Endocytosed HIV-1 Envelope Glycoprotein Traffics to Rab14+ Late Endosomes and Lysosomes to Regulate Surface Levels in T-Cell Lines.

Production of infectious HIV-1 particles requires incorporation of the viral envelope glycoprotein (Env) at the plasma membrane (PM) of infected CD4+ T cells. Env trafficking to the PM exposes viral epitopes that can be exploited by the host immune system; however, HIV-1 can evade this response by endocytosis of excess Env from the PM. The fate of Env after internalization remains unclear, with evidence suggesting several different vesicular trafficking steps may be involved, including recycling pathways. To date, there have been very few studies documenting the trafficking pathways of native Env in infected T cells. Furthermore, it remains unclear whether there are T-cell-specific endosomal pathways regulating the fate of endocytic Env. Here, we use a pulse-labeling approach with a monovalent anti-Env Fab probe to characterize the trafficking of internalized Env within infected CD4+ T-cell lines, together with CRISPR/Cas9-mediated endogenous protein tagging, to assess the role of host cell Rab GTPases in Env trafficking. We show that endocytosed Env traffics to Rab14+ compartments that possess hallmarks of late endosomes and lysosomes. We also demonstrate that Env can recycle back to the PM, although we find that recycling does not occur at high rates when compared to the model recycling protein transferrin. These results help to resolve open questions about the fate and relevance of endocytosed Env in HIV-infected cells and suggest a novel role for Rab14 in a cell-type-specific late-endosomal/lysosomal trafficking pathway in T cells. IMPORTANCE HIV-1 envelope glycoprotein (Env) evades immune neutralization through many mechanisms. One immune evasion strategy may result from the internalization of excess surface-exposed Env to prevent antibody-dependent cellular cytotoxicity or neutralization. Characterization of the fate of endocytosed Env is critical to understand which vesicular pathways could be targeted to promote display of Env epitopes to the immune system. In this study, we characterize the endocytic fate of native Env, expressed from infected human T-cell lines. We demonstrate that Env is rapidly trafficked to a late-endosome/lysosome-like compartment and can be recycled to the cell surface for incorporation into virus assembly sites. This study implicates a novel intracellular compartment, marked by host-cell Rab14 GTPases, for the sequestration of Env. Therapeutic approaches aimed at mobilizing this intracellular pool of Env could lead to stronger immune control of HIV-1 infection via antibody-dependent cell-mediated cytotoxicity.

Critical Comparison of MaxCal and Other Stochastic Modeling Approaches in Analysis of Gene Networks.

Learning the underlying details of a gene network with feedback is critical in designing new synthetic circuits. Yet, quantitative characterization of these circuits remains limited. This is due to the fact that experiments can only measure partial information from which the details of the circuit must be inferred. One potentially useful avenue is to harness hidden information from single-cell stochastic gene expression time trajectories measured for long periods of time-recorded at frequent intervals-over multiple cells. This raises the feasibility vs. accuracy dilemma while deciding between different models of mining these stochastic trajectories. We demonstrate that inference based on the Maximum Caliber (MaxCal) principle is the method of choice by critically evaluating its computational efficiency and accuracy against two other typical modeling approaches: (i) a detailed model (DM) with explicit consideration of multiple molecules including protein-promoter interaction, and (ii) a coarse-grain model (CGM) using Hill type functions to model feedback. MaxCal provides a reasonably accurate model while being significantly more computationally efficient than DM and CGM. Furthermore, MaxCal requires minimal assumptions since it is a top-down approach and allows systematic model improvement by including constraints of higher order, in contrast to traditional bottom-up approaches that require more parameters or ad hoc assumptions. Thus, based on efficiency, accuracy, and ability to build minimal models, we propose MaxCal as a superior alternative to traditional approaches (DM, CGM) when inferring underlying details of gene circuits with feedback from limited data.

Molecular Structure of 1-Isocyano-1-silacyclopent-3-ene: A Combined Microwave Spectral and Theoretical Study.

The microwave spectrum of 1-isocyano-1-silacyclopent-3-ene has been obtained from broad-band chirped-pulse Fourier transform microwave spectroscopy. The rotational constants (RCs) for the standard abundant isotopic species are A = 3328.4182(23), B = 1017.69404(53), and C = 1012.33297(58) MHz. The symmetric quartic centrifugal distortion constants, using the Ir representation in CS symmetry for ΔJ,ΔJK, ΔK, and δJ, have been evaluated; similarly, the 14N nuclear quadrupole coupling has been determined. Several singly substituted isotopologues observed in natural abundance enabled most of the heavy atom substructure to be determined. The five-membered ring is close to planar, but the orientation of the isocyanate unit, derived from the N13CO spectrum, unexpectedly lies above the ring center in a cis C2,5-Si-N═C conformation. Our initial equilibrium structural searches led to a trans orientation of the C2,5-Si-N═C unit, i.e., bending away from the ring. When the cis conformation was applied, the final equilibrium structure, assuming CS symmetry, gave RC values of 3221.3 ( A), 1037.0 ( B), and 1031.3 ( C) MHz, very close to the MW values. This enabled the full-equilibrium structure to be determined with confidence. The principal bond lengths were 1.7157 (Si-N), 1.8696 (Si-C), 1.1998 (N═C), and 1.1737 (C═O) Å, with angles 163.3 (Si-N═C), 178.1 (N═C═O), 96.5 (C-Si-C), and 118.7° (C-C═C), respectively. The extensive widening of the SiNC angle is particularly notable; the SiNCO unit has a trans dihedral angle. The cis orientation implies a (weak) attractive force between the ring and isocyanate groups by a through-space interaction. An atoms in molecule study, where the local minima of electron density are determined, fails to disclose the exact nature of the interaction; however, a highly polarized skeleton was obtained. A systematic theoretical study of the Si-N═C angle potential energy surface (PES) relative to the ring gave a very shallow double minimum with the barrier being less than 1 cm-1; a polynomial fit to the surface shows major contributions of both harmonic and quartic components. A similar study of the XSiN angle, where X is at the ring center, also gave a PES with considerable quartic character.

Identifying groups at-risk to extreme heat: Intersections of age, race/ethnicity, and socioeconomic status.

Anthropogenic climate change has resulted in a significant rise in extreme heat events, exerting considerable but unequal impacts on morbidity and mortality. Numerous studies have identified inequities in heat exposure across different groups, but social identities have often been viewed in isolation from each other. Children (5 and under) and older adults (65 and older) also face elevated risks of heat-related health impacts. We employ an intersectional cross-classificatory approach to analyze the distribution of heat exposure between sociodemographic categories split into age groups in the contiguous US. We utilize high-resolution daily air temperature data to establish three census tract-level heat metrics (i.e., average summer temperature, heat waves, and heat island days). We pair those metrics with American Community Survey estimates on racial/ethnic, socioeconomic, and disability status by age to calculate population weighted mean exposures and absolute disparity metrics. Our findings indicate few substantive differences between age groups overall, but more substantial differences between sociodemographic categories within age groups, with children and older adults from socially marginalized backgrounds facing greater exposure than adults from similar backgrounds. When looking at sociodemographic differences by age, people of color of any age and older adults without health insurance emerge as the most exposed groups. This study identifies groups who are most exposed to extreme heat. Policy and program interventions aimed at reducing the impacts of heat should take these disparities in exposure into account to achieve health equity objectives.

Impact of Early Admission Surveys on Physicians' Communication Skills on Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS) Scores: A Quality Improvement Initiative.

Introduction Effective communication between doctors and patients is one of the 19 core questions on the Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS) survey. Positive patient experiences, as reflected in HCAHPS scores, are associated with higher quality of care, reduced mortality, and fewer readmissions. These scores also influence a hospital's financial bonuses or penalties from the Centers for Medicare & Medicaid Services. Methods This quality improvement (QI) initiative evaluated how early surveys of patients' feedback on doctors' communication skills during admission impacted discharge communication scores. The approach involved promptly sharing patients' feedback with physicians and focusing on improving communication skills for those with less-than-perfect scores. Results This QI initiative involved 41 patients. By surveying patients early during admission and addressing communication issues for those with less-than-perfect scores, the percentage of patients achieving a perfect score on all survey questions increased from 18 (43.9%) at admission to 34 (82.9%) at discharge. The largest improvement was seen in perfect scores for doctors' explanations of patients' conditions, which rose from 59% at admission to 90% at discharge. Conversely, the increase in perfect scores for doctors' respectful treatment was smaller, rising from 88% at admission to 98% at discharge. Conclusion Surveying patients about their doctors' communication skills early in admission to identify those with less-than-perfect scores and improving communication for this group resulted in a 39% increase in satisfaction with doctors' communication.

Bupropion Mediated Effects on Depression, Attention Deficit Hyperactivity Disorder, and Smoking Cessation.

Bupropion had been in use since the late 1980s as an unconventional treatment for depression. Unlike other antidepressants, bupropion has no serotonergic activity and inhibits the reuptake of norepinephrine and dopamine. The drug has been used to treat depression, Attention Deficit Hyperactivity Disorder (ADHD), and smoking cessation. This investigation reviews the pharmacokinetic and pharmacodynamic effects of bupropion and its mechanisms of action and interactions with other drugs. We evaluated the efficacy of major on and off-label uses of bupropion, focusing on the indications, benefits, and adverse effects. Our review demonstrates that bupropion is superior to placebo and non-inferior to SSRIs such as escitalopram in treating major depressive disorder. More research is needed to determine positive patient-centered outcomes such as increases in quality of life. In the case of ADHD, the evidence for efficacy is mixed with poorly conducted randomized clinical trials, small sample sizes, and a lack of long-term assessments. The same is true in the case of bipolar disorder in which there is still limited and controversial data available on bupropion's safety and efficacy. In the case of smoking cessation, bupropion is found to be an effective anti-smoking drug with synergistic benefits when used as a combination therapy. We conclude that bupropion has the potential to provide benefit for a subset of patients who do not tolerate other typical antidepressants or anti-smoking therapies or for those whose treatment goals align with bupropion's unique side effect profile, such as smokers who wish to quit and lose weight. Additional research is needed to determine the drug's full clinical potential, particularly in the areas of adolescent depression and combination therapy with varenicline or dextromethorphan. Clinicians should use this review to understand the varied uses of the drug and identify the situations and patient populations in which bupropion can lend its greatest benefit.

Successful mechanical thrombectomy for terminal ICA bullet embolism: A case report.

Bullet embolism after high velocity penetrating trauma is a rare event that can have devastating and wide-ranging effects distant from the original site of injury. A 29-year-old presented with multiple gunshot wounds to the chest, back, abdomen, and lower extremities but no penetrating head injury. After proper resuscitation, the patient was noted to have left-sided hemiparesis and computed tomography angiography of the head showed a bullet fragment that had traveled to the right M1 segment of the middle cerebral artery resulting in occlusion of the vessel. Mechanical thrombectomy was performed in an attempt to remove the bullet fragment but this was unsuccessful as the fragment was firmly lodged in the blood vessel. Aspiration of clot distal to the fragment was then performed in hopes of preventing a large volume ischemic event which was angiographically successful resulting in TICI 2c revascularization. This case demonstrates that thrombectomy can be safely and successfully performed distal to a lodged foreign body.

Reduced length of nodes of Ranvier and altered proteoglycan immunoreactivity in prefrontal white matter in major depressive disorder and chronically stressed rats.

Major depressive disorder (MDD) and chronic unpredictable stress (CUS) in animals feature comparable cellular and molecular disturbances that involve neurons and glial cells in gray and white matter (WM) in prefrontal brain areas. These same areas demonstrate disturbed connectivity with other brain regions in MDD and stress-related disorders. Functional connectivity ultimately depends on signal propagation along WM myelinated axons, and thus on the integrity of nodes of Ranvier (NRs) and their environment. Various glia-derived proteoglycans interact with NR axonal proteins to sustain NR function. It is unclear whether NR length and the content of associated proteoglycans is altered in prefrontal cortex (PFC) WM of human subjects with MDD and in experimentally stressed animals. The length of WM NRs in histological sections from the PFC of 10 controls and 10 MDD subjects, and from the PFC of control and CUS rats was measured. In addition, in WM of the same brain region, five proteoglycans, tenascin-R and NR protein neurofascin were immunostained or their levels measured with western blots. Analysis of covariance and t-tests were used for group comparisons. There was dramatic reduction of NR length in PFC WM in both MDD and CUS rats. Proteoglycan BRAL1 immunostaining was reduced at NRs and in overall WM of MDD subjects, as was versican in overall WM. Phosphacan immunostaining and levels were increased in both in MDD and CUS. Neurofascin immunostaining at NRs and in overall WM was significantly increased in MDD. Reduced length of NRs and increased phosphacan and neurocan in MDD and stressed animals suggest that morphological and proteoglycan changes at NRs in depression may be related to stress exposure and contribute to connectivity alterations. However, differences between MDD and CUS for some NR related markers may point to other mechanisms affecting the structure and function of NRs in MDD.

Preventing Medical Device-Related Pressure Injuries Due to Noninvasive Ventilation Masks and Nasal Cannulas.

BACKGROUND: Medical device-related pressure injuries attributed to oxygen delivery devices are common yet preventable. Pressure injuries increase hospital costs, patients' length of stay, and mortality rates. OBJECTIVE: To decrease medical device-related pressure injuries by transitioning patients from an over-the-nose noninvasive ventilation mask to a single-headset, interchangeable under-the-nose and over-the-nose noninvasive ventilation mask; replacing polyvinyl chloride nasal cannulas with a softer nasal cannula; and providing staff education on preventing pressure injuries related to oxygen delivery devices. METHODS: The project was implemented on 4 adult inpatient units with a total of 75 inpatient beds at an academic medical center. Interventions included implementing the trial noninvasive ventilation masks and nasal cannulas, alternating masks every 4 hours, relocating protective dressings, promoting the use of protective dressings for over-the-nose oral-nasal masks, and educating health care staff. RESULTS: In the 2 months before implementation (September and October 2020), 1 medical device-related pressure injury was caused by a noninvasive ventilation mask and 4 injuries were caused by nasal cannulas. During the 2 months of trial implementation (November and December 2020), no pressure injuries developed in patients using the trial devices. DISCUSSION: The interventions implemented had clinically relevant results. A larger sample size would be necessary to determine statistical significance. Postintervention data indicated a need for further education on evidence-based practice guidelines on mask alternation and use of preventive dressings to bolster compliance. CONCLUSION: Following institutional approval, all noninvasive ventilation masks and nasal cannulas were transitioned to the trial devices at the study institution.

Patient-Specific Characteristics Associated with Favorable Response to Vagus Nerve Stimulation.

OBJECTIVE: The expansion in treatments for medically refractory epilepsy heightens the importance of identifying patients who are likely to benefit from vagus nerve stimulation (VNS). Here, we identify predictors with a positive VNS response. METHODS: We present a retrospective analysis of 158 patients with medically refractory epilepsy. Patients were categorized as VNS responders or nonresponders. Baseline characteristics and time to VNS response were recorded. Univariate and multivariate Cox regression were used to identify predictors of response. Recursive partitioning analysis was used to identify likely VNS responders. RESULTS: Eighty-nine (56.3%) patients achieved ≥50% seizure frequency reduction. Left-hand dominance (hazard ratio [HR] 1.703, P = 0.038), age at epilepsy onset ≥15 years (HR 2.029, P = 0.005), duration of epilepsy ≥8 years (HR 1.968, P = 0.007) and age at implantation ≥35 years (HR 1.809, P = 0.020), and baseline seizure frequency <5/month (HR 1.569, P = 0.044) were significant univariate predictors of VNS response. Following multivariate Cox regression, left-hand dominance, age at epilepsy onset ≥15 years, and duration of epilepsy ≥8 years remained significant. With recursive partitioning analysis, patients with either age at epilepsy onset ≥15 years, left-hand dominance, or baseline seizure frequency <5/month were stratified into Group A and had a 73.9% responder rate; the remaining patients stratified into Group B had a 43.8% responder rate. CONCLUSIONS: Patients with age at epilepsy onset ≥15 years, left-hand dominance, or baseline seizure frequency <5/month are ideal candidates for VNS.

Sequence-specific destabilization of azurin by tetramethylguanidinium-dipeptide ionic liquids.

The thermal unfolding of the copper redox protein azurin was studied in the presence of four different dipeptide-based ionic liquids (ILs) utilizing tetramethylguanidinium as the cation. The four dipeptides have different sequences including the amino acids Ser and Asp: TMG-AspAsp, TMG-SerSer, TMG-SerAsp, and TMG-AspSer. Thermal unfolding curves generated from temperature-dependent fluorescence spectroscopy experiments showed that TMG-AspAsp and TMG-SerSer have minor destabilizing effects on the protein while TMG-AspSer and TMG-SerAsp strongly destabilize azurin. Red-shifted fluorescence signatures in the 25 °C correlate with the observed protein destabilization in the solutions with TMG-AspSer and TMG-SerAsp. These signals could correspond to interactions between the Asp residue in the dipeptide and the azurin Trp residue in the unfolded state. These results, supported by appropriate control experiments, suggest that dipeptide sequence-specific interactions lead to selective protein destabilization and motivate further studies of TMG-dipeptide ILs.

The impact of sub specialization within functional neurosurgery on patient outcomes in a comprehensive epilepsy center.

BACKGROUND: One in three patients with epilepsy are medication-refractory and may benefit from investigations and operative treatment at a comprehensive epilepsy center. However, while these centers have capabilities for advanced seizure monitoring and surgical intervention, they are not required to have a functional neurosurgeon who is primarily focused in epilepsy surgery. Therefore, the objective of this study is to determine the impact of having a sub-specialized, epilepsy-focused functional neurosurgeon on patient outcomes. METHODS: We conducted a retrospective chart review for all patients who underwent surgical intervention for medically refractory epilepsy at a Level 4 comprehensive Epilepsy Center from 2008 through 2019. Data was divided into two groups: group 1 comprised patients who had surgery before the hiring of a dedicated epilepsy-focused functional neurosurgeon in 2016, and group 2 was afterwards. We compared surgical procedures, significant complications, and seizure outcomes. RESULTS: A total of 101 patients underwent 105 operations (52 in group 1 and 53 in group 2), not including intracranial EEG insertion. Compared to group 1, group 2 had more surgeries performed per year (15.1 vs. 6.5), and a significantly lower Engel score at last follow-up (1.78 vs. 2.57; p < 0.001). There was no difference in percentage of cases undergoing iEEG, and no difference in complication rates. CONCLUSIONS: In this series, the hiring of a sub-specialized functional neurosurgeon dedicated to epilepsy surgery in a comprehensive epilepsy center was associated with an increase in surgical volume and improved seizure outcomes.

Thermodynamic destabilization of azurin by four different tetramethylguanidinium amino acid ionic liquids.

The thermal unfolding of the copper redox protein azurin was studied in the presence of four different amino acid-based ionic liquids (ILs), all of which have tetramethylguanidium as cation. The anionic amino acid includes two with alcohol side chains, serine and threonine, and two with carboxylic acids, aspartate and glutamate. Control experiments showed that amino acids alone do not significantly change protein stability and pH changes anticipated by the amino acid nature have only minor effects on the protein. With the ILs, the protein is destabilized and the melting temperature is decreased. The two ILs with alcohol side chains strongly destabilize the protein while the two ILs with acid side chains have weaker effects. Unfolding enthalpy (ΔHunf°) and entropy (ΔSunf°) values, derived from fits of the unfolding data, show that some ILs increase ΔHunf°while others do not significantly change this value. All ILs, however, increase ΔSunf°. MD simulations of both the folded and unfolded protein conformations in the presence of the ILs provide insight into the different IL-protein interactions and how they affect the ΔHunf° values. The simulations also confirm that the ILs increase the unfolded state entropies which can explain the increased ΔSunf° values.

Complexation of Mo in FLiNaK Molten Salt: Insight from Ab Initio Molecular Dynamics.

Online extraction of fission products, such as the medical isotope Mo-99, is a key advantage of the proposed molten salt nuclear reactor design. The chemical and structural behavior of Mo solvated in fluoride salt has been relatively unknown. Ab initio molecular dynamics simulations were employed to examine the behavior of molybdenum in the molten salt FLiNaK (LiF-NaF-KF) for oxidation states between 0 and 6+. Mo complexation was found to vary with the Mo oxidation state, with lower oxidation states tending to result in complexes with more molybdenum ions. Complexes containing multiple Mo ions were observed for all Mo oxidation states studied except 5+ and 6+. A relationship between the solubility of a complex and electronic isolation of a complex in a molten salt is explored using the Bader atoms in molecule electron density partitioning scheme, with more volatile complexes exhibiting greater electronic isolation. The impacts of UF4 and H2O on the predominant molybdenum species are also considered. While no impacts on Mo behavior by UF4 were observed, Mo-O interactions may inhibit the formation of complexes containing multiple Mo ions.

Weight management program for first responders: Feasibility study and lessons learned.

BACKGROUND: Overweight and obesity are significant concerns for first responders (firefighters and emergency medical service providers). OBJECTIVE: This pilot study examines the feasibility of a weight loss program for career first responders. METHODS: Participants were career first responders with BMI≥30 and interest in losing weight. Baseline height, weight, and body composition using dual x-ray absorptiometry, Pittsburgh Sleep Quality Index (PSQI), and a Functional Movement Screen (FMS) with follow-up weight and body composition were assessed. A 10-week weight loss program used evidence-based lifestyle goals (encouraged increasing activity levels, using tailored calorie goals, following the DASH diet, tracking with a phone app) and weekly in-person sessions with a student coach-in-training who reviewed weight and tracking information. RESULTS: Participants (N = 10) were 39.0±9.9 years old with a BMI of 35.6±4.7 kg/m2. There were significant reductions in weight -4.8±2.3 kg and percent body fat -3.9±1.7%. All participants had a PSQI score >5 indicating poor quality of sleep. Nine participants had FMS scores ≤14 indicating increased risk of injury. CONCLUSION: The use of commercially available apps with support from a student coach-in-training and evidence-based weight loss recommendations is a feasible approach to support career firefighters in their weight loss efforts.