Effect of Ligands and Transducers on the Neurotensin Receptor 1 Conformational Ensemble.

Using a discrete, intracellular 19F nuclear magnetic resonance (NMR) probe on transmembrane helix 6 of the neurotensin receptor 1 (NTS1), we aim to understand how ligands and transducers modulate the receptor's structural ensemble in a solution. For apo NTS1, 19F NMR spectra reveal an ensemble of at least three conformational substates (one inactive and two active-like) in equilibrium that exchange on the millisecond to second timescale. Dynamic NMR experiments reveal that these substates follow a linear three-site exchange process that is both thermodynamically and kinetically remodeled by orthosteric ligands. As previously observed in other G protein-coupled receptors (GPCRs), the full agonist is insufficient to completely stabilize the active-like state. The inactive substate is abolished upon coupling to ß-arrestin-1 (ßArr1) or the C-terminal helix of Gαq, which comprises ≳60% of the GPCR/G protein interface surface area. Whereas ßArr1 exclusively selects for pre-existing active-like substates, the Gαq peptide induces a new substate. Both transducer molecules promote substantial line broadening of active-like states, suggesting contributions from additional microsecond to millisecond exchange processes. Together, our study suggests that (i) the NTS1 allosteric activation mechanism may be alternatively dominated by induced fit or conformational selection depending on the coupled transducer, and (ii) the available static structures do not represent the entire conformational ensemble observed in a solution.

Sulfonate/Nitro Bearing Methylmalonyl-Thioester Isosteres Applied to Methylmalonyl-CoA Decarboxylase Structure-Function Studies.

Malonyl-thioesters are reactive centers of malonyl-CoA and malonyl- S-acyl carrier protein, essential to fatty acid, polyketide and various specialized metabolite biosynthesis. Enzymes that create or use malonyl-thioesters spontaneously hydrolyze or decarboxylate reactants on the crystallographic time frame preventing determination of structure-function relationships. To address this problem, we have synthesized a panel of methylmalonyl-CoA analogs with the carboxylate represented by a sulfonate or nitro and the thioester retained or represented by an ester or amide. Structures of Escherichia coli methylmalonyl-CoA decarboxylase in complex with our analogs affords insight into substrate binding and the catalytic mechanism. Counterintuitively, the negatively charged sulfonate and nitronate functional groups of our analogs bind in an active site hydrophobic pocket. Upon decarboxylation the enolate intermediate is protonated by a histidine preventing CO2-enolate recombination, yielding propionyl-CoA. Activity assays support a histidine catalytic acid and reveal the enzyme displays significant hydrolysis activity. Our structures also provide insight into this hydrolysis activity. Our analogs inhibit decarboxylation/hydrolysis activity with low micromolar Ki values. This study sets precedents for using malonyl-CoA analogs with carboxyate isosteres to study the complicated structure-function relationships of acyl-CoA carboxylases, trans-carboxytransferases, malonyltransferases and ß-ketoacylsynthases.

Squamous cell carcinoma in the setting of hidradenitis suppurativa: a retrospective review of the literature.

BACKGROUND: Marjolin ulcer is an SCC arising from chronic inflammatory tissue. Such ulcers pose a high risk for metastasis; the 5-year survival rate of 40% to 69% suggests that improvement is possible with early diagnosis. OBJECTIVE: To raise awareness and update education in the medical community regarding this manifestation of SCC. METHODS: The authors performed a retrospective literature review specifically for SCC arising in the context of HS. A PubMed search yielded 81 articles reporting SCC in the context of HS from 1958 to November 2022, with a total of 123 cases. The authors' additional patient, who was treated in practice, is included as Case 1, bringing the total number of patients to 124. RESULTS: Seventy-three percent of patients were male. The mean age at the time of SCC diagnosis was 53 years ± 10.25 standard deviation. The mean interval between HS diagnosis and SCC development was 24.4 years ± 11.33. Thirty-six percent of patients had metastatic disease. Of the 102 cases that included details on life and death, 58 were noted to have died; of those 58, 60% died within the first year following diagnosis. CONCLUSION: Given the poor prognosis of Marjolin ulcer, with rapid progression after diagnosis, frequent visual examination and biopsies with a high index of suspicion for HS are recommended to identify SCC prior to metastatic transformation and subsequent unresectable disease. If surgical intervention is performed in the management of HS, tissue should always be sent for pathologic examination.

Modeling tissue-specific breakpoint proximity of structural variations from whole-genomes to identify cancer drivers.

Structural variations (SVs) in cancer cells often impact large genomic regions with functional consequences. However, identification of SVs under positive selection is a challenging task because little is known about the genomic features related to the background breakpoint distribution in different cancers. We report a method that uses a generalized additive model to investigate the breakpoint proximity curves from 2,382 whole-genomes of 32 cancer types. We find that a multivariate model, which includes linear and nonlinear partial contributions of various tissue-specific features and their interaction terms, can explain up to 57% of the observed deviance of breakpoint proximity. In particular, three-dimensional genomic features such as topologically associating domains (TADs), TAD-boundaries and their interaction with other features show significant contributions. The model is validated by identification of known cancer genes and revealed putative drivers in cancers different than those with previous evidence of positive selection.

Two unique cases of Bouveret syndrome with review of literature.

Bouveret syndrome is a rare form of gallstone ileus in which a proximally lodged gallstone in the duodenum causes a gastric outlet obstruction. It is a rare condition that can be challenging to manage. Although endoscopic management remains first line, a surgical approach can be needed. We present two cases of Bouveret syndrome. A 65-year-old man with oral squamous cell carcinoma treated with endoscopic management and a 63-year-old woman treated with surgery.

A method for selective 19 F-labeling absent of probe sequestration (SLAPS).

Fluorine (19 F) offers several distinct advantages for biomolecular nuclear magnetic resonance spectroscopy such as no background signal, 100% natural abundance, high sensitivity, and a large chemical shift range. Exogenous cysteine-reactive 19 F-probes have proven especially indispensable for characterizing large, challenging systems that are less amenable to other isotopic labeling strategies such as G protein-coupled receptors. As fluorine linewidths are inherently broad, limiting reactions with offsite cysteines is critical for spectral simplification and accurate deconvolution of component peaks-especially when analyzing systems with intermediate to slow timescale conformational exchange. Here, we uncovered noncovalent probe sequestration by detergent proteomicelles as a second source of offsite labeling when using the popular 19 F-probe BTFMA (2-bromo-N-(4-[trifluoromethyl]phenyl)acetamide). The chemical shift and relaxation rates of these unreacted 19 F-BTFMA molecules are insufficient to distinguish them from protein-conjugates, but they can be easily identified using mass spectrometry. We present a simple four-step protocol for Selective Labeling Absent of Probe Sequestration (SLAPS): physically disrupt cell membranes in the absence of detergent, incubate membranes with cysteine-reactive 19 F-BTFMA, remove excess unreacted 19 F-BTFMA molecules via ultracentrifugation, and finally solubilize in the detergent of choice. Our approach builds upon the in-membrane chemical modification method with the addition of one crucial step: removal of unreacted 19 F-probes by ultracentrifugation prior to detergent solubilization. SLAPS is broadly applicable to other lipophilic cysteine-reactive probes and membrane protein classes solubilized in detergent micelles or lipid mimetics.

fastMRI+, Clinical pathology annotations for knee and brain fully sampled magnetic resonance imaging data.

Improving speed and image quality of Magnetic Resonance Imaging (MRI) using deep learning reconstruction is an active area of research. The fastMRI dataset contains large volumes of raw MRI data, which has enabled significant advances in this field. While the impact of the fastMRI dataset is unquestioned, the dataset currently lacks clinical expert pathology annotations, critical to addressing clinically relevant reconstruction frameworks and exploring important questions regarding rendering of specific pathology using such novel approaches. This work introduces fastMRI+, which consists of 16154 subspecialist expert bounding box annotations and 13 study-level labels for 22 different pathology categories on the fastMRI knee dataset, and 7570 subspecialist expert bounding box annotations and 643 study-level labels for 30 different pathology categories for the fastMRI brain dataset. The fastMRI+ dataset is open access and aims to support further research and advancement of medical imaging in MRI reconstruction and beyond.

Acupuncture for acute musculoskeletal pain management in the emergency department and continuity clinic: a protocol for an adaptive pragmatic randomised controlled trial.

INTRODUCTION: Chronic musculoskeletal pain causes a significant burden on health and quality of life and may result from inadequate treatment of acute musculoskeletal pain. The emergency department (ED) represents a novel setting in which to test non-pharmacological interventions early in the pain trajectory to prevent the transition from acute to chronic pain. Acupuncture is increasingly recognised as a safe, affordable and effective treatment for pain and anxiety in the clinic setting, but it has yet to be established as a primary treatment option in the ED. METHODS AND ANALYSIS: This pragmatic clinical trial uses a two-stage adaptive randomised design to determine the feasibility, acceptability and effectiveness of acupuncture initiated in the ED and continued in outpatient clinic for treating acute musculoskeletal pain. The objective of the first (treatment selection) stage is to determine the more effective style of ED-based acupuncture, auricular acupuncture or peripheral acupuncture, as compared with no acupuncture. All arms will receive usual care at the discretion of the ED provider blinded to treatment arm. The objective of the second (effectiveness confirmation) stage is to confirm the impact of the selected acupuncture arm on pain reduction. An interim analysis is planned at the end of stage 1 based on probability of being the best treatment, after which adaptations will be considered including dropping the less effective arm, sample size re-estimation and unequal treatment allocation ratio (eg, 1:2) for stage 2. Acupuncture treatments will be delivered by licensed acupuncturists in the ED and twice weekly for 1 month afterward in an outpatient clinic. ETHICS AND DISSEMINATION: This study has been reviewed and approved by the Duke University Health System Institutional Review Board. Informed consent will be obtained from all participants. Results will be disseminated through peer-review publications and public and conference presentations. TRIAL REGISTRATION NUMBER: NCT04290741.

Defense by exploitation in Negev gerbils.

In this study, we addressed how frequently a non-traplining animal should visit food patches. More specifically, we investigate if non-traplining animals engage in a behavior called "defense by exploitation", which is characterized by an increase in visitation rates with increased intra-specific competition. We ran four tests with two gerbil species in the Negev Desert. Firstly, we measured patch use of Gerbillus pyramidum and Gerbillus andersoni allenbyi. We assumed that activity and competition would decrease through the night and that patch use would decrease with number of visits. Secondly, we measured how the number of visits to resource patches increased with the addition of individuals. Thirdly, we repeated this experiment, but instead removed individuals. Lastly, we conducted a simulation to compare these results against theoretical expectations. In the first test, we found support for defense by exploitation in G. pyramidum. The second and third test found no support. The fourth test found support for this increase visitation, but only if costs of locomotion are relatively small.

Divergent behavior amid convergent evolution: A case of four desert rodents learning to respond to known and novel vipers.

Desert communities world-wide are used as natural laboratories for the study of convergent evolution, yet inferences drawn from such studies are necessarily indirect. Here, we brought desert organisms together (rodents and vipers) from two deserts (Mojave and Negev). Both predators and prey in the Mojave have adaptations that give them competitive advantage compared to their middle-eastern counterparts. Heteromyid rodents of the Mojave, kangaroo rats and pocket mice, have fur-lined cheek pouches that allow them to carry larger loads of seeds under predation risk compared to gerbilline rodents of the Negev Deserts. Sidewinder rattlesnakes have heat-sensing pits, allowing them to hunt better on moonless nights when their Negev sidewinding counterpart, the Saharan horned vipers, are visually impaired. In behavioral-assays, we used giving-up density (GUD) to gauge how each species of rodent perceived risk posed by known and novel snakes. We repeated this for the same set of rodents at first encounter and again two months later following intensive "natural" exposure to both snake species. Pre-exposure, all rodents identified their evolutionarily familiar snake as a greater risk than the novel one. However, post-exposure all identified the heat-sensing sidewinder rattlesnake as a greater risk. The heteromyids were more likely to avoid encounters with, and discern the behavioral difference among, snakes than their gerbilline counterparts.

Improving retrieval rates for retrievable inferior vena cava filters.

The introduction of retrievable inferior vena cava (IVC) filters was an important step in the evolution of deep vein thrombosis/pulmonary embolism management. Their removability makes them preferred to permanent filters in many cases. IVC filter retrieval often occurs at a suboptimal rate, leading to complications associated with long-term placement. Improving retrievability includes solutions for patients being lost to follow-up, filter malpositioning, need arising for permanent IVC filtration, filtration requiring longer than the filter's window of retrievability, and filter compromise by the presence of a large trapped clot. This review explores these strategies for retrieval in detail in hopes of improving IVC filter retrieval rates.