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Platinum exhibits desirable catalytic properties, but it is scarce and expensive. Optimizing its use in key applications such as emission control catalysis is important to reduce our reliance on such a rare element. Supported Pt nanoparticles (NPs) used in emission control systems deactivate over time because of particle growth in sintering processes. In this work, we shed light on the stability against sintering of Pt NPs supported on and encapsulated in Al2O3 using a combination of nanocrystal catalysts and atomic layer deposition (ALD) techniques. We find that small amounts of alumina overlayers created by ALD on preformed Pt NPs can stabilize supported Pt catalysts, significantly reducing deactivation caused by sintering, as previously observed by others. Combining theoretical and experimental insights, we correlate this behavior to the decreased propensity of oxidized Pt species to undergo Ostwald ripening phenomena because of the physical barrier imposed by the alumina overlayers. Furthermore, we find that highly stable catalysts can present an abundance of under-coordinated Pt sites after restructuring of both Pt particles and alumina overlayers at a high temperature (800 °C) in C3H6 oxidation conditions. The enhanced stability significantly improves the Pt utilization efficiency after accelerated aging treatments, with encapsulated Pt catalysts reaching reaction rates more than two times greater than those of a control supported Pt catalyst.
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Introduction: The use of telemedicine and ultrasound is emerging and novel in the field of community paramedicine. However, there is a paucity of data supporting its use and even less evidence that shows a morbidity and mortality benefit. This case highlights a unique way to diagnose a common medical emergency, which can lead to a good outcome. Case Report: We describe the use of lung point-of-care ultrasound by a trained community paramedic that led to the identification of a pneumothorax in an 86-year-old male at a scheduled home visit. The images were interpreted over telehealth in real-time by an emergency physician, and the patient was transported to the emergency department where the diagnosis was confirmed by chest radiography. He underwent chest tube placement and was discharged five days later after returning to his baseline. Conclusion: Despite minimal data to support or refute the use of paramedic tele-ultrasound, this case highlights a unique opportunity to expand the use of telemedicine and ultrasound in community paramedicine to improve patient outcomes.
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The Metabolic Bone Health Department, Cardiff and Vale University Health Board, serves a local population of approximately 445 000 people. A retrospective audit of attendance data regarding the denosumab treatment clinic (the traditional treatment pathway) and the denosumab Self-Injection Program (SIP) was conducted to determine whether the SIP is both cost-effective and environmentally beneficial, compared to the traditional treatment pathway. Cost analysis was then conducted by the Finance Department. The audit was conducted over 3 years following the implementation of the service development; 233 patients had enrolled in the program at the time of the audit and 69 had completed 3 years of self-injected treatment. A control group of 497 patients were identified by the service. This group remained on the historical pathway and had consistent attendance activity over the 3-yr period from 2017 to 2019. Pre- and post-period activity of all patients on the program was compared, together with the activity for the independent control group. The SIP resulted in a reduction in clinical contacts, with financial analysis showing a total opportunity cost saving per patient of £420 per annum. There were obvious benefits to the patient of a reduced number of visits to a clinical site, which also resulted in an estimated carbon footprint reduction of 59 kg CO2 per patient per annum. The cost analysis is based on our organization's 2022 charges. The SIP demonstrates that by focusing on care "closer to home", it is possible to maximize resources, improve the patient experience through reduced travel, and reduce the environmental impact of healthcare.
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Multicomponent catalysts can be designed to synergistically combine reaction intermediates at interfacial active sites, but restructuring makes systematic control and understanding of such dynamics challenging. We here unveil how reducibility and mobility of indium oxide species in Ru-based catalysts crucially control the direct, selective conversion of CO2 to ethanol. When uncontrolled, reduced indium oxide species occupy the Ru surface, leading to deactivation. With the addition of steam as a mild oxidant and using porous polymer layers to control In mobility, Ru-In2O3 interface sites are stabilized, and ethanol can be produced with superior overall selectivity (70%, rest CO). Our work highlights how engineering of bifunctional active ensembles enables cooperativity and synergy at tailored interfaces, which unlocks unprecedented performance in heterogeneous catalysts.
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Background: The role of biceps tenodesis (BT) during open reduction internal fixation (ORIF) of proximal humerus fractures (PHFs) remains unclear. A subset of patients undergoing ORIF have persistent pain with unclear etiology. The purpose of our study was to compare outcomes of ORIF of PHFs with and without concomitant BT. We hypothesize patients undergoing BT at the time of ORIF will have improved patient-reported outcome scores with fewer secondary procedures related to treatment of the biceps. Methods: In this retrospective cohort study, all patients undergoing ORIF for a PHF at a single level one trauma center from January 2019 to June 2022 were reviewed. Patients under the age of 18 were excluded. Primary outcomes were patient-reported outcomes measurement information system physical function, depression, and pain interference scores at 5 time points up to final follow-up. Secondary outcomes included total operative time, complications, subsequent procedures, steroid injections, and range of motion. Chi-square tests were performed for categorical values and paired t-tests for continuous variables. Results: 71 patients met inclusion criteria: 41 undergoing ORIF without BT and 30 undergoing ORIF with BT. Average follow-up was 11 months. There were no statistically significant differences in patient demographics between groups. There were no differences in patient-reported outcomes measurement information system scores at any time point postoperatively. At final follow-up patients in the ORIF with BT group had higher forward flexion than those who did not undergo BT (142 vs. 123 degrees, respectively, P < .02). There were no differences in surgical time, revision rates, postsurgical complications, or postoperative injections between groups. Conclusion: BT performed during ORIF of PHFs did not result in significantly different functional or patient-reported outcomes between groups, except for greater forward flexion at final follow-up. Although BT was done more commonly in severe PHFs, patients in both groups had similar rates of subsequent biceps-related procedures and revision surgery.
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ABSTRACT: Stone, MH, Hornsby, G, Mizuguchi, S, Sato, K, Gahreman, D, Duca, M, Carroll, K, Ramsey, MW, Stone, ME, and Haff, GG. The use of free weight squats in sports: a narrative review-squatting movements, adaptation, and sports performance: physiological. J Strength Cond Res 38(8): 1494-1508, 2024-The squat and its variants can provide numerous benefits including positively affecting sports performance and injury prevention, injury severity reduction, and rehabilitation. The positive benefits of squat are likely the result of training-induced neural alterations and mechanical and morphological adaptations in tendons, skeletal muscles, and bones, resulting in increased tissue stiffness and cross-sectional area (CSA). Although direct evidence is lacking, structural adaptations can also be expected to occur in ligaments. These adaptations are thought to beneficially increase force transmission and mechanical resistance (e.g., resistance to mechanical strain) and reduce the likelihood and severity of injuries. Adaptations such as these, also likely play an important role in rehabilitation, particularly for injuries that require restricted use or immobilization of body parts and thus lead to a consequential reduction in the CSA and alterations in the mechanical properties of tendons, skeletal muscles, and ligaments. Both volume and particularly intensity (e.g., levels of loading used) of training seem to be important for the mechanical and morphological adaptations for at least skeletal muscles, tendons, and bones. Therefore, the training intensity and volume used for the squat and its variations should progressively become greater while adhering to the concept of periodization and recognized training principles.
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Adaptação Fisiológica , Desempenho Atlético , Músculo Esquelético , Humanos , Desempenho Atlético/fisiologia , Adaptação Fisiológica/fisiologia , Músculo Esquelético/fisiologia , Treinamento Resistido/métodos , Movimento/fisiologia , Tendões/fisiologia , Fenômenos BiomecânicosRESUMO
Reductive catalytic fractionation (RCF) is a promising method to extract and depolymerize lignin from biomass, and bench-scale studies have enabled considerable progress in the past decade. RCF experiments are typically conducted in pressurized batch reactors with volumes ranging between 50 and 1000 mL, limiting the throughput of these experiments to one to six reactions per day for an individual researcher. Here, we report a high-throughput RCF (HTP-RCF) method in which batch RCF reactions are conducted in 1 mL wells machined directly into Hastelloy reactor plates. The plate reactors can seal high pressures produced by organic solvents by vertically stacking multiple reactor plates, leading to a compact and modular system capable of performing 240 reactions per experiment. Using this setup, we screened solvent mixtures and catalyst loadings for hydrogen-free RCF using 50 mg poplar and 0.5 mL reaction solvent. The system of 1:1 isopropanol/methanol showed optimal monomer yields and selectivity to 4-propyl substituted monomers, and validation reactions using 75 mL batch reactors produced identical monomer yields. To accommodate the low material loadings, we then developed a workup procedure for parallel filtration, washing, and drying of samples and a 1H nuclear magnetic resonance spectroscopy method to measure the RCF oil yield without performing liquid-liquid extraction. As a demonstration of this experimental pipeline, 50 unique switchgrass samples were screened in RCF reactions in the HTP-RCF system, revealing a wide range of monomer yields (21-36%), S/G ratios (0.41-0.93), and oil yields (40-75%). These results were successfully validated by repeating RCF reactions in 75 mL batch reactors for a subset of samples. We anticipate that this approach can be used to rapidly screen substrates, catalysts, and reaction conditions in high-pressure batch reactions with higher throughput than standard batch reactors.
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BACKGROUND: Artificial intelligence (AI) uses computer systems to simulate cognitive capacities to accomplish goals like problem-solving and decision-making. Machine learning (ML), a branch of AI, makes algorithms find connections between preset variables, thereby producing prediction models. ML can aid shoulder surgeons in determining which patients may be susceptible to worse outcomes and complications following shoulder arthroplasty (SA) and align patient expectations following SA. However, limited literature is available on ML utilization in total shoulder arthroplasty (TSA) and reverse TSA. METHODS: A systematic literature review in accordance with PRISMA guidelines was performed to identify primary research articles evaluating ML's ability to predict SA outcomes. With duplicates removed, the initial query yielded 327 articles, and after applying inclusion and exclusion criteria, 12 articles that had at least 1 month follow-up time were included. RESULTS: ML can predict 30-day postoperative complications with a 90% accuracy, postoperative range of motion with a higher-than-85% accuracy, and clinical improvement in patient-reported outcome measures above minimal clinically important differences with a 93%-99% accuracy. ML can predict length of stay, operative time, discharge disposition, and hospitalization costs. CONCLUSION: ML can accurately predict outcomes and complications following SA and healthcare utilization. Outcomes are highly dependent on the type of algorithms used, data input, and features selected for the model. LEVEL OF EVIDENCE: III.
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The base excision repair (BER) pathway is a precise and versatile mechanism of DNA repair that is initiated by DNA glycosylases. Endonuclease VIII-like 1 (NEIL1) is a bifunctional glycosylase/abasic site (AP) lyase that excises a damaged base and subsequently cleaves the phosphodiester backbone. NEIL1 is able to recognize and hydrolyze a broad range of oxidatively-induced base lesions and substituted ring-fragmented guanines, including aflatoxin-induced 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl-formamido)-9-hydroxyaflatoxin B1 (AFB1-FapyGua). Due to NEIL1's protective role against these and other pro-mutagenic lesions, it was hypothesized that naturally occurring single nucleotide polymorphic (SNP) variants of NEIL1 could increase human risk for aflatoxin-induced hepatocellular carcinoma (HCC). Given that populations in South Asia experience high levels of dietary aflatoxin exposures and hepatitis B viral infections that induce oxidative stress, investigations on SNP variants of NEIL1 that occur in this region may have clinical implications. In this study, the most common South Asian variants of NEIL1 were expressed, purified, and functionally characterized. All tested variants exhibited activities and substrate specificities similar to wild type (wt)-NEIL1 on high-molecular weight DNA containing an array of oxidatively-induced base lesions. On short oligodeoxynucleotides (17-mers) containing either a site-specific apurinic/apyrimidinic (AP) site, thymine glycol (ThyGly), or AFB1-FapyGua, P206L-NEIL1 was catalytically comparable to wt-NEIL1, while the activities of NEIL1 variants Q67K and T278I on these substrates were ≈2-fold reduced. Variant T103A had a greatly diminished ability to bind to 17-mer DNAs, limiting the subsequent glycosylase and lyase reactions. Consistent with this observation, the rate of excision by T103A on 17-mer oligodeoxynucleotides containing ThyGly or AFB1-FapyGua could not be measured. However, the ability of T103A to excise ThyGly was improved on longer oligodeoxynucleotides (51-mers), with ≈7-fold reduced activity compared to wt-NEIL1. Our studies suggest that NEIL1 variant T103A may present a pathogenic phenotype that is limited in damage recognition, potentially increasing human risk for HCC.
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DNA Glicosilases , Reparo do DNA , Polimorfismo de Nucleotídeo Único , DNA Glicosilases/metabolismo , DNA Glicosilases/genética , DNA Glicosilases/química , Humanos , Aflatoxina B1/metabolismo , Dano ao DNA , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/enzimologia , Especificidade por Substrato , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/enzimologiaRESUMO
Background: Medial ulnar collateral ligament (mUCL) injury can cause significant pain and alter throwing mechanics. Common autograft options for mUCL reconstruction (UCLR) include the palmaris longus (PL) and hamstring tendons. Allograft use may reduce donor site morbidity and decrease function related to PL autografts. Purpose: To compare varus stability and load to failure between a novel allograft for UCLR-knee medial collateral ligament (kMCL)-and a PL autograft in human donor elbow specimens. Study Design: Controlled laboratory study. Methods: A total of 24 fresh-frozen human elbows were dissected to expose the mUCL. Medial elbow stability was tested with the mUCL intact (native), deficient, and reconstructed utilizing the humeral single-docking technique with either a (1) kMCL allograft (n = 12) or (2) a PL autograft (n = 12). A 3-N·m valgus torque was applied to the elbow, and valgus rotation of the ulna was recorded via motion tracking cameras. The elbow was cycled through a full range of motion 5 times. After kinematic testing, specimens were loaded to failure at 70° of elbow flexion, and failure modes were recorded. Results: The mUCL-deficient elbows demonstrated significantly greater valgus rotation compared with the intact and reconstructed elbows at every flexion angle tested (10°-120°) (P <.001). Both kMCL- and PL-reconstructed elbows exhibited significantly higher mean valgus rotation compared with the intact state between 10° and 40° of flexion (P < .01). There were no significant differences in valgus rotation at any flexion angle between the kMCL and PL graft groups. When loaded to failure, elbows reconstructed with both kMCL and PL grafts failed at similar torque values (18.6 ± 4 and 18.1 ± 3.4 N·m, respectively; P = .765). Conclusion: Fresh-frozen and aseptically processed kMCL allografts demonstrated similar kinematic and failure properties to PL tendon autografts in UCL-reconstructed elbows, although neither graft fully restored kinematics between 10° and 40°. Clinical Relevance: Prepared kMCL ligament allografts may provide a viable graft material when reconstructing elbow ligaments while avoiding the potential complications related to PL autografts- including donor site morbidity.
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Aflatoxin B1 (AFB1) is a potent human liver carcinogen produced by certain molds, particularly Aspergillus flavus and Aspergillus parasiticus, which contaminate peanuts, corn, rice, cottonseed, and ground and tree nuts, principally in warm and humid climates. AFB1 undergoes bioactivation in the liver to produce AFB1-exo-8,9-epoxide, which forms the covalently bound cationic AFB1-N7-guanine (AFB1-N7-Gua) DNA adduct. This adduct is unstable and undergoes base-catalyzed opening of the guanine imidazolium ring to form two ring-opened diastereomeric 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl-formamido)-9-hydroxy-aflatoxin B1 (AFB1-FapyGua) adducts. The AFB1 formamidopyrimidine (Fapy) adducts induce G â T transversion mutations and are likely responsible for the carcinogenic effects of AFB1. Quantitative liquid chromatography-mass spectrometry (LC-MS) methods have shown that AFB1-N7-Gua is eliminated in rodent and human urine, whereas ring-opened AFB1-FapyGua adducts persist in rodent liver. However, fresh frozen biopsy tissues are seldom available for biomonitoring AFB1 DNA adducts in humans, impeding research advances in this potent liver carcinogen. In contrast, formalin-fixed paraffin-embedded (FFPE) specimens used for histopathological analysis are often accessible for molecular studies. However, ensuring nucleic acid quality presents a challenge due to incomplete reversal of formalin-mediated DNA cross-links, which can preclude accurate quantitative measurements of DNA adducts. In this study, employing ion trap or high-resolution accurate Orbitrap mass spectrometry, we demonstrate that ring-opened AFB1-FapyGua adducts formed in AFB1-exposed newborn mice are stable to the formalin fixation and DNA de-cross-linking retrieval processes. The AFB1-FapyGua adducts can be detected at levels comparable to those in a match of fresh frozen liver. Orbitrap MS2 measurements can detect AFB1-FapyGua at a quantification limit of 4.0 adducts per 108 bases when only 0.8 µg of DNA is assayed on the column. Thus, our breakthrough DNA retrieval technology can be adapted to screen for AFB1 DNA adducts in FFPE human liver specimens from cohorts at risk of this potent liver carcinogen.
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Aflatoxina B1 , Adutos de DNA , Camundongos , Humanos , Animais , Aflatoxina B1/química , Inclusão em Parafina , DNA/metabolismo , Carcinógenos/metabolismo , Espectrometria de Massas , Guanina , FormaldeídoRESUMO
Urea lesions in DNA arise from thymine glycol (Tg) or 8-oxo-dG; their genotoxicity is thought to arise in part due to their potential to accommodate the insertion of all four dNTPs during error-prone replication. Replication bypass with human DNA polymerase η (hPol η) confirmed that all four dNTPs were inserted opposite urea lesions but with purines exhibiting greater incorporation efficiency. X-ray crystal structures of ternary replication bypass complexes in the presence of Mg2+ ions with incoming dNTP analogs dAMPnPP, dCMPnPP, dGMPnPP, and dTMPnPP bound opposite urea lesions (hPol η·DNA·dNMPnPP complexes) revealed all were accommodated by hPol η. In each, the Watson-Crick face of the dNMPnPP was paired with the urea lesion, exploiting the ability of the amine and carbonyl groups of the urea to act as H-bond donors or acceptors, respectively. With incoming dAMPnPP or dGMPnPP, the distance between the imino nitrogen of urea and the N9 atoms of incoming dNMPnPP approximated the canonical distance of 9 Å in B-DNA. With incoming dCMPnPP or dTMPnPP, the corresponding distance of about 7 Å was less ideal. Improved base-stacking interactions were also observed with incoming purines vs pyrimidines. Nevertheless, in each instance, the α-phosphate of incoming dNMPnPPs was close to the 3'-hydroxyl group of the primer terminus, consistent with the catalysis of nucleotidyl transfer and the observation that all four nucleotides could be inserted opposite urea lesions. Preferential insertion of purines by hPol η may explain, in part, why the urea-directed spectrum of mutations arising from Tg vs 8-oxo-dG lesions differs.
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Dano ao DNA , DNA Polimerase Dirigida por DNA , Humanos , 8-Hidroxi-2'-Desoxiguanosina , DNA Polimerase Dirigida por DNA/metabolismo , DNA/química , Replicação do DNA , Nucleotídeos , Adutos de DNARESUMO
OBJECTIVES: Because of its low frequency in adult populations and clinical and laboratory overlap with hemophagocytic lymphohistiocytosis and other T-cell lymphomas, T-cell/natural killer (NK) cell systemic, chronic, active Epstein-Barr virus (EBV) (T/NK sCAEBV) infection remains underdiagnosed, preventing critical, prompt therapeutic interventions. METHODS: We report a 5-case series that included 2 adult patients with T/NK sCAEBV and 3 additional adult patients with T/NK lymphomas with concomitant systemic EBV infection to review these entities' overlapping diagnostic and clinical features. RESULTS: Approximately 95% of the world population has been infected with EBV during their lifetime, and infection is usually asymptomatic, with symptomatic cases eventually resolving spontaneously. A small subset of immunocompetent patients develops CAEBV, a life-threatening complication resulting from EBV-infected T-cell or NK cell neoplastic lymphocytes. The sites of end-organ damage in T/NK sCAEBV demonstrate pathologic findings such as reactive lymphoid proliferations, making the diagnosis difficult to establish, with the only curative option being an allogeneic hematopoietic stem cell transplant. CONCLUSIONS: This diagnosis is most prevalent in Asia, with few cases reported in Western countries. Adult age is an independent risk factor for poor outcomes, and most cases are diagnosed in pediatric populations.
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Infecções por Vírus Epstein-Barr , Humanos , Infecções por Vírus Epstein-Barr/complicações , Infecções por Vírus Epstein-Barr/diagnóstico , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Células Matadoras Naturais/patologia , Células Matadoras Naturais/imunologia , Transtornos Linfoproliferativos/virologia , Transtornos Linfoproliferativos/diagnóstico , Transtornos Linfoproliferativos/patologia , Herpesvirus Humano 4/isolamento & purificação , Idoso , Doença Crônica , Linfoma Extranodal de Células T-NK/patologia , Linfoma Extranodal de Células T-NK/virologia , Linfoma Extranodal de Células T-NK/diagnósticoRESUMO
ABSTRACT: Chae, S, Long, SA, Lis, RP, McDowell, KW, Wagle, JP, Carroll, KM, Mizuguchi, S, and Stone, MH. Combined accentuated eccentric loading and rest redistribution in high-volume back squat: Acute stimulus and fatigue. J Strength Cond Res 38(4): 648-655, 2024-The purpose of this study was to examine acute stimulus and fatigue responses to combined accentuated eccentric loading and rest redistribution (AEL + RR). Resistance-trained men ( n = 12, 25.6 ± 4.4 years, 1.77 ± 0.06 m, and 81.7 ± 11.4 kg) completed a back squat (BS) 1 repetition maximum (1RM) and weight releaser familiarization session. Three BS exercise conditions (sets × repetitions × eccentric-concentric loading) consisted of (a) 3 × (5 × 2) × 110/60% (AEL + RR 5), (b) 3 × (2 × 5) × 110/60% (AEL + RR 2), and (c) 3 × 10 × 60/60% 1RM (traditional sets [TS]). Weight releasers (50% 1RM) were attached to every first repetition of each cluster set (every first, third, fifth, seventh, and ninth repetition in AEL + RR 5 and every first and sixth repetition in AEL + RR 2). The AEL + RR 5 resulted in greater total volume load (sets × repetitions × eccentric + concentric loading) (6,630 ± 1,210 kg) when compared with AEL + RR 2 (5,944 ± 1,085 kg) and TS (5,487 ± 1,002 kg). In addition, AEL + RR 5 led to significantly ( p < 0.05) greater rating of perceived exertion (RPE) after set 2 and set 3 and lower blood lactate (BL) after set 3 and 5, 15, and 25 minutes postexercise than AEL + RR 2 and TS. There was a main effect of condition for BL between AEL + RR 5 (5.11 ± 2.90 mmol·L -1 ), AEL + RR 2 (6.23 ± 3.22 mmol·L -1 ), and TS (6.15 ± 3.17 mmol·L -1 ). In summary, AEL + RR 5 results in unique stimulus and fatigue responses. Although it may increase perceived exertion, coaches could use AEL + RR 5 to achieve greater back squat total volume load while reducing BL accumulation.
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Músculo Esquelético , Treinamento Resistido , Masculino , Humanos , Músculo Esquelético/fisiologia , Treinamento Resistido/métodos , Exercício Físico/fisiologia , Terapia por Exercício , Descanso/fisiologia , Força Muscular/fisiologiaRESUMO
Dietary exposure to aflatoxin B1 (AFB1) is a recognized risk factor for developing hepatocellular carcinoma. The mutational signature of AFB1 is characterized by high-frequency base substitutions, predominantly G>T transversions, in a limited subset of trinucleotide sequences. The 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl-formamido)-9-hydroxyaflatoxin B1 (AFB1-FapyGua) has been implicated as the primary DNA lesion responsible for AFB1-induced mutations. This study evaluated the mutagenic potential of AFB1-FapyGua in four sequence contexts, including hot- and cold-spot sequences as apparent in the mutational signature. Vectors containing site-specific AFB1-FapyGua lesions were replicated in primate cells and the products of replication were isolated and sequenced. Consistent with the role of AFB1-FapyGua in AFB1-induced mutagenesis, AFB1-FapyGua was highly mutagenic in all four sequence contexts, causing G>T transversions and other base substitutions at frequencies of ~80%-90%. These data suggest that the unique mutational signature of AFB1 is not explained by sequence-dependent fidelity of replication past AFB1-FapyGua lesions.
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Neoplasias Hepáticas , Mutagênicos , Animais , Mutagênicos/toxicidade , Aflatoxina B1/toxicidade , Adutos de DNA/genética , Guanina , Mutagênese , Neoplasias Hepáticas/patologia , Imidazóis/efeitos adversosRESUMO
Status epilepticus (SE) is a life-threatening development of self-sustaining seizures that becomes resistant to benzodiazepines when treatment is delayed. Benzodiazepine pharmacoresistance is thought in part to result from internalization of synaptic GABAA receptors, which are the main target of the drug. The naturally occurring neurosteroid allopregnanolone is a therapy of interest against SE for its ability to modulate all isoforms of GABAA receptors. Ketamine, an N-methyl-D-aspartate (NMDA) receptor antagonist, has been partially effective in combination with benzodiazepines in mitigating SE-associated neurotoxicity. In this study, allopregnanolone as an adjunct to midazolam or midazolam-ketamine combination therapy was evaluated for efficacy against cholinergic-induced SE. Adult male rats implanted with electroencephalographic (EEG) telemetry devices were exposed to the organophosphorus chemical (OP) soman (GD) and treated with an admix of atropine sulfate and HI-6 at 1 minute after exposure followed by midazolam, midazolam-allopregnanolone, or midazolam-ketamine-allopregnanolone 40 minutes after seizure onset. Neurodegeneration, neuronal loss, and neuroinflammation were assessed 2 weeks after GD exposure. Seizure activity, EEG power integral, and epileptogenesis were also compared among groups. Overall, midazolam-ketamine-allopregnanolone combination therapy was effective in reducing cholinergic-induced toxic signs and neuropathology, particularly in the thalamus and hippocampus. Higher dosage of allopregnanolone administered in combination with midazolam and ketamine was also effective in reducing EEG power integral and epileptogenesis. The current study reports that there is a promising potential of neurosteroids in combination with benzodiazepine and ketamine treatments in a GD model of SE. SIGNIFICANCE STATEMENT: Allopregnanolone, a naturally occurring neurosteroid, reduced pathologies associated with soman (GD) exposure such as epileptogenesis, neurodegeneration, and neuroinflammation, and suppressed GD-induced toxic signs when used as an adjunct to midazolam and ketamine in a delayed treatment model of soman-induced status epilepticus (SE) in rats. However, protection was incomplete, suggesting that further studies are needed to identify optimal combinations of antiseizure medications and routes of administration for maximal efficacy against cholinergic-induced SE.
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Ketamina , Neuroesteroides , Soman , Estado Epiléptico , Ratos , Masculino , Animais , Midazolam/farmacologia , Midazolam/uso terapêutico , Ketamina/farmacologia , Ketamina/uso terapêutico , Pregnanolona/efeitos adversos , Soman/toxicidade , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Doenças Neuroinflamatórias , Neuroesteroides/uso terapêutico , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/tratamento farmacológico , Convulsões/tratamento farmacológico , Benzodiazepinas , Colinérgicos/efeitos adversos , Receptores de GABA-A , Ácido gama-AminobutíricoRESUMO
Pathogenic variants in the human Factor VIII (F8) gene cause Hemophilia A (HA). Here, we investigated the impact of 97 HA-causing single-nucleotide variants on the splicing of 11 exons from F8. For the majority of F8 exons, splicing was insensitive to the presence of HA-causing variants. However, splicing of several exons, including exon-16, was impacted by variants predicted to alter exonic splicing regulatory sequences. Using exon-16 as a model, we investigated the structure-function relationship of HA-causing variants on splicing. Intriguingly, RNA chemical probing analyses revealed a three-way junction structure at the 3'-end of intron-15 (TWJ-3-15) capable of sequestering the polypyrimidine tract. We discovered antisense oligonucleotides (ASOs) targeting TWJ-3-15 partially rescue splicing-deficient exon-16 variants by increasing accessibility of the polypyrimidine tract. The apical stem loop region of TWJ-3-15 also contains two hnRNPA1-dependent intronic splicing silencers (ISSs). ASOs blocking these ISSs also partially rescued splicing. When used in combination, ASOs targeting both the ISSs and the region sequestering the polypyrimidine tract, fully rescue pre-mRNA splicing of multiple HA-linked variants of exon-16. Together, our data reveal a putative RNA structure that sensitizes F8 exon-16 to aberrant splicing.
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Fator VIII , Íntrons , Splicing de RNA , Humanos , Processamento Alternativo , Éxons , Fator VIII/genética , RNA , Precursores de RNARESUMO
Mitoxantrone (1,4-dihydroxy-5,8-bis[2-(2-hydroxyethylamino)ethylamino]-anthracene-9,10-dione) is a clinically-relevant synthetic anthracenedione that functions as a topoisomerase II poison by trapping DNA double-strand break intermediates. Mitoxantrone binds to DNA via both stacking interactions with DNA bases and hydrogen bonding with the sugar-phosphate backbone. It has been shown that mitoxantrone inhibits apurinic/apyrimidinic (AP) endonuclease 1 (APE1)-catalyzed incision of DNA containing a tetrahydrofuran (THF) moiety and more recently, that mitoxantrone forms Schiff base conjugates at AP sites in DNA. In this study, mitoxantrone-mediated inhibition of APE1 at THF sites was shown to be consistent with preferential binding to, and thermal stabilization of DNA containing a THF site as compared to non-damaged DNA. Investigations into the properties of mitoxantrone at AP and 3' α,ß-unsaturated aldehyde sites demonstrated that in addition to being a potent inhibitor of APE1 at these biologically-relevant substrates (â¼ 0.5 µM IC50 on AP site-containing DNA), mitoxantrone also incised AP site-containing DNA by catalyzing ß- and ß/δ-elimination reactions. The efficiency of these reactions to generate the 3' α,ß-unsaturated aldehyde and 3' phosphate products was modulated by DNA structure. Although these cell-free reactions revealed that mitoxantrone can generate 3' phosphates, cells lacking polynucleotide kinase phosphatase did not show increased sensitivity to mitoxantrone treatment. Consistent with its ability to inhibit APE1 activity on DNAs containing either an AP site or a 3' α,ß-unsaturated aldehyde, combined exposures to clinically-relevant concentrations of mitoxantrone and a small molecule APE1 inhibitor revealed additive cytotoxicity. These data suggest that in a cellular context, mitoxantrone may interfere with APE1 DNA repair functions.
Assuntos
DNA , Mitoxantrona , Mitoxantrona/farmacologia , DNA/metabolismo , Reparo do DNA , Aldeídos , Fosfatos , Endonucleases/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismoRESUMO
Benzodiazepine pharmacoresistance develops when treatment of status epilepticus (SE) is delayed. This response may result from gamma-aminobutyric acid A receptors (GABAAR) internalization that follows prolonged SE; this receptor trafficking results in fewer GABAAR in the synapse to restore inhibition. Increase in synaptic N-methyl-D-aspartate receptors (NMDAR) also occurs in rodent models of SE. Lacosamide, a third-generation antiseizure medication (ASM), acts on the slow inactivation of voltage-gated sodium channels. Another ASM, rufinamide, similarly acts on sodium channels by extending the duration of time spent in the inactivation stage. Combination therapy of the benzodiazepine midazolam, NMDAR antagonist ketamine, and ASMs lacosamide (or rufinamide) was investigated for efficacy against soman (GD)-induced SE and neuropathology. Adult male rats implanted with telemetry transmitters for monitoring electroencephalographic (EEG) activity were exposed to a seizure-inducing dose of GD and treated with an admix of atropine sulfate and HI-6 1 minute later and with midazolam monotherapy or combination therapy 40 minutes after EEG seizure onset. Rats were monitored continuously for seizure activity for two weeks, after which brains were processed for assessment of neurodegeneration, neuronal loss, and neuroinflammatory responses. Simultaneous administration of midazolam, ketamine, and lacosamide (or rufinamide) was more protective against GD-induced SE compared with midazolam monotherapy. In general, lacosamide triple therapy had more positive outcomes on measures of epileptogenesis, EEG power integral, and the number of brain regions protected from neuropathology compared with rats treated with rufinamide triple therapy. Overall, both drugs were well tolerated in these combination models. SIGNIFICANCE STATEMENT: We currently report on improved efficacy of antiseizure medications lacosamide and rufinamide, each administered in combination with ketamine (NMDAR antagonist) and midazolam (benzodiazepine), in combatting soman (GD)-induced seizure, epileptogenesis, and brain pathology over that provided by midazolam monotherapy, or dual therapy of midazolam and lacosamide (or rufinamide) in rats. Administration of lacosamide as adjunct to midazolam and ketamine was particularly effective against GD-induced toxicity. However, protection was incomplete, suggesting the need for further study.