This research employs pepsin-containing membranes to digest proteins online after a capillary electrophoresis (CE) separation and prior to tandem mass spectrometry. Proteolysis after the separation ...allows the peptides from a given protein to enter the mass spectrometer in a single plug. Thus, migration time can serve as an additional criterion for confirming the identification of a peptide. The membrane resides in a sheath-flow electrospray ionization (ESI) source to enable digestion immediately before spray into the mass spectrometer, thus limiting separation of the digested peptides. Using the same membrane, digestion occurred reproducibly during 20 consecutive CE analyses performed over a 10 h period. Additionally, after separating a mixture of six unreduced proteins with CE, online digestion facilitated protein identification with at least 2 identifiable peptides for all the proteins. Sequence coverages were >75% for myoglobin and carbonic anhydrase II but much lower for proteins containing disulfide bonds. Development of methods for efficient separation of reduced proteins or identification of cross-linked peptides should enhance sequence coverages for proteins with disulfide bonds. Migration times for the peptides identified from a specific protein differed by <∼30 s, which allows for rejection of some spurious peptide identifications.
This research employs pepsin-containing membranes to digest proteins online after a capillary electrophoresis separation and prior to tandem mass spectrometry, allowing the peptides from a protein to enter the mass spectrometer in a single plug.
Interleukin (IL)-2 is a pleiotropic cytokine that is necessary to prevent chronic inflammation in the gastrointestinal tract
. The protective effects of IL-2 involve the generation, maintenance and ...function of regulatory T (T
) cells
, and the use of low doses of IL-2 has emerged as a potential therapeutic strategy for patients with inflammatory bowel disease
. However, the cellular and molecular pathways that control the production of IL-2 in the context of intestinal health are undefined. Here we show, in a mouse model, that IL-2 is acutely required to maintain T
cells and immunological homeostasis throughout the gastrointestinal tract. Notably, lineage-specific deletion of IL-2 in T cells did not reduce T
cells in the small intestine. Unbiased analyses revealed that, in the small intestine, group-3 innate lymphoid cells (ILC3s) are the dominant cellular source of IL-2, which is induced selectively by IL-1β. Macrophages in the small intestine produce IL-1β, and activation of this pathway involves MYD88- and NOD2-dependent sensing of the microbiota. Our loss-of-function studies show that ILC3-derived IL-2 is essential for maintaining T
cells, immunological homeostasis and oral tolerance to dietary antigens in the small intestine. Furthermore, production of IL-2 by ILC3s was significantly reduced in the small intestine of patients with Crohn's disease, and this correlated with lower frequencies of T
cells. Our results reveal a previously unappreciated pathway in which a microbiota- and IL-1β-dependent axis promotes the production of IL-2 by ILC3s to orchestrate immune regulation in the intestine.
Objective
A novel peer facilitation model was used to deliver a two session, dissonance‐based, inclusive body image intervention that critically examines how internalized size‐based oppression ...intersects with race, class, gender, sexuality, and ability.
Method
The EVERYbody Project was open to all college students and delivered by “expert” peer facilitators with body image and diversity experience and advanced facilitation skills. Recruitment was halted due to COVID‐19; 90 students in the Northwest US (M age = 19.83 years, SD = 2.38; 80% female‐identified, 13% male‐identified, 7% gender expansive) were randomized to receive the EVERYbody Project or a video‐based, low‐dissonance comparison intervention.
Results
Around half of students (56%) held one or more specific socially marginalized identity (26% with a racial or ethnic identity other than white and non‐Hispanic, 39% with a sexual identity other than straight, 7% with a gender identity other than cisgender). The EVERYbody Project produced greater reductions in three outcomes associated with poor body image (internalized appearance norms, body dissatisfaction, and negative affect) compared to the video intervention through 3‐month follow‐up (ps < .003) with medium between group effects. Both conditions experienced small reductions in eating disorder psychopathology over time.
Discussion
Expert peer facilitation may be a viable delivery model for inclusive, diversity‐focused dissonance‐based body image programs.
Public Significance
This study explored a novel facilitation approach for a diversity‐focused body image program for college students. A brief (4 h) dissonance‐based program was open to all college students and delivered by “expert” diverse peer leaders who were screened for facilitation readiness. Body image and related outcomes were improved through 3‐month follow‐up relative to a comparison condition, suggesting that expert peer facilitation may be a viable option for delivering universal, inclusive body image programming.
Light sheet microscopy of the gerbil cochlea Hutson, Kendall A.; Pulver, Stephen H.; Ariel, Pablo ...
Journal of comparative neurology (1911),
March 2021, Letnik:
529, Številka:
4
Journal Article
Recenzirano
Odprti dostop
Light sheet fluorescence microscopy (LSFM) provides a rapid and complete three‐dimensional image of the cochlea. The method retains anatomical relationships—on a micrometer scale—between internal ...structures such as hair cells, basilar membrane (BM), and modiolus with external surface structures such as the round and oval windows. Immunolabeled hair cells were used to visualize the spiraling BM in the intact cochlea without time intensive dissections or additional histological processing; yet material prepared for LSFM could be rehydrated, the BM dissected out and reimaged at higher resolution with the confocal microscope. In immersion‐fixed material, details of the cochlear vasculature were seen throughout the cochlea. Hair cell counts (both inner and outer) as well as frequency maps of the BM were comparable to those obtained by other methods, but with the added dimension of depth. The material provided measures of angular, linear, and vector distance between characteristic frequency regions along the BM. Thus, LSFM provides a unique ability to rapidly image the entire cochlea in a manner applicable to model and interpret physiological results. Furthermore, the three‐dimensional organization of the cochlea can be studied at the organ and cellular level with LSFM, and this same material can be taken to the confocal microscope for detailed analysis at the subcellular level.
Light sheet microcopy is used to map frequency along the basilar membrane within intact cochleae, retaining spatial orientation and relationships with internal and external structural landmarks. Three‐dimensional measures of how frequency domains relate to one another gives insight into how the geometric organization of the cochlea may influence its physiology.
Abstract
Objectives
We evaluated how the changes in Gleason grading affected the long-term outcomes of a large prostatectomy cohort.
Methods
We obtained long-term follow-up (16.7 years) in 581 ...patients having undergone radical retropubic prostatectomy between 1985 and 1995. We excluded those with seminal vesicle and/or lymphatic involvement. We regraded the specimens according to contemporary guidelines and compared how this affected outcomes compared with their original (pre-1995) Gleason scoring. In total, 499 patients were evaluable.
Results
A Gleason score of 6 or less declined from 73% to 29%, and the number increased from 25% to 63% for a Gleason score of 7 and from 5% to 8% for a Gleason score of 8 to 9. As a result, for a Gleason score less than 7, biochemical failure decreased from 28% to 23%, metastatic disease 5% to 2%, and prostate cancer death from 5% to 3%. The same results were 50% to 37%, 11% to 7%, and 10% to 6% for a Gleason score of 7 and 86% to 71%, 43% to 32%, and 29% to 26% for a Gleason score more than 7, respectively. With the most recent grade grouping, for groups 1 to 5, biochemical failure occurred in 23%, 32%, 45%, 69%, and 78%, respectively. Metastatic disease occurred in 2%, 4%, 12%, 24%, and 56%, respectively. Prostate cancer–related death occurred in 2%, 4%, 9%, 21%, and 44%, respectively.
Conclusions
The revised Gleason scores improved the outcomes in all risk groups. Based on Gleason score, patients with prostate cancer will appear to have better outcomes than they did before 2005, making any comparison tenable. The current grading system shows a consistent increased risk in biochemical failure, metastatic disease, and prostate cancer–related death with each successive grade.
Organic electronic materials have the potential to impact almost every aspect of modern life including how we access information, light our homes, and power personal electronics. Nevertheless, weak ...intermolecular interactions and disorder at junctions of different organic materials limit the performance and stability of organic interfaces and hence the applicability of organic semiconductors to electronic devices. Here, we demonstrate control of donor–acceptor heterojunctions through microphase-separated conjugated block copolymers. When utilized as the active layer of photovoltaic cells, block copolymer-based devices demonstrate efficient photoconversion well beyond devices composed of homopolymer blends. The 3% block copolymer device efficiencies are achieved without the use of a fullerene acceptor. X-ray scattering results reveal that the remarkable performance of block copolymer solar cells is due to self-assembly into mesoscale lamellar morphologies with primarily face-on crystallite orientations. Conjugated block copolymers thus provide a pathway to enhance performance in excitonic solar cells through control of donor–acceptor interfaces.
Multidrug-resistant (MDR) tuberculosis can be acquired through de-novo mutation during tuberculosis treatment or through transmission from other individuals with active MDR tuberculosis. ...Understanding the balance between these two mechanisms is essential when allocating resources for MDR tuberculosis. We aimed to create a dynamic transmission model of an MDR tuberculosis epidemic to estimate the contributions of treatment-related acquisition and person-to-person transmission of resistance to incident MDR tuberculosis cases.
In this modelling analysis, we constructed a dynamic transmission model of an MDR tuberculosis epidemic, allowing for both treatment-related acquisition and person-to-person transmission of resistance. We used national tuberculosis notification data to inform Bayesian estimates of the proportion of each country's 2013 MDR tuberculosis incidence that resulted from MDR transmission rather than treatment-related MDR acquisition.
Global estimates of 3·5% MDR tuberculosis prevalence among new tuberculosis notifications and 20·5% among re-treatment notifications translate into an estimate that resistance transmission rather than acquisition accounts for a median 95·9% (95% uncertainty range UR 68·0-99·6) of all incident MDR tuberculosis, and 61·3% (16·5-95·2) of incident MDR tuberculosis in previously treated individuals. The estimated proportion of MDR tuberculosis resulting from transmission varied substantially with different countries' notification data-ranging from 48% (95% UR 30-75) in Bangladesh to 99% (91-100) in Uzbekistan. Estimates were most sensitive to estimates of the transmissibility of MDR strains, the probability of acquiring MDR during tuberculosis treatment, and the responsiveness of MDR tuberculosis to first-line treatment.
Notifications of MDR prevalence from most high-burden settings are consistent with most incident MDR tuberculosis resulting from transmission rather than new treatment-related acquisition of resistance. Merely improving the treatment of drug-susceptible tuberculosis is unlikely to greatly reduce future MDR tuberculosis incidence. Improved diagnosis and treatment of MDR tuberculosis-including new tests and drug regimens-should be highly prioritised.
National Institutes of Health and the Bill & Melinda Gates Foundation.
The field of second language studies is using increasingly sophisticated methodological approaches to address a growing number of urgent, real-world problems. These methodological developments bring ...both new challenges and opportunities. This article briefly reviews recent ontological and methodological debates in the field, then builds on these insights to consider some of the current dilemmas faced by researchers of second language teaching and learning, including concerns regarding fragmentation, generalizability, and replication. Through a review of recent research, we argue that one means of addressing these ongoing questions is to continue to focus collectively and collaboratively on solving realworld problems of language learning, while also layering our perspectives. By layering, we mean considering the central philosophical challenges, often those that are basic values in our methodological approaches, such as objectivity and bias, from varied epistemological stances. We argue that recognizing these differences and using a layered approach will enhance and improve our attempts to address the pressing problems in our field.
Capacity building may play an important role in improving classroom teachers' and early childhood educators' (ECE) capacity to implement physical activity and FMS interventions. Capacity building is ...the development of knowledge, skills, and structures to improve the capability of individuals and organisations to achieve effective health promotion. This review aimed to determine the efficacy of capacity building interventions on teachers' and ECEs' perceived capabilities, knowledge, and attitudes relating to physical activity and fundamental movement skills.
An exhaustive literature search of six electronic databases was conducted. Controlled, single-group pre-post studies were included if they measured the effect of a capacity building intervention on in-service or pre-service classroom teachers' (primary or secondary) or ECEs' physical activity or fundamental skills related perceived capabilities, knowledge, or attitudes. The effects of interventions were synthesised using random effects meta-analysis. Subgroup analysis and meta-regression was conducted to determine if the effects differed based on study design, type of teacher (ECE vs. primary school), or teacher level (pre-service vs. in-service).
A total of 22 studies reporting on 25 unique samples were included in the meta-analyses. Only studies reporting on ECEs and primary school teachers were identified. Interventions most commonly included training/professional development, resources and toolkits, communities of practice, mentorships, and ongoing support. Results showed that capacity building interventions significantly improved teachers' and ECEs' perceived capabilities (g = 0.614, 95% CI = 0.442, 0.786), knowledge (g = 0.792 95% CI = 0.459, 1.125), and attitudes (g = 0.376 95% CI = 0.181, 0.571). The effects did not differ significantly as a function of any of the moderators examined.
Findings from this review provide strong support that capacity building interventions are efficacious at improving teachers' and ECEs' perceived capabilities, knowledge, and attitudes related to promoting physical activity and teaching fundamental movement skills. Pre-service teachers and ECEs should be provided training in physical activity and fundamental movement skills as part of their degrees, and continual professional development and capacity building should be offered to in-service teachers and ECEs to promote physical activity and fundamental movement skills in children.
Signal peptidase I (SPase I) is critical for the release of translocated preproteins from the membrane as they are transported from a cytoplasmic site of synthesis to extracytoplasmic locations. ...These proteins are synthesized with an amino‐terminal extension, the signal sequence, which directs the preprotein to the Sec‐ or Tat‐translocation pathway. Recent evidence indicates that the SPase I cleaves preproteins as they emerge from either pathway, though the steps involved are unclear. Now that the structure of many translocation pathway components has been elucidated, it is critical to determine how these components work in concert to support protein translocation and cleavage. Molecular modeling and NMR studies have provided insight on how the preprotein docks on SPase I in preparation for cleavage. This is a key area for future work since SPase I enzymes in a variety of species have now been identified and the inhibition of these enzymes by antibiotics is being pursued. The eubacterial SPase I is essential for cell viability and belongs to a unique group of serine endoproteases which utilize a Ser‐Lys catalytic dyad instead of the prototypical Ser‐His‐Asp triad used by eukaryotes. As such, SPase I is a desirable antimicrobial target. Advances in our understanding of how the preprotein interfaces with SPase I during the final stages of translocation will facilitate future development of inhibitors that display a high efficacy against SPase I function.
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