T cell cholesterol transport is a metabolic checkpoint that links intestinal immune responses to dietary lipid absorption.

The intrinsic pathways that control membrane organization in immune cells and the impact of such pathways on cellular function are not well defined. Here we report that the non-vesicular cholesterol transporter Aster-A links plasma membrane (PM) cholesterol availability in T cells to immune signaling and systemic metabolism. Aster-A is recruited to the PM during T-cell receptor (TCR) activation, where it facilitates the removal of newly generated "accessible" membrane cholesterol. Loss of Aster-A leads to excess PM cholesterol accumulation, resulting in enhanced TCR nano-clustering and signaling, and Th17 cytokine production. Finally, we show that the mucosal Th17 response is restrained by PM cholesterol remodeling. Ablation of Aster-A in T cells leads to enhanced IL-22 production, reduced intestinal fatty acid absorption, and resistance to diet-induced obesity. These findings delineate a multi-tiered regulatory scheme linking immune cell lipid flux to nutrient absorption and systemic physiology.

IL7 in combination with radiotherapy stimulates a memory T-cell response to improve outcomes in HNSCC models.

Clinically approved head and neck squamous cell carcinoma (HNSCC) immunotherapies manipulate the immune checkpoint blockade (ICB) axis but have had limited success outside of recurrent/metastatic disease. Interleukin-7 (IL7) has been shown to be essential for effector T-cell survival, activation, and proliferation. Here, we show that IL7 in combination with radiotherapy (RT) is effective in activating CD8 + T-cells for reducing tumor growth. Our studies were conducted using both human papillomavirus related and unrelated orthotopic HNSCC murine models. Immune populations from the tumor, draining lymph nodes, and blood were compared between treatment groups and controls using flow cytometry, proteomics, immunofluorescence staining, and RNA sequencing. Treatment with RT and IL7 (RT + IL7) resulted in significant tumor growth reduction, high CD8 T-cell tumor infiltration, and increased proliferation of T-cell progenitors in the bone marrow. IL7 also expanded a memory-like subpopulation of CD8 T-cells. These results indicate that IL7 in combination with RT can serve as an effective immunotherapy strategy outside of the conventional ICB axis to drive the antitumor activity of CD8 T-cells.

Sensory nerve release of CGRP increases tumor growth in HNSCC by suppressing TILs.

BACKGROUND: Perineural invasion (PNI) and nerve density within the tumor microenvironment (TME) have long been associated with worse outcomes in head and neck squamous cell carcinoma (HNSCC). This prompted an investigation into how nerves within the tumor microenvironment affect the adaptive immune system and tumor growth. METHODS: We used RNA sequencing analysis of human tumor tissue from a recent HNSCC clinical trial, proteomics of human nerves from HNSCC patients, and syngeneic orthotopic murine models of HPV-unrelated HNSCC to investigate how sensory nerves modulate the adaptive immune system. FINDINGS: Calcitonin gene-related peptide (CGRP) directly inhibited CD8 T cell activity in vitro, and blocking sensory nerve function surgically, pharmacologically, or genetically increased CD8 and CD4 T cell activity in vivo. CONCLUSIONS: Our data support sensory nerves playing a role in accelerating tumor growth by directly acting on the adaptive immune system to decrease Th1 CD4 T cells and activated CD8 T cells in the TME. These data support further investigation into the role of sensory nerves in the TME of HNSCC and points toward the possible treatment efficacy of blocking sensory nerve function or specifically inhibiting CGRP release or activity within the TME to improve outcomes. FUNDING: 1R01DE028282-01, 1R01DE028529-01, 1P50CA261605-01 (to S.D.K.), 1R01CA284651-01 (to S.D.K.), and F31 DE029997 (to L.B.D.).

PPARα Agonism Enhances Immune Response to Radiotherapy while Dietary Oleic Acid Results in Counteraction.

INTRODUCTION: Head and neck cancer (HNC) improvements are stagnant, even with advances in immunotherapy. Our previous clinical trial data show altered fatty acid (FA) metabolism correlates with outcome. We hypothesized that pharmacologic and dietary modulation of FA catabolism will impact therapeutic efficacy. METHODS: We performed in vivo and in vitro experiments employing PPARα agonism with fenofibrate (FF) or high oleic acid diets (OAD) with radiotherapy, generating metabolomic, proteomic, stable isotope tracing, extracellular flux analysis, and flow cytometric data to investigate these alterations. RESULTS: FF improved anti-tumor efficacy of high dose per fraction radiotherapy in HNC murine models, while the OAD reversed this effect. FF treated mice on the control diet had evidence of increased FA catabolism. Stable isotope tracing showed less glycolytic utilization by ex vivo CD8+ T cells. Improved efficacy correlated with intratumoral alterations in eicosanoid metabolism and downregulated mTOR and CD36. CONCLUSION: Metabolic intervention with increased FA catabolism improves efficacy of HNC therapy and enhance anti-tumoral immune response.

Patient and Caregiver Perception of Adenoidectomies: A Non-Real-World Social Media Analysis.

Objective: To survey the social media outlets Twitter and Instagram for public posts related to adenoidectomy surgery. This study aims to investigate the attitudes and perceptions of patients and caregivers on social media, through thematic content-analysis of social media posts regarding adenoidectomy. Study Design: Non-real world qualitative study. Setting: Twitter and Instagram social media platforms. Methods: Public posts uploaded between February, 2021 and February, 2023 using the hashtags "#adenoidectomy," and "#adenoidectomyrecovery" were searched. Posts were excluded if they were unrelated to adenoidectomy or were in a non-English language. Relevant posts were stratified demographically as patient or caregiver and pre- or postoperative, and categorized into relevant themes for analysis. Outcomes were measured as the total number of posts. Results: A total of 394 relevant posts were analyzed. A significance threshold of P < 0.05 was used. Patients posted significantly more posts regarding procedure pain (P = 0.002) and concern for appearance (P = 0.048) compared to caregivers. Caregivers posted significantly (P < 0.001) more posts regarding condition awareness and were significantly (P < 0.001) more likely to spread positivity in their posts compared to patients themselves. Posts made by female caregivers were more likely to reference fear, while those made by male caregivers were more likely to provide education (P = 0.002). Conclusion: Patients may worry about appearance and mental health while caregivers are more likely to spread information and positivity. Male and female caregivers may also use social media differently. A better understanding of patient and caregiver concerns may optimize physician interaction and involvement.

Aster-dependent nonvesicular transport facilitates dietary cholesterol uptake.

Intestinal absorption is an important contributor to systemic cholesterol homeostasis. Niemann-Pick C1 Like 1 (NPC1L1) assists in the initial step of dietary cholesterol uptake, but how cholesterol moves downstream of NPC1L1 is unknown. We show that Aster-B and Aster-C are critical for nonvesicular cholesterol movement in enterocytes. Loss of NPC1L1 diminishes accessible plasma membrane (PM) cholesterol and abolishes Aster recruitment to the intestinal brush border. Enterocytes lacking Asters accumulate PM cholesterol and show endoplasmic reticulum cholesterol depletion. Aster-deficient mice have impaired cholesterol absorption and are protected against diet-induced hypercholesterolemia. Finally, the Aster pathway can be targeted with a small-molecule inhibitor to manipulate cholesterol uptake. These findings identify the Aster pathway as a physiologically important and pharmacologically tractable node in dietary lipid absorption.

Impact of community-based technology training with low-income older adults.

OBJECTIVES: Compared with younger and middle-aged adults, older adults are less likely to adopt new computer technology, potentially limiting access to healthcare and many other important resources available online. This limitation could impact cognitive abilities, well-being, and mental health outcomes of older adults. The aims of the present study were to increase access to online county and healthcare resources, while also assessing the impact of technology access on cognitive functioning and multiple well-being domains. METHODS: A pilot community collaboration provided a two-month tablet training intervention, focused on increasing digital independence via tablet navigation, resources access, and fraud and scam prevention, to 20 low-income older adult participants (75% female, Mage = 70.85). Pre- and post-test phone interviews were conducted to measure any changes in digital independence, cognitive abilities, well-being, mental health, and mindset. RESULTS: Linear mixed effects models revealed no significant changes in outcome measures from pre- to post-test. However, we found effects of digital independence on several well-being measures, providing important information for the impact of technology access and training for low-income older adults. CONCLUSION: This pilot intervention offers limited but promising results, inspiring further investigations that may inform public health and policy services to address barriers to access and potentially improve psychological health.

Headguard use in combat sports: position statement of the Association of Ringside Physicians.

Headguard use is appropriate during some combat sports activities where the risks of injury to the face and ears are elevated. Headguards are highly effective in reducing the incidence of facial lacerations in studies of amateur boxers and are just as effective in other striking sports. They should be used in scenarios - especially sparring prior to competitions - where avoidance of laceration and subsequent exposure to potential blood-borne pathogens is important. Headguards are appropriate where avoidance of auricular injury is deemed important; limited data show a marked reduction in incidence of auricular injury in wrestlers wearing headguards.Headguards should not be relied upon to reduce the risk of concussion or other traumatic brain injury. They have not been shown to prevent these types of injuries in combat sports or other sports, and human studies on the effect of headguards on concussive injury are lacking. While biomechanical studies suggest they reduce linear and rotational acceleration of the cranium, changes in athlete behavior to more risk-taking when wearing headguards may offset any risk reduction. In the absence of high-quality studies on headguard use, the Association of Ringside Physicians recommends that further research be conducted to clarify the role of headguards in all combat sports, at all ages of participation. Furthermore, in the absence of data on gender differences, policies should be standardized for men and women.

Aster-dependent non-vesicular transport facilitates dietary cholesterol uptake.

Intestinal cholesterol absorption is an important contributor to systemic cholesterol homeostasis. Niemann-Pick C1 Like 1 (NPC1L1), the target of the drug ezetimibe (EZ), assists in the initial step of dietary cholesterol uptake. However, how cholesterol moves downstream of NPC1L1 is unknown. Here we show that Aster-B and Aster-C are critical for non-vesicular cholesterol movement in enterocytes, bridging NPC1L1 at the plasma membrane (PM) and ACAT2 in the endoplasmic reticulum (ER). Loss of NPC1L1 diminishes accessible PM cholesterol in enterocytes and abolishes Aster recruitment to the intestinal brush border. Enterocytes lacking Asters accumulate cholesterol at the PM and display evidence of ER cholesterol depletion, including decreased cholesterol ester stores and activation of the SREBP-2 transcriptional pathway. Aster-deficient mice have impaired cholesterol absorption and are protected against diet-induced hypercholesterolemia. Finally, we show that the Aster pathway can be targeted with a small molecule inhibitor to manipulate dietary cholesterol uptake. These findings identify the Aster pathway as a physiologically important and pharmacologically tractable node in dietary lipid absorption. One-Sentence Summary: Identification of a targetable pathway for regulation of dietary cholesterol absorption.

Leptomeningeal Metastasis: A Review of the Pathophysiology, Diagnostic Methodology, and Therapeutic Landscape.

The present review aimed to establish an understanding of the pathophysiology of leptomeningeal disease as it relates to late-stage development among different cancer types. For our purposes, the focused metastatic malignancies include breast cancer, lung cancer, melanoma, primary central nervous system tumors, and hematologic cancers (lymphoma, leukemia, and multiple myeloma). Of note, our discussion was limited to cancer-specific leptomeningeal metastases secondary to the aforementioned primary cancers. LMD mechanisms secondary to non-cancerous pathologies, such as infection or inflammation of the leptomeningeal layer, were excluded from our scope of review. Furthermore, we intended to characterize general leptomeningeal disease, including the specific anatomical infiltration process/area, CSF dissemination, manifesting clinical symptoms in patients afflicted with the disease, detection mechanisms, imaging modalities, and treatment therapies (both preclinical and clinical). Of these parameters, leptomeningeal disease across different primary cancers shares several features. Pathophysiology regarding the development of CNS involvement within the mentioned cancer subtypes is similar in nature and progression of disease. Consequently, detection of leptomeningeal disease, regardless of cancer type, employs several of the same techniques. Cerebrospinal fluid analysis in combination with varied imaging (CT, MRI, and PET-CT) has been noted in the current literature as the gold standard in the diagnosis of leptomeningeal metastasis. Treatment options for the disease are both varied and currently in development, given the rarity of these cases. Our review details the differences in leptomeningeal disease as they pertain through the lens of several different cancer subtypes in an effort to highlight the current state of targeted therapy, the potential shortcomings in treatment, and the direction of preclinical and clinical treatments in the future. As there is a lack of comprehensive reviews that seek to characterize leptomeningeal metastasis from various solid and hematologic cancers altogether, the authors intended to highlight not only the overlapping mechanisms but also the distinct patterning of disease detection and progression as a means to uniquely treat each metastasis type. The scarcity of LMD cases poses a barrier to more robust evaluations of this pathology. However, as treatments for primary cancers have improved over time, so has the incidence of LMD. The increase in diagnosed cases only represents a small fraction of LMD-afflicted patients. More often than not, LMD is determined upon autopsy. The motivation behind this review stems from the increased capacity to study LMD in spite of scarcity or poor patient prognosis. In vitro analysis of leptomeningeal cancer cells has allowed researchers to approach this disease at the level of cancer subtypes and markers. We ultimately hope to facilitate the clinical translation of LMD research through our discourse.

Synthesis and crystal structure of 2-[(2,3,5,6-tetra-fluoro-pyridin-4-yl)amino]-ethyl methacrylate.

In the title compound, C11H10F4N2O2, the conformation about the N-C-C-O bond is gauche [torsion angle = 61.84 (13)°]. In the crystal, N-H⋯O hydrogen bonds link the mol-ecules into [010] chains, which are cross-linked by C-H⋯F and C-H⋯π contacts. Hirshfeld surface analysis was conducted to aid in the visualization of these various influences on the packing. This analysis showed that the largest contribution to the surface contacts arises from F⋯H/H⋯F inter-actions (35.6%), followed by O⋯H/H⋯O (17.8%) and H⋯H (12.7%).

The ATPase activity of the phosphatidylethanolamine flippase TAT-5 inhibits extracellular vesicle budding from the plasma membrane.

Cells release extracellular vesicles (EVs) from their surface, but the mechanisms that govern EV release by plasma membrane budding are poorly understood. The lipid flippase TAT-5 inhibits EV release from the plasma membrane in C. elegans , but how the level of flippase activity regulates EV release was unknown. We generated point mutations in the DGET motif of TAT-5 predicted to lead to a partial or complete loss of ATPase activity. We discovered that tat-5(E246Q) mutants were sterile, while tat-5(D244T) mutants produced embryos that arrested during development. Using degron-based reporters, we found that EV release was increased in tat-5(D244T) mutant embryos and that phagocytosis was also disrupted. These data suggest that a low level of flippase activity can promote fertility, while a higher level of flippase activity is required to inhibit EV release, allow phagocytosis, and carry out embryonic development.

Top-down patterning of topological surface and edge states using a focused ion beam.

The conducting boundary states of topological insulators appear at an interface where the characteristic invariant ℤ2 switches from 1 to 0. These states offer prospects for quantum electronics; however, a method is needed to spatially-control ℤ2 to pattern conducting channels. It is shown that modifying Sb2Te3 single-crystal surfaces with an ion beam switches the topological insulator into an amorphous state exhibiting negligible bulk and surface conductivity. This is attributed to a transition from ℤ2 = 1 → ℤ2 = 0 at a threshold disorder strength. This observation is supported by density functional theory and model Hamiltonian calculations. Here we show that this ion-beam treatment allows for inverse lithography to pattern arrays of topological surfaces, edges and corners which are the building blocks of topological electronics.

Neutrophil to Lymphocyte Ratio as a Predictor of Postoperative Outcomes in Traumatic Brain Injury: A Systematic Review and Meta-Analysis.

(1) Introduction: Traumatic brain injury (TBI) is a leading cause of injury and mortality worldwide, carrying an estimated cost of $38 billion in the United States alone. Neutrophil to lymphocyte ratio (NLR) has been investigated as a standardized biomarker that can be used to predict outcomes of TBI. The aim of this review was to determine the prognostic utility of NLR among patients admitted for TBI. (2) Methods: A literature search was conducted in PubMed, Scopus, and Web of Science in November 2022 to retrieve articles regarding the use of neutrophil to lymphocyte ratio (NLR) as a prognostic measure in traumatic brain injury (TBI) patients. Inclusion criteria included studies reporting outcomes of TBI patients with associated NLR values. Exclusion criteria were studies reporting only non-primary data, those insufficiently disaggregated to extract NLR data, and non-English or cadaveric studies. The Newcastle-Ottawa Scale was utilized to assess for the presence of bias in included studies. (3) Results: Following the final study selection 19 articles were included for quantitative and qualitative analysis. The average age was 46.25 years. Of the 7750 patients, 73% were male. Average GCS at presentation was 10.51. There was no significant difference in the NLR between surgical vs. non-surgical cohorts (SMD 2.41 95% CI -1.82 to 6.63, p = 0.264). There was no significant difference in the NLR between bleeding vs. non-bleeding cohorts (SMD 4.84 95% CI -0.26 to 9.93, p = 0.0627). There was a significant increase in the NLR between favorable vs. non-favorable cohorts (SMD 1.31 95% CI 0.33 to 2.29, p = 0.0090). (4) Conclusions: Our study found that NLR was only significantly predictive for adverse outcomes in TBI patients and not surgical treatment or intracranial hemorrhage, making it nonetheless an affordable alternative for physicians to assess patient prognosis.

Hepatic nonvesicular cholesterol transport is critical for systemic lipid homeostasis.

In cell models, changes in the 'accessible' pool of plasma membrane (PM) cholesterol are linked with the regulation of endoplasmic reticulum sterol synthesis and metabolism by the Aster family of nonvesicular transporters; however, the relevance of such nonvesicular transport mechanisms for lipid homeostasis in vivo has not been defined. Here we reveal two physiological contexts that generate accessible PM cholesterol and engage the Aster pathway in the liver: fasting and reverse cholesterol transport. During fasting, adipose-tissue-derived fatty acids activate hepatocyte sphingomyelinase to liberate sequestered PM cholesterol. Aster-dependent cholesterol transport during fasting facilitates cholesteryl ester formation, cholesterol movement into bile and very low-density lipoprotein production. During reverse cholesterol transport, high-density lipoprotein delivers excess cholesterol to the hepatocyte PM through scavenger receptor class B member 1. Loss of hepatic Asters impairs cholesterol movement into feces, raises plasma cholesterol levels and causes cholesterol accumulation in peripheral tissues. These results reveal fundamental mechanisms by which Aster cholesterol flux contributes to hepatic and systemic lipid homeostasis.

Effect of simulated body fluid formulation on orthopedic device apatite-forming ability assessment.

Integration of native bone into orthopedic devices is a key factor in long-term implant success. The material-tissue interface is generally accepted to consist of a hydroxyapatite layer so bioactive materials that can spontaneously generate this hydroxyapatite layer after implantation may improve patient outcomes. Per the ISO 22317:2014 standard, "Implants for surgery - In vitro evaluation for apatite-forming ability of implant materials," bioactivity performance statements can be assessed by soaking the material in simulated body fluid (SBF) and evaluating the surface for the formation of a hydroxyapatite layer; however, variations in test methods may alter hydroxyapatite formation and result in false-positive assessments. The goal of this study was to identify the effect of SBF formulation on bioactivity assessment. Bioglass® (45S5 and S53P4) and non-bioactive Ti-6Al-4V were exposed to SBF formulations varying in calcium ion and phosphate concentrations as well as supporting ion concentrations. Scanning electron microscopy and X-ray powder diffraction evaluation of the resulting hydroxyapatite layers revealed that SBF enriched with double or quadruple the calcium and phosphate ion concentrations increased hydroxyapatite crystal size and quantity compared to the standard formulation and can induce hydroxyapatite crystallization on surfaces traditionally considered non-bioactive. Altering concentrations of other ions, for example, bicarbonate, changed hydroxyapatite induction time, quantity, and morphology. For studies evaluating the apatite-forming ability of a material to support bioactivity performance statements, test method parameters must be adequately described and controlled. It is unclear if apatite formation after exposure to any of the SBF formulations is representative of an in vivo biological response. The ISO 23317 standard test method should be further developed to provide additional guidance on apatite characterization and interpretation of the results.

Conserved N- and C-terminal motifs of PAD-1 are required to inhibit extracellular vesicle release.

Cells release extracellular vesicles (EVs) carrying cargos that can influence development and disease, but the mechanisms that govern EV release by plasma membrane budding are poorly understood. We previously showed that the Dopey protein PAD-1 inhibits EV release from the plasma membrane in C. elegans . However, PAD-1 is large, and the domains required to regulate EV release were unknown. Here, we reveal that the conserved N-terminal EWAD motif and C-terminal leucine zippers are required to inhibit EV release from the plasma membrane. Revealing a role for these domains is an important first step to identifying how EV release is regulated.

Arterial Occlusion and Acute Deep Vein Thrombosis-Induced Acute Limb Ischemia in a COVID-19 Patient.

Coronavirus disease 2019 (COVID-19) is a viral illness known to elicit a hypercoagulable state leading to a myriad of vascular pathologies. Over the course of the COVID-19 pandemic, widespread insults to the venous system have been well documented, with an increasing number of arterial events being reported. Despite the rising incidence of both pathological manifestations, these events are rare, but when present, serve as significant life threats to the patient in question. We report and discuss a case of a 69-year-old female with no thromboembolic risk factors or systemic signs of illness who presented with signs and symptoms consistent with acute limb ischemia (ALI). The patient was ultimately found to have occlusion of multiple arterial and venous vessels. She tested positive for COVID-19 despite being otherwise asymptomatic from a viral syndrome standpoint. To our knowledge, there are no reports in the medical literature of ALI - in the setting of arterial occlusion and concomitant deep vein thrombosis (DVT) - as the sole clinical manifestation in an asymptomatic patient without thrombotic risk factors who was only incidentally found to be COVID-19-positive. This case underscores the atypical manifestations and deleterious effects associated with COVID-19 and the need to have a high index of suspicion for a multitude of pathologies when facing this viral illness.

Cannot Ventilate: An Unexpected Cause of Respiratory Failure in a Ten-Year-Old Child.

Granulomatosis with polyangiitis (GPA) is a necrotizing vasculitis known to affect the respiratory and renal systems. There are a multitude of clinical manifestations, many of which are not specific to the disease, such as dysfunction of the nasal, sinus, auditory, tracheal, pulmonary, ocular, renal, cardiac, and nervous systems. As a multisystemic illness without a "classic" presentation and insidious progression, it is often a challenging diagnosis. We report and discuss a case of a 10-year-old female with no significant past medical history who presented to the emergency department with a 10-day course of worsening respiratory symptoms. As her respiratory and clinical status began to precipitously decline, the decision was made to intubate the patient, which was performed without issue. Unfortunately, attempts at oxygenating and ventilating the patient were met with extreme resistance and difficulty-an airway situation that could have been catastrophic if not for quick reaction maneuvers performed that would ultimately go on to remedy the issue at hand. We hope to raise awareness regarding the airway challenges posed by GPA and delve into its management as a means of improving recognition and preparing clinicians to treat this condition.

Micromolding of Amphotericin-B-Loaded Methoxyethylene-Maleic Anhydride Copolymer Microneedles.

Biocompatible and biodegradable materials have been used for fabricating polymeric microneedles to deliver therapeutic drug molecules through the skin. Microneedles have advantages over other drug delivery methods, such as low manufacturing cost, controlled drug release, and the reduction or absence of pain. The study examined the delivery of amphotericin B, an antifungal agent, using microneedles that were fabricated using a micromolding technique. The microneedle matrix was made from GantrezTM AN-119 BF, a benzene-free methyl vinyl ether/maleic anhydride copolymer. The GantrezTM AN-119 BF was mixed with water; after water evaporation, the polymer exhibited sufficient strength for microneedle fabrication. Molds cured at room temperature remained sharp and straight. SEM images showed straight and sharp needle tips; a confocal microscope was used to determine the height and tip diameter for the microneedles. Nanoindentation was used to obtain the hardness and Young's modulus values of the polymer. Load-displacement testing was used to assess the failure force of the needles under compressive loading. These two mechanical tests confirmed the mechanical properties of the needles. In vitro studies validated the presence of amphotericin B in the needles and the antifungal properties of the needles. Amphotericin B GantrezTM microneedles fabricated in this study showed appropriate characteristics for clinical translation in terms of mechanical properties, sharpness, and antifungal properties.