articular elbow fractures. Methods The elbow synovial fluid fracture hematoma of 11 patients with intra-articular elbow fractures was analyzed for CTXII (C-terminal telopeptides of type II collagen [a cartilage breakdown product]) as well as 15 cytokines and matrix metalloproteinases (MMPs) including interferon γ, interleukin (IL) 1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, tumor necrosis factor α, MMP-1, MMP-2, MMP-3, MMP-9, and MMP-10. The uninjured, contralateral elbow served as a matched control. Mean concentrations of each factor were compared between the fluid from fractured elbows and the fluid from control elbows. Results The levels of 14 of 15 measured cytokines and MMPs—interferon γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, tumor necrosis factor α, MMP-1, MMP-3, MMP-9, and MMP-10—were significantly higher in the fractured elbows. In addition, post hoc power analysis revealed that 10 of 14 significant differences were detected with greater than 90% power. The mean concentration of CTXII was not significantly different between groups. Conclusions These results demonstrate a proinflammatory environment after fracture that may be the catalyst to the development of post-traumatic elbow joint contracture. The cytokines with elevated levels were similar, although not identical, to the cytokines with elevated levels in studies of other weight-bearing joints, indicating the elbow responds uniquely to trauma.

Context Younger age at treatment onset with conventional therapy (phosphate salts and active vitamin D; Pi/D) is associated with improved growth and skeletal outcomes in children with X-linked hypophosphatemia (XLH). The effect of age on burosumab efficacy and safety in XLH is unknown. Objective This work aimed to explore the efficacy and safety of burosumab vs Pi/D in younger ( Methods This post hoc analysis of a 64-week, open-label, randomized controlled study took place at 16 academic centers. Sixty-one children aged 1 to 12 years with XLH (younger, n = 26; older, n = 35) participated. Children received burosumab starting at 0.8 mg/kg every 2 weeks (younger, n = 14; older, n = 15) or continued Pi/D individually titrated per recommended guidelines (younger, n = 12; older, n = 20). The main outcome measure included the least squares means difference (LSMD) in Radiographic Global Impression of Change (RGI-C) rickets total score from baseline to week 64. Results The LSMD in outcomes through 64 weeks on burosumab vs conventional therapy by age group were as follows: RGI-C rickets total score (younger, +0.90; older, +1.07), total Rickets Severity Score (younger, −0.86; older, −1.44), RGI-C lower limb deformity score (younger, +1.02; older, +0.91), recumbent length or standing height Z-score (younger, +0.20; older, +0.09), and serum alkaline phosphatase (ALP) (younger, −31.15% of upper normal limit [ULN]; older, −52.11% of ULN). On burosumab, dental abscesses were not reported in younger children but were in 53% of older children. Conclusion Burosumab appears to improve outcomes both in younger and older children with XLH, including rickets, lower limb deformities, growth, and ALP, compared with Pi/D.

The Dragons of Inaction Psychological Barrier (DIPB) scale, a measure of psychological barriers, was developed and validated in three studies across six environmental domains: food choices, transportation, energy use, water use, purchasing, and waste. A preliminary 65-item version was created based on past studies. In Study 1, it was examined using a large sample with exploratory factor analysis; five factors emerged. In Study 2, using a different sample, the factors were supported with confirmatory factor analyses. In Study 3, measurement invariance of the scale was supported across high and low difficulty behaviors. The final 22-item DIPB includes five subscales: Change Unnecessary, Conflicting Goals and Aspirations, Interpersonal Relations, Lacking Knowledge, and Tokenism. No significant DIPB differences across environmental domains were found for Tokenism, but the Change Unnecessary barrier was weakest for buying green, whereas the Interpersonal Relations barrier was strongest for food choices, the Conflicting Goals and Aspirations barrier was strongest for driving less, and the Lacking Knowledge barrier was strongest for buying green.

The development of antibodies that target specific glycan structures on cancer cells or human pathogens poses a significant challenge due to the immense complexity of naturally occurring glycans. Automated glycan assembly enables the production of structurally homogeneous glycans in amounts that are difficult to derive from natural sources. Nanobodies (Nbs) are the smallest antigen-binding domains of heavy-chain-only antibodies (hcAbs) found in camelids. To date, the development of glycan-specific Nbs using synthetic glycans has not been reported. Here, we use defined synthetic glycans for alpaca immunization to elicit glycan-specific hcAbs, and describe the identification, isolation, and production of a Nb specific for the tumor-associated carbohydrate antigen Globo-H. The Nb binds the terminal fucose of Globo-H and recognizes synthetic Globo-H in solution and native Globo-H on breast cancer cells with high specificity. These results demonstrate the potential of our approach for generating glycan-targeting Nbs to be used in biomedical and biotechnological applications.

Summary Introduction Bladder bowel dysfunction (BBD), defined by the International Children's Continence Society (ICCS) as a spectrum of lower urinary tract and bowel symptoms, represents to up to 40% of pediatric urology consults. Management of BBD involves strict behavioral management with frequent follow ups by urology advanced practice providers (APPs). If left untreated, patients may develop secondary comorbidities that impact their renal and/or bladder function, bowel function, and psychosocial well-being. Previous studies have reported feasibility for virtual post-operative visits and prenatal consultations, however, telehealth management of BBD, or TeleBBD, has not yet been studied. The goal of this study is to survey APPs in pediatric urology to understand how TeleBBD compares to in-person visits, and identify benefits and limitations of TeleBBD. Study design An online survey via Qualtrics was designed based on current practice guidelines for BBD management and telehealth considerations. Survey was distributed in September 2020 via the Pediatric Urology Nurses & Specialists listserv. Group qualitative coding was completed by the authors to generate themes that emerged from the results. Results/Discussion A total of 53 APPs from across 21 states in the US completed the online survey, with 49 (92%) APPs reporting currently providing TeleBBD. Those who did not provide TeleBBD typically do not manage BBD patients. Summary Table shows the comparison of TeleBBD with in-person visits, with many elements of TeleBBD better or the same as in-person visits. APPs reported that TeleBBD has been most beneficial in increasing access and overall improvement in follow up which is significant for management of this chronic condition. Barriers include access to technology/internet and inability to perform full physical exams via televisit. Limitations of the study include lack of validated survey and small sample size. Overall increased access and improved patient adherence and resolution from BBD is significant for this patient population for prevention of secondary comorbidities. Conclusion This is the first study to survey APPs across the US specifically exploring indication of TeleBBD as well as comparison of TeleBBD with in-person visits. The perception of improved access to care and less no-show rates is significant especially during the pandemic in providing continuity of care and prevent secondary comorbidities. Additionally, TeleBBD was felt to be just as effective as in-person visits for patient adherence to treatment, and other components of care, with the exception of ability to provide physical exams. Providers leveraged the benefits of TeleBBD and shared strategies for best practices. Summary Table . APP Perspective of TeleBBD Compared to In-Person Visits (n = 49). Question Better than in-person visits % (n) Same as in-person visits % (n) Worse than in-person visits % (n) Total response per question (n) Frequency of follow up visits 50.00% (22) 47.73% (21) 2.27% (1) 44 No show rates 47.73% (21) 45.45% (20) 6.82% (3) 44 Patient/Parent/Caretaker satisfaction 29.27% (12) 68.29% (28) 2.44% (1) 41 Adherence to treatment regimen 7.32% (3) 80.49% (33) 12.20% (5) 41 Ability to discuss recommendations and teaching with parents/caretakers 6.82% (3) 79.55% (35) 13.64% (6) 44 Ability to collect history 6.98% (3) 79.07% (34) 13.95% (6) 43 Timing to resolution/discharge from clinic 17.65% (6) 67.65% (23) 14.71% (5) 34 Ability to review studies/imaging with patients/parents/caretakers 13.64% (6) 63.64% (28) 22.73% (10) 44 Ability to discuss recommendations and teaching with patients 9.09% (4) 68.18% (30) 22.73% (10) 44 Ability to perform PE 0.00% (0) 0.00% (0) 100.00% (42) 42 PE: Physical Examination.

Supplemental Digital Content is Available in the Text., Clinical training is a key component of nurse practitioner (NP) education. The rapid shift to telehealth necessitated by the pandemic has also created a need for socially distanced education and precepting. This article presents teleprecepting as a modality for clinical opportunities and connecting students to previously limited experiences, such as training in small specialties, in rural areas, and with interdisciplinary teams. Precepting NP students using telehealth follows similar principles as in-person teaching, but some modifications and additional considerations are needed to transition to the virtual environment. At a time when many NPs have swiftly adapted to telehealth in practice, this article will offer a brief “how-to” for teleprecepting. Prior to COVID-19, teleprecepting was piloted with less than 2% of NP students in the school's pilot teleprecepting project. Seven months after the initial surge of cases and restrictions, 72% of students (n = 151) in the family nurse practitioner, psychiatric mental health nurse practitioner, and pediatric nurse practitioner specialties had transitioned to teleprecepting. This project was implemented rapidly during the pandemic, and thus, evaluations comparing competency outcomes and experiences of students and preceptors are still in process. Additionally, feasibility of this educational model may change as telehealth regulations continue to evolve. COVID-19 poses challenges for both patient care and clinical training of NP students across specialties. With some adaptation, clinical placements can be transitioned to the virtual environment of telehealth. Future studies should examine student competencies based on teleprecepting experiences and preceptor training to support teleprecepting roles.

DRAM scaling has been slowed down. Emerging non-volatile memories (e.g., Phase-Change Memory) promises higher density, better scalability, and persistence. However, endurance is a fundamental issue that hinders the broad adoption of PCM---after repeated writes, a PCM cell can get stuck at a value and be no longer programmable. The prevalence of this stuck-at fault issue requires error detection and correction mechanisms for PCM. Existing solutions such as verify-after-write adds additional latency to PCM writes, which degrades overall system performance. Other solutions like in-memory error-correcting code (ECC) requires a high storage overhead and introduces more reliability issue because ECC bits tend to wear out faster than the protected data bits. In this paper, a novel stuck-at faults detection technique is proposed to improve performance and reliability simultaneously. Since stuck-at faults can only be detected after new writes, ECC does not need to be stored permanently and can be deleted immediately after a one-time detection, which helps to reduce ECC storage overhead. Therefore, this work proposes to use a small on-chip ECC cache to store the temporary ECC entries, which does not suffer from endurance issue. To maximize the utilization of the limited cache space, this work optimizes ECC entry insertion and deletion mechanisms and exploits memory bank-level parallelism to minimize performance impact. For the evaluated workloads, the proposed ECC cache achieves an average of 9.4% of the performance improvement over the baseline with a verify-after-write detection.

Changing to burosumab, a monoclonal antibody targeting fibroblast growth factor 23, significantly improved phosphorus homeostasis, rickets, lower-extremity deformities, mobility, and growth versus continuing oral phosphate and active vitamin D (conventional therapy) in a randomized, open-label, phase 3 trial involving children aged 1–12 years with X-linked hypophosphatemia. Patients were randomized (1:1) to subcutaneous burosumab or to continue conventional therapy. We present patient-reported outcomes (PROs) from this trial for children aged ≥ 5 years at screening (n = 35), using a Patient-Reported Outcomes Measurement Information System (PROMIS) questionnaire and SF-10 Health Survey for Children. PROMIS pain interference, physical function mobility, and fatigue scores improved from baseline with burosumab at weeks 40 and 64, but changed little with continued conventional therapy. Pain interference scores differed significantly between groups at week 40 (− 5.02, 95% CI − 9.29 to − 0.75; p = 0.0212) but not at week 64. Between-group differences were not significant at either week for physical function mobility or fatigue. Reductions in PROMIS pain interference and fatigue scores from baseline were clinically meaningful with burosumab at weeks 40 and 64 but not with conventional therapy. SF-10 physical health scores (PHS-10) improved significantly with burosumab at week 40 (least-squares mean [standard error] + 5.98 [1.79]; p = 0.0008) and week 64 (+ 5.93 [1.88]; p = 0.0016) but not with conventional therapy (between-treatment differences were nonsignificant). In conclusion, changing to burosumab improved PRO measures, with statistically significant differences in PROMIS pain interference at week 40 versus continuing with conventional therapy and in PHS-10 at weeks 40 and 64 versus baseline.Trial registration: ClinicalTrials.gov NCT02915705

In XLH, excess circulating FGF23 causes hypophosphatemia, rickets, lower limb deformity, and impaired growth and mobility. An active-controlled, phase 3 trial (CL301; NCT02915705) showed treatment with burosumab, a fully human monoclonal antibody against FGF23, resulted in significantly greater improvements in all of these outcomes in children with XLH, compared with continuing oral phosphate and active vitamin D as conventional therapy (Pi/D) per established guidelines. In a post-hoc analysis, we compared children who received burosumab vs those who received an average 64-week oral phosphate daily dose >40 mg/kg (higher-dose Pi) vs ≤40 mg/kg (lower-dose Pi). Sixty-one children with XLH (1–12 years-old) were randomized 1:1 after a 7-day Pi/D washout to receive burosumab (n=29) starting at 0.8 mg/kg subcutaneously Q2W or to resume Pi/D (n=32) titrated by their investigator, for 64 weeks. Eligibility criteria included Rickets Severity Score (RSS) ≥2.0 despite prior Pi/D treatment. Of the 32 subjects randomized to Pi/D, 12 received average higher-dose Pi and 20 received average lower-dose Pi (as specified above). The primary endpoint was rickets healing, using the Radiographic Global Impression of Change (RGI-C) Scale. At week 64, the improvement in the least square (LS) mean (LS mean [±SE; 95%CI]) RGI-C Global Score for rickets was greater on burosumab (+2.06 [0.072; 1.92, 2.20]) compared with either higher-dose (+1.02 [0.241; 0.55, 1.50]) or lower-dose (+1.04 [0.162; 0.73, 1.36]) Pi. The mean decrease in the total RSS from baseline was also greater on burosumab (-2.23 [0.117; -2.46, -2.00]) compared with higher-dose (-0.87 [0.264; -1.39, -0.35] or lower-dose (-1.09 [0.180; -1.45, -0.74]) Pi. Similarly, the mean RGI-C Lower Limb Deformity Score was greater on burosumab (+1.25 [0.170; 0.92, 1.59]) compared with either higher-dose (+0.32 [0.188; -0.05, 0.69]) or lower-dose (+0.26 [0.146; -0.02, 0.55]) Pi. Adverse events including hypersensitivity and injection site reactions, were more frequent with burosumab, and were mild to moderate in severity overall. No discontinuations occurred. In conclusion, children with XLH treated with burosumab had greater improvements in rickets and lower limb deformity compared with subjects receiving higher or lower doses of Pi.

Quantitative cross-linking/mass spectrometry (QCLMS/QXL-MS) probes structural changes of proteins in solution. This method has revealed induced conformational changes, composition shifts in conformational ensembles and changes in protein interactions. It illuminates different structural states of proteins or protein complexes by comparing which residue pairs can be cross-linked in these states. Cross-links provide information about structural changes that may be inaccessible by alternative technologies. Small local conformational changes affect relative abundances of nearby cross-links, whereas larger conformational changes cause new cross-links to be formed. Distinguishing between minor and major changes requires a robust analysis based on carefully selected replicates and, when using isotope-labeled cross-linkers, replicated analysis with a permutated isotope-labeling scheme. A label-free workflow allows for application of a wide range of cross-linking chemistries and enables parallel comparison of multiple conformations. In this protocol, we demonstrate both label-free and isotope-labeled cross-linker-based workflows using the cross-linker bis(sulfosuccinimidyl)suberate (BS3). The software XiSearch, developed by our group, is used to identify cross-linked residue pairs, although the workflow is not limited to this search software. The open-access software Skyline is used for automated quantitation. Note that additional manual correction greatly enhances quantitation accuracy. The protocol has been applied to purified multi-protein complexes but is not necessarily limited to that level of sample complexity. Optimizing the cross-linker/protein ratio and fractionating peptides increase the data density of quantified cross-links, and thus the resolution of QCLMS. The entire procedure takes ~1-3 weeks.

Author(s): Pierstorff, Erik; Chen, Shanshan; Chaparro, Maria Paola; Cortez, John M; Chen, Yen-Jung; Ryu, Su Young; Tsai, Sherry M; Baum, Marc M; Yang, Wan Wan; Kalinec, Federico; Smith, Thomas; Ludwig, Stacey; Slattery, William H | Abstract: ObjectiveInvestigate a new polymer-based drug coating suitability for safe intracochlear delivery and ability to maintain long-term physiologically active levels of the corticosteroid fluticasone propionate.Study designIn vitro dissolution study to evaluate release profiles of polymer-coated drug particles and in vivo studies using a guinea pig model to measure perilymph drug concentrations at specific time points after implantation with polymer-coated drug particles and evaluate their effect on hearing function.MethodsPolymer-coated fluticasone propionate (FP) particles were surgically implanted in guinea pigs through the round window membrane into the cochlear scala tympani. In the pilot study, pre- and post-op hearing thresholds were conducted on days 7, 14, and 42. In a second study, post-op hearing thresholds were conducted on days 90, 120, and 180. Perilymph drug concentrations were measured on the same time points.ResultsIn 15 of 16 animals from day 7 through day 90, drug levels were within the targeted range, with no initial burst release detected. Drug was present in all animals on day 90 and was detected in some animals at 120 and 180 days. Hearing was tested and compared with non-implanted ears. Very good hearing preservation was observed in ears implanted with intracochlear particles when compared with contralateral ears.ConclusionsThe polymer-based extended release system is effective in providing long-term, stable drug delivery for at least 90 days with good hearing outcomes. The results of this study support the potential for achieving long-term drug delivery with a single intracochlear administration.

Background and purpose Massive Open Online Courses (MOOCs) offer an innovative approach to pharmacy education and are expected to challenge traditional pedagogy and foundational knowledge acquisition practices. A survey of the literature reveals no current publications describing implementation of MOOCs in pharmacy education and limited information about MOOC implementation in other healthcare disciplines in the United States. Educational activity and setting A few colleges of pharmacy (COPs) and other health professions’ educational programs have recently started offering MOOCs. Findings Herein we provide an overview of MOOCs and describe the early implementation stages of MOOCs being conducted at two COPs, an interprofessional MOOC, and a variety of MOOCs offered by a public health program. This overview and the four case studies on MOOC implementation in healthcare education provide practical information about course development, descriptions of selected course engagement outcomes, insight into lessons learned by the institutions, and practical considerations for development of future MOOCs. Discussion MOOCs prompt diversification of models of teaching and learning, transformation of pedagogical frameworks, and innovation in the scholarship of teaching and learning. Summary MOOCs offer exciting opportunities to distribute knowledge on a massive and global scale to a diverse population of learners.

INTRODUCTION: The incidence of CRC continues to increase at a rate of 2 1% per year in both men and women Screening for CRC improves prognosis by identifying early-stages for treatment, resulting in lower mortality Compliance rates for CRC screening have been low, and in optimized clinical settings, range from 50% to 70% Fecal Immunochemical Test (FIT) has advantages over colonoscopy as it does not require bowel preparation and anesthesia, and is an easier test to complete We hypothesize that CRC screening rates at our academic community hospital-affiliated clinic will improve by increasing resident education about FIT METHODS: We reviewed the charts of patients aged 50-75 seen by resident physicians at our clinic from June 1, 2019 to December 31, 2019 Patients who did not have a follow-up or comprehensive visit within the mentioned timeframe, had an active diagnosis of CRC or history of colorectal surgery, or with risk factors placing them at high lifetime risk for CRC were excluded Variables included patient age, gender, ethnicity, colonoscopy referral, FIT order, and reasons why CRC screening was not offered Resident physicians were given an educational presentation and an informational e-mail addressing self-reported barriers to ordering FIT for patients We then conducted a post-intervention chart review of patients aged 50-75 evaluated by residents from January 1, 2020 to March 31, 2020 and assessed the association of each variable with multivariate logistic regression RESULTS: There was no significant difference in age, gender, ethnicity and overall CRC screening rates between the pre- and post-intervention groups (Figure 1 ) There was a significant increase in FIT testing over colonoscopy (P=0 012, OR=2 10, 95% CI: 1 104-3 999) and in CRC screening rates in unscreened patients post-intervention (P=0 037, OR=1 55, 95% CI: 0 960-2 501) (Figure 2 ) There was a significant reduction in amount of resident physicians forgetting to screen patients (P=0 003, OR=4 44, 95% CI: 1 664-11 82) (Figure 3 ) CONCLUSION: Education improved the utilization of FIT testing over colonoscopy as well as the overall CRC screening rates in previously unscreened patients at our outpatient clinic Recent EHR data showed a decrease of approximately 86% in CRC screenings across the U S - presumably due to access disruptions due to COVID-19 However, our clinic after resident education shifted towards theutilization of FIT, and ultimately increasing our CRC screening rates for previously unscreened patients (Figure Presented)

Objectives Surgical treatment of proximal biceps tendinopathy remains a source of great controversy. Current surgical treatment debates center around the fixation location and not the quality of the tendon because little is known about the intra-tendinous pathology to guide this debate. Therefore, this study aims to evaluate two distinct anatomic regions of pathologic biceps tendons with regard to intra-tendinous architecture, painful neurotransmitters, and inflammatory modulators: the intra-articular portion and the extra-articular portion lying in the bicipital groove. Methods Eleven human degenerative biceps tendons were obtained post-tenodesis. They were divided into intra- and extra-articular portions. They underwent histologic staining with hematoxylin and eosin and Alcian blue as well as immunohistochemistry staining for substance P, tyrosine hydroxylase, and neurofilament. Tendon architecture was assessed according to the modified BONAR tendon score. Immunohistochemistry staining was quantified using ImageJ density analysis. Portions of the tendon sections were cultured for 48 hours and the resultant secretome was assessed for Substance P secretion with ELISA analysis. Results Composite BONAR scores demonstrated pathologic tendons with no significant difference between the intra-articular biceps tendon and the portion of the tendon within the bicipital groove (p= 0.31). Immunohistochemistry (IHC) analysis demonstrated significantly (p0.05). Stained substance P and tyr hydroxylase was intimately associated with intra-tendinous vascularity. ELISA results demonstrated significantly increased secretion of substance P in the bicipital groove tendon compared to the intraarticular portion. Conclusion This study shows abnormal architecture, increased release and staining of painful neurotransmitters, and secretion of substance P from the proximal biceps tendon up to the distal aspect of the bicipital groove. Despite varied surgical techniques for tenodesis, this study supports the removal of the entirety of the long head of the biceps to the distal portion of the bicipital groove. [Figure: see text][Figure: see text]

In two different mouse liver cancer models, we recently showed that a switch from oxidative phosphorylation (Oxphos) to glycolysis (the Warburg effect) is invariably accompanied by a marked decline in fatty acid oxidation (FAO) and a reciprocal increase in the activity of pyruvate dehydrogenase (PDH), which links glycolysis to the TCA cycle. We now show that short-term implementation of either medium-chain (MC) or long-chain (LC) high fat diets (HFDs) nearly doubled the survival of mice with c-Myc oncoprotein-driven hepatocellular carcinoma (HCC). Mechanistically, HFDs forced tumors to become more reliant on fatty acids as an energy source, thus normalizing both FAO and PDH activities. More generally, both MC- and LC-HFDs partially or completely normalized the expression of 682 tumor-dysregulated transcripts, a substantial fraction of which are involved in cell cycle control, proliferation and metabolism. That these same transcripts were responsive to HFDs in livers strongly suggested that the changes were the cause of tumor inhibition rather than its consequence. In seven different human cancer cohorts, patients with tumors containing high ratios of FAO-related:glycolysis-related transcripts had prolonged survival relative to those with low ratios. Furthermore, in 13 human cancer types, the expression patterns of transcripts encoding enzymes participating in FAO and/or cholesterol biosynthesis also correlated with significantly prolonged survival. Collectively, our results support the idea that the survival benefits of HFDs are due to a reversal of the Warburg effect and other tumor-associated metabolic and cell cycle abnormalities. They also suggest that short-term dietary manipulation, either alone or in combination with more traditional chemotherapeutic regimens, might be employed as a relatively non-toxic and cost-effective means of enhancing survival in certain cancer types.

Objective Cisplatin is a chemotherapeutic drug known to induce hearing loss. Although corticosteroids may help to mitigate the ototoxic side effects of cisplatin, there are complications associated with their systemic and prolonged use. The goal of this study is to test the efficacy of extended-release fluticasone propionate intracochlear implant particles to protect against cisplatin-induced hearing loss. Methods We used guinea pigs (n = 9) injected with cisplatin (IP, 12 mg/kg weight). Fluticasone particles were delivered to the cochlear scala tympani through the round window membrane into the right ears of the guinea pigs (left ears being used as a control) two weeks prior to cisplatin administration, and hearing function was evaluated by ABR and DPOAE before implantation, immediately before cisplatin administration, and 2 weeks after the challenge with cisplatin. Data was statistically evaluated using paired t-test analysis. Results No significant differences were observed in ABR threshold between control and implanted ears on day 14 (23.9 ± 2.3 dB vs. 25.6 ± 1.3 dB, P = 0.524), whereas the significant cisplatin-induced hearing loss in control animals (23.9 ± 2.3 dB at day 14 vs. 40.7 ± 2.5 dB at day 28, P ≤ 0.0001) was prevented in implanted animals (25.6 ± 1.3 dB at day 14 vs. 25.0 ± 3.1 at day 28, P ≥ 0.85). A similar, though not statistically significant, trend was observed in DPOAE responses in untreated ears (7.9 ± 5.8 dB at day14 vs. −0.5 ± 5.3 dB at day 28, P = 0.654) as compared to treatment (11.1 ± 3.4 dB at day 14 vs. 13.6 ± 4.8 dB at day 28, P = 0.733). Conclusion These results suggest that fluticasone intracochlear implants are safe and able to provide effective otoprotection against cisplatin-induced hearing loss in the guinea pig model.

Rapid climate warming has widely been considered as the main driver of recent increases in Arctic tundra productivity. Field observations and remote sensing both show that tundra “greening” has been widespread, but heterogeneity in regional and landscape-scale trends suggest that additional controls are mediating the response of tundra vegetation to warming. In this study, we examined the relationship between changes in vegetation productivity in the western Canadian Arctic and biophysical variables by analyzing trends in the Enhanced Vegetation Index (EVI) obtained from nonparametric regression of annual Landsat surface reflectance composites. We used Random Forests classification and regression tree modelling to predict the trajectory and magnitude of greening from 1984 to 2016 and identify biophysical controls. More than two-thirds of our study area showed statistically significant increases in vegetation productivity, but observed changes were heterogeneous, occurring most rapidly within areas of the Southern Arctic that were: (1) dominated by dwarf and upright shrub cover types, (2) moderately sloping, and (3) located at lower elevation. These findings suggest that the response of tundra vegetation to warming is mediated by regional- and landscape-scale variation in microclimate, topography and soil moisture, and physiological differences among plant functional groups. Our work highlights the potential of the joint analysis of annual remotely sensed vegetation indices and broad-scale biophysical data to understand spatial variation in tundra vegetation change.

XLH is the most common heritable rickets. Affected children have high levels of circulating FGF23 that cause hypophosphatemia with consequent rickets and lower limb deformity. Burosumab, a fully human monoclonal antibody that binds FGF23, is FDA-approved for the treatment of XLH in children ≥6 months old and adults. Herein, we report final, 3-year safety and efficacy data from an open-label, phase 2 study of burosumab in children 1 to Eligibility required hypophosphatemia and radiographic evidence of rickets. The primary efficacy endpoint was change from baseline in fasting serum phosphorus (Pi). Secondary endpoints included Rickets Severity Score (RSS) and Radiographic Global Impression of Change (RGI-C). Patients received burosumab subcutaneous Q2W starting at 0.8 mg/kg for 160 weeks (64-week treatment + 96-week treatment extension periods). All 13 enrolled patients completed the 64-week treatment period; 1 left the study to transition to commercially available burosumab, and 12 completed all 160 weeks. Baseline mean (SD) age was 2.9 (1.1) years; 69% were boys; all had previously received oral phosphate salts and active vitamin D. Burosumab rapidly corrected fasting serum Pi with mean (SD) levels of 2.5 (0.3) mg/dL at Baseline, 3.7 (0.5) mg/dL at Week 1 (W1), 3.4 (0.5) mg/dL at W64, and 3.4 (0.5) mg/dL at W160 (normal range: 3.2–6.1 mg/dL). Lower RSS indicated improved rickets. Total RSS decreased from 2.9 (1.4) at Baseline to 1.2 (0.5) at W40 and to 0.9 (0.5) at W64 and was maintained through W160 [1.0 (0.6)]. Positive RGI-C scores indicate healing rickets relative to Baseline. Global RGI-C scores indicating substantial healing (≥+2) at W40 [+2.2 (0.3)] and W64 [+2.2 (0.4)] were maintained through W160 [+2.2 (0.4)]. Similarly, lower limb deformity RGI-C scores were +1.2 (0.6) at W40 and +1.5 (0.5) at W64, and sustained healing was evident at W160 [+2.0 (0.3)]. Wrist and knee RSSs and RGI-C scores similarly improved. The upper limit of normal for serum ALP ranged from 297 to 345 U/L depending on the child’s age and sex. Mean ALP was 549 (194) U/L at Baseline, normalized by W40 [335 (88) U/L], and was sustained through W160 [302 (71) U/L]. The burosumab safety profile over 160 weeks resembled previous pediatric studies; no new safety concerns emerged. All patients had ≥1 treatment-emergent adverse event (TEAE). All TEAEs were mild (Grade 1) or moderate (Grade 2) except for one patient with a grade 3 TEAE (food allergy) and one with a grade 3 TEAE (increased serum amylase, 92% salivary/8% pancreatic). One patient had a serious TEAE (dental abscess leading to hospitalization). These grade 3 and serious TEAEs were considered unrelated to study drug. Burosumab rapidly restored Pi homeostasis, improved rickets, and normalized serum ALP in children with XLH aged 1 to

Humans emit large quantities of greenhouse gases, driving climate change and putting all species at risk. Psychologists can assist in changing these harmful greenhouse gas-emitting behaviors. However, structural and psychological obstacles exist, hindering behavioral efforts. Examining the question of what limits proenvironmental behavior for individuals that have the capacity to act (i.e., those not facing important structural barriers), Gifford (2011) gathered a list of psychological barriers called the dragons of inaction. The dragons of inaction comprise 30 psychological barriers, organized under 7 categories. Studies measuring the dragons of inaction in the context of mitigative food choices and household energy use are reviewed here. These studies suggest that the dragons of inaction comprise four to six barrier factors. Building from these findings, a recent study is discussed, which proposes a new measurement scale for psychological barriers based on exploratory and confirmatory analyses. This psychometric scale, called the Dragons of Inaction Psychological Barrier (DIPB) instrument, is validated for use across six domains of environmental behavior. Practical applications of the DIPB are presented. Theoretical applications of the DIPB are also presented; specifically, its use in measuring the perception of barriers in a (proposed) theory of behavioral choice (TBC). The TBC improves upon the well-known theory of planned behavior by incorporating psychological barriers, structural barriers, desire, and the distinction between actual and reported behavior. The DIPB and the TBC provide a valuable contribution to climate change mitigation and adaptation efforts.