Digital Pathology is a Fast and Effective Platform for Providing Head and Neck Pathology Consultations.

Surgical pathology of the head and neck is one of the more challenging areas in all of diagnostic pathology. Its unparalleled diversity and complexity renders it highly vulnerable to diagnostic error compelling unconstrained access to specialized diagnostic expertise. Digital pathology (DP) is a state-of-the-art tool that could facilitate access to specialized expertise, but it is relatively untested in the context of pathology consultations. In a collaboration between Labcorp Dianon and a large academic hospital with subspecialized surgical pathology, DP was implemented to provide the pathology community access to head and neck pathology expertise. From this collaborative experience, glass slides from consecutive consult cases that had been previously diagnosed using DP were reviewed by an expert consultant in a blinded manner following an extended wash-out period. The intraobserver discrepancy rate was recorded. Major discrepancies were defined as those resulting in significant impact on clinical management and/or prognosis, whereas minor discrepancies were those with no impact on care or prognosis. Slides from 57 cases were available for review. The average wash-out period was 19 months. Five discrepancies were recorded (intraobserver concordance rate of 91%). All discrepancies were minor (major discrepancy rate, 0%; minor discrepancy rate, 9%). On appraisal of the discrepant cases, discordant diagnoses were attributed to subjective differences in interpretation rather than objective differences related to the inferiority of DP. DP decreased the median turnaround time by 97% (from 70 h 26 min to 2 h 25 min). DP provides efficient and fast access to expert consultants. The speed of case delivery does not compromise diagnostic precision. Discrepancies are uncommon, minor, and reflect subjective interpretative differences inherent to difficult and ambiguous head and neck cases, and not the inferiority of DP as a diagnostic platform. High concordance can be achieved even for those difficult and complex cases that are concentrated in the consultation practice. This observation carries profound implications regarding universal health care access to specialized diagnostic expertise.

Histidine-assisted reduction of arylnitrenes upon photo-activation of phenyl azide chromophores in GFP-like fluorescent proteins.

The photochemically active sites of the proteins sfGFP66azF and Venus66azF, members of the green fluorescent protein (GFP) family, contain a non-canonical amino acid residue p-azidophenylalanine (azF) instead of Tyr66. The light-induced decomposition of azF at these sites leads to the formation of reactive arylnitrene (nF) intermediates followed by the formation of phenylamine-containing chromophores. We report the first study of the reaction mechanism of the reduction of the arylnitrene intermediates in sfGFP66nF and Venus66nF using molecular modeling methods. The Gibbs energy profiles for the elementary steps of the chemical reaction in sfGFP66nF are computed using molecular dynamics simulations with quantum mechanics/molecular mechanics (QM/MM) potentials. Structures and energies along the reaction pathway in Venus66nF are evaluated using a QM/MM approach. According to the results of the simulations, arylnitrene reduction is coupled with oxidation of the histidine side chain on the His148 residue located near the chromophore.

Histology and clinical correlations in autoimmune hepatitis, primary biliary cholangitis, and autoimmune hepatitis-primary biliary cholangitis overlap syndrome.

OBJECTIVES: The diagnosis of autoimmune hepatitis (AIH), primary biliary cholangitis (PBC), and AIH-PBC overlap syndrome (OS) relies on their histologic features and clinical findings. In this study, we aimed to identify specific morphologic features of these diseases and evaluate their clinical correlation. METHODS: We included initial biopsies from untreated patients with AIH (n = 14), PBC (n = 10), and OS (n = 7). Histologic features of the portal tract, portal-lobular interface, and hepatic lobule, fibrosis, as well as clinical data including serology, autoantibodies, treatment, and prognosis were reviewed and analyzed. RESULTS: Our results showed that several histologic features differed significantly between AIH and PBC (p < 0.05). Among these features, OS cases were more likely to present with bile duct-centered processes (presence of bile duct damage while absence of inflammation gradient from bile duct to interface, plasma cell cluster and pericentral inflammation) unlike those seen in AIH (p < 0.05), and interface-centered processes (unequivocal interface hepatitis, ductular reaction, and periportal fibrosis) which were not seen in PBC (p < 0.05). We observed a significant correlation between transaminase levels and lobular inflammation, including numbers of lymphocyte, plasma cell and eosinophil. Our study also found that anti-smooth muscle antibody positivity was associated with interface hepatitis (p < 0.01), while antimitochondrial antibody positivity was associated with duct damage (including ductopenia) and granulomas (p < 0.05). CONCLUSION: Our results highlight distinctive morphological features between AIH and PBC. The possibility of overlap syndrome should be considered when encountering AIH with bile duct-centered processes or PBC with interface-centered processes in morphology and correlation with autoantibodies.

Multiscale Simulations of the Covalent Inhibition of the SARS-CoV-2 Main Protease: Four Compounds and Three Reaction Mechanisms.

We report the results of computational modeling of the reactions of the SARS-CoV-2 main protease (MPro) with four potential covalent inhibitors. Two of them, carmofur and nirmatrelvir, have shown experimentally the ability to inhibit MPro. Two other compounds, X77A and X77C, were designed computationally in this work. They were derived from the structure of X77, a non-covalent inhibitor forming a tight surface complex with MPro. We modified the X77 structure by introducing warheads capable of reacting with the catalytic cysteine residue in the MPro active site. The reaction mechanisms of the four molecules with MPro were investigated by quantum mechanics/molecular mechanics (QM/MM) simulations. The results show that all four compounds form covalent adducts with the catalytic cysteine Cys 145 of MPro. From the chemical perspective, the reactions of these four molecules with MPro follow three distinct mechanisms. The reactions are initiated by a nucleophilic attack of the thiolate group of the deprotonated cysteine residue from the catalytic dyad Cys145-His41 of MPro. In the case of carmofur and X77A, the covalent binding of the thiolate to the ligand is accompanied by the formation of the fluoro-uracil leaving group. The reaction with X77C follows the nucleophilic aromatic substitution SNAr mechanism. The reaction of MPro with nirmatrelvir (which has a reactive nitrile group) leads to the formation of a covalent thioimidate adduct with the thiolate of the Cys145 residue in the enzyme active site. Our results contribute to the ongoing search for efficient inhibitors of the SARS-CoV-2 enzymes.

Modeling Light-Induced Chromophore Hydration in the Reversibly Photoswitchable Fluorescent Protein Dreiklang.

We report the results of a computational study of the mechanism of the light-induced chemical reaction of chromophore hydration in the fluorescent protein Dreiklang, responsible for its switching from the fluorescent ON-state to the dark OFF-state. We explore the relief of the charge-transfer excited-state potential energy surface in the ON-state to locate minimum energy conical intersection points with the ground-state energy surface. Simulations of the further evolution of model systems allow us to characterize the ground-state reaction intermediate tentatively suggested in the femtosecond studies of the light-induced dynamics in Dreiklang and finally to arrive at the reaction product. The obtained results clarify the details of the photoswitching mechanism in Dreiklang, which is governed by the chemical modification of its chromophore.

Quantum-based Modeling of Protein-ligand Interaction: The Complex of RutA with Uracil and Molecular Oxygen.

Modern quantum-based methods are employed to model interaction of the flavin-dependent enzyme RutA with the uracil and oxygen molecules. This complex presents the structure of reactants for the chain of chemical reactions of monooxygenation in the enzyme active site, which is important in drug metabolism. In this case, application of quantum-based approaches is an essential issue, unlike conventional modeling of protein-ligand interaction with force fields using molecular mechanics and classical molecular dynamics methods. We focus on two difficult problems to characterize the structure of reactants in the RutA-FMN-O2 -uracil complex, where FMN stands for the flavin mononucleotide species. First, location of a small O2 molecule in the triplet spin state in the protein cavities is required. Second, positions of both ligands, O2 and uracil, must be specified in the active site with a comparable accuracy. We show that the methods of molecular dynamics with the interaction potentials of quantum mechanics/molecular mechanics theory (QM/MM MD) allow us to characterize this complex and, in addition, to surmise possible reaction mechanism of uracil oxygenation by RutA.

"I Refuse to Wear a Mask!" Examining the Relationship Between Grit and Preventative Behaviors and Well-Being: Mediating Role of Motivation.

Despite government mandates to wear face masks in some public locations, why do some people still refuse to wear a face mask in public? This paper examines the relationship between grit, COVID-19 preventative health measure, and distress. Further, the mediating role of COVID-19 autonomous and controlled motivation is also investigated. The results suggest that grit is directly related to decreased distress and increased preventative behaviors. COVID-19 autonomous and controlled motivated mediated the relationship between grit, distress and preventive behaviors, such that COVID-19 autonomous motivation mediated the association between grit and preventive behaviors; while COVID-19 controlled motivation mediated the relationship between grit and psychological distress. These findings suggest that grit can be an important growth mindset in increasing preventative behaviors and individual well-being during the pandemic. Implications for human resource managers in attempting to help employees cope effectively during the pandemic are discussed.

Esofagotomía y cierre primario por cuerpo extraño en esófago superior / Esophagotomy and primary closure due to foreign body in the upper esophagus

RESUMEN Los cuerpos extraños en el esófago son un problema frecuente en urgencias, debido la mayoría de las veces a ingestión accidental. Presentamos el caso de una paciente femenina de 50 años, sin antecedentes patológicos, que consulta por presentar la ingesta de prótesis dental mientras almorzaba, con posterior disfagia, odinofagia y dolor cervical. Se le realizan radiografías y tomografía computarizada (TC) donde se evidencia imagen radiopaca en esófago cervical. Se interconsulta con el Servicio de Gastroenterología, dada la imposibilidad de extracción de la prótesis por vía endoscópica. Se realiza luego cervicotomía lateral izquierda de emergencia, con esofagotomía, extracción del cuerpo extraño y cierre primario del esófago. La obstrucción del esófago por cuerpos extraños puede ocasionar muchas complicaciones. La consecuente demora en el tratamiento puede llevar a un marcado aumento de la mortalidad. Por eso, en este caso, se prioriza la cirugía ante un riesgo elevado de perforación y hemorragia.

ABSTRACT Esophageal foreign bodies are a common problem in the emergency department, usually due to accidental ingestion. We report the case of a 50-year-old otherwise healthy female patient who sought medical care due to dysphagia, odynophagia and cervical pain after swallowing a partial denture while having lunch. The patient underwent X-rays and a computed tomography (CT) scan which showed a radiopaque image in the upper esophagus. After a failed attempt to remove the denture through endoscopy, surgery was decided via a left lateral emergency cervicotomy with esophagotomy, foreign body removal and primary closure of the esophagus. Esophageal obstruction due to foreign bodies may cause many complications. Treatment delays may result in increased mortality. Thus, in this case we prioritized surgery due to high risk of perforation and bleeding.

How Reproducible Are QM/MM Simulations? Lessons from Computational Studies of the Covalent Inhibition of the SARS-CoV-2 Main Protease by Carmofur.

This work explores the level of transparency in reporting the details of computational protocols that is required for practical reproducibility of quantum mechanics/molecular mechanics (QM/MM) simulations. Using the reaction of an essential SARS-CoV-2 enzyme (the main protease) with a covalent inhibitor (carmofur) as a test case of chemical reactions in biomolecules, we carried out QM/MM calculations to determine the structures and energies of the reactants, the product, and the transition state/intermediate using analogous QM/MM models implemented in two software packages, NWChem and Q-Chem. Our main benchmarking goal was to reproduce the key energetics computed with the two packages. Our results indicate that quantitative agreement (within the numerical thresholds used in calculations) is difficult to achieve. We show that rather minor details of QM/MM simulations must be reported in order to ensure the reproducibility of the results and offer suggestions toward developing practical guidelines for reporting the results of biosimulations.

[Ocular abnormalities of children born prematurely at the Yaoundé Gynaeco-Obstetrics And Pediatric Hospital]. / Anomalies oculaires des enfants nés prématurément à l'Hôpital gynéco-obstétrique et pédiatrique de Yaoundé.

PURPOSE: To identify ocular abnormalities in premature children in order to treat reversible visual disorders and help prevent low vision and blindness in this population at risk. METHODOLOGY: This was a cross-sectional, analytical study carried out from November 1, 2018 to July 31, 2019 at the Yaoundé Gyneco-Obstetric And Pediatric Hospital, including premature children, aged 3 to 15 years. The variables studied were age, sex, prenatal, birth and past ophthalmological history, visual acuity, oculomotor examination and fundus examination. For statistical analysis, we used the epi-info software 3.5.4, the Chi2 test, odds ratio and a 95% confidence interval with a significance P<0.05. RESULTS: Of the 50 patients examined, the mean age was 6.02 years±2.58. Of the 22 optically corrected patients, hyperopia and hyperopic astigmatism were predominant (70.5%) (n=31). Distance visual acuity, measured in 31 cooperative patients (62 eyes), was between 3/10 and 8/10 in 14 eyes (22.6%), and<3/10 in two eyes (3.2%). Strabismus was present in 21 patients (42%), of which 13 cases were esotropia (61.9%). Tropical endemic limbo-conjunctivitis was found in eight eyes (8%). There were no cases of retinopathy of prematurity. Very low birth weight (<1500g) and neonatal resuscitation were associated with strabismus. CONCLUSION: Ocular abnormalities in preterm infants are dominated by strabismus, which is associated with very low birth weight and neonatal resuscitation.

Examining the relationship between fear of COVID-19, intolerance for uncertainty, and cyberloafing: A mediational model.

After the COVID-19 pandemic began, organizations had to pivot and move to online remote work. As companies moved to digital platforms and technologies for remote working, a key concern was the increase in workplace withdrawal behaviors during the pandemic, including cyberloafing, a form of workplace deviance. Cyberloafing can be described as the action of using the internet for non-work-related activities or personal use during working hours. Given its effect on organizational effectiveness and efficiency, organizations must take measures to minimize cyberloafing. We examined how two factors-fear of COVID-19 and intolerance for uncertainty-were related to cyberloafing during the third lockdown in Israel. A sample of 322 adults who were enrolled in professional courses at a university in Israel were surveyed. Based on Conservation of Resources Theory, our findings suggest that distress significantly mediated the relationship between fear of COVID-19, intolerance for uncertainty, and cyberloafing. In an attempt to deal with the stress and depletion of personal resources during the COVID-19 lockdown, individuals engaged in cyberloafing as a way to handle the stress. Our results suggest that organizations should take measures to reduce fear and uncertainty in order to decrease distress, which, in turn, will reduce cyberloafing.

Protonation States of Molecular Groups in the Chromophore-Binding Site Modulate Properties of the Reversibly Switchable Fluorescent Protein rsEGFP2.

The role of protonation states of the chromophore and its neighboring amino acid side chains of the reversibly switching fluorescent protein rsEGFP2 upon photoswitching is characterized by molecular modeling methods. Numerous conformations of the chromophore-binding site in computationally derived model systems are obtained using the quantum chemistry and QM/MM approaches. Excitation energies are computed using the extended multiconfigurational quasidegenerate perturbation theory (XMCQDPT2). The obtained structures and absorption spectra allow us to provide an interpretation of the observed structural and spectral properties of rsEGFP2 in the active ON and inactive OFF states. The results demonstrate that in addition to the dominating anionic and neutral forms of the chromophore, the cationic and zwitterionic forms may participate in the photoswitching of rsEGFP2. Conformations and protonation forms of the Glu223 and His149 side chains in the chromophore-binding site play an essential role in stabilizing specific protonation forms of the chromophore.

Stalling chromophore synthesis of the fluorescent protein Venus reveals the molecular basis of the final oxidation step.

Fluorescent proteins (FPs) have revolutionised the life sciences, but the mechanism of chromophore maturation is still not fully understood. Here we show that incorporation of a photo-responsive non-canonical amino acid within the chromophore stalls maturation of Venus, a yellow FP, at an intermediate stage; a crystal structure indicates the presence of O2 located above a dehydrated enolate form of the imidazolone ring, close to the strictly conserved Gly67 that occupies a twisted conformation. His148 adopts an "open" conformation so forming a channel that allows O2 access to the immature chromophore. Absorbance spectroscopy supported by QM/MM simulations suggests that the first oxidation step involves formation of a hydroperoxyl intermediate in conjunction with dehydrogenation of the methylene bridge. A fully conjugated mature chromophore is formed through release of H2O2, both in vitro and in vivo. The possibility of interrupting and photochemically restarting chromophore maturation and the mechanistic insights open up new approaches for engineering optically controlled fluorescent proteins.

Modeling photophysical properties of the bacteriophytochrome-based fluorescent protein IFP1.4.

An enhanced interest in the phytochrome-based fluorescent proteins is explained by their ability to absorb and emit light in the far-red and infra-red regions particularly suitable for bioimaging. The fluorescent protein IFP1.4 was engineered from the chromophore-binding domain of a bacteriophytochrome in attempts to increase the fluorescence quantum yield. We report the results of simulations of structures in the ground S0 and excited S1 electronic states of IFP1.4 using the methods of quantum chemistry and quantum mechanics/molecular mechanics. We construct different protonation states of the biliverdin (BV) chromophore in the red-absorbing form of the protein by moving protons from the BV pyrrole rings to a suitable acceptor within the system and show that these structures are close in energy but differ by absorption bands. For the first time, we report structures of the minimum energy conical intersection points S1/S0 on the energy surfaces of BV in the protein environment and describe their connection to the local minima in the excited S1 state. These simulations allow us to characterize the deactivation routes in IFP1.4.

Interplay between Locally Excited and Charge Transfer States Governs the Photoswitching Mechanism in the Fluorescent Protein Dreiklang.

We present the results of high-level electronic structure and dynamics simulations of the photoactive protein Dreiklang. With the goal of understanding the details of the Dreiklang photocycle, we carefully characterize the excited states of the ON- and OFF-forms of Dreiklang. The key finding of our study is the existence of a low-lying excited state of a charge-transfer character in the neutral ON form and that population of this state, which is nearly isoenergetic with the locally excited bright state, initiates a series of steps that ultimately lead to the formation of the hydrated dark chromophore (OFF state). These results allow us to refine the mechanistic picture of Dreiklang's photocycle and photoactivation.

CYP450 polymorphisms and clinical pharmacogenetics of ibuprofen after lower third molar extraction.

PURPOSE: This study hypothesized that drugs accumulate in the bloodstream of poor-metabolizing patients and may have more adverse effects and different pain perceptions and aimed to investigate the influence of CYP450 polymorphisms on acute postoperative pain, swelling, and trismus controlled by ibuprofen (600 mg) in 200 volunteers after dental extraction. In addition, surgical outcomes can determine pain, edema, and trismus and indicate inflammatory reactions after oral surgeries. METHODS: Genetic sequencing was performed to identify CYP450 polymorphisms and the surgical parameters evaluated: pre and postoperative swelling, trismus, and temperature; self-reported postoperative pain with visual analog scale (VAS); rescue medication consumed; and severity of adverse reactions. RESULTS: A multiple linear regression model with independent variables [single nucleotide polymorphisms (SNPs), BMI (body mass index), duration, and difficulty of surgery] and dependent variables [postoperative pain by sum of pain intensity difference (SPID), trismus, and swelling] was used for analysis. The duration of surgery was a predictor for pain at 8 h and 96 h after surgery, and BMI was a predictor for both swelling and trismus on the 2nd postoperative day. When evaluating CYP2C8 and C9 genotyped SNPs, it was observed that normal metabolizers showed higher pain levels than the intermediate/poor metabolizers on the postoperative periods as compared with time 0 h. In another analysis, the poor metabolizers for CYP2C8 and C9 presented lower levels of postoperative pain after 8 h and used rescue medication earlier than normal metabolizers. CONCLUSION: Ibuprofen 600 mg was very effective in controlling inflammatory pain after lower third molar surgeries, without relevant adverse reactions; although in a very subtle way, patients with poor metabolism had higher levels of pain in the first hours, and no longer after 8 h, and used pain relief medication earlier. TRIAL REGISTRATION: The study was registered with ClinicalTrials.gov ID (NCT03169127), on March 16th, 2017.

Structure of the Brain N-Acetylaspartate Biosynthetic Enzyme NAT8L Revealed by Computer Modeling.

We report the results of computational modeling of a three-dimensional all-atom structure of the membrane-associated protein encoded by the NAT8L gene, aspartate N-acetyltransferase, which is essential for brain synthesis of N-acetyl-l-aspartate (NAA). The lack of experimentally derived three-dimensional structures of NAT8L poses one of the obstacles in studies of the mechanism of NAA formation and understanding the precise role of NAA in neurological disorders. We apply a computational protocol employing the contact map prediction, ab initio folding, homology modeling, and refinement to obtain a structure of NAT8L with the aspartate and acetyl coenzyme A cofactors in the protein molecule. To verify the computational protocol, we check its predictive power by reproducing the crystal structure of a related N-acetyltransferase domain, specifically, that from the bacterial N-acetylglutamate synthase. We show that the constructed NAT8L model correlates with structural features of the protein revealed in rare experimental studies. The obtained structure of the enzyme active site with the trapped reactants suggests a mechanism of the acetyl transfer upon NAA formation.

AT1 receptor antagonism promotes bone loss attenuation in experimental periodontitis, blocks inflammatory mediators, and upregulates antioxidant enzymes and bone formation markers.

BACKGROUND: The initiation and progression of periodontitis might involve a local renin-angiotensin system in periodontal tissue. This study hypothesized that Losartan treatment could promote protection to rats submitted to experimental periodontitis (EP) by attenuating alveolar bone loss due to reduction in inflammatory cytokines, better reactive oxidant species regulation and maintenance of the balance between bone formation and resorption factors. METHODS: One hundred and thirty rats were submitted to EP with a silk suture thread (4.0) placed around the lower right first molar for 1, 3, 7, and 14 consecutive days. The study comprised four groups: G1-control without EP; G2-animals with EP treated with water; G3-Losartan-treated animals (treatment started at the same day of EP induction), and G4-animals previously treated with Losartan for 30 days followed by induction of EP and continuity of treatment. RESULTS: G2 rats had greater bone loss volume, increased number, and thickness and decreased separation of trabeculae. On the other hand, G4 animals showed significant improvements in these parameters. Histological analysis revealed that EP favors inflammatory cell infiltration and junctional epithelium, cementum with alveolar bone crest destruction, but animals pretreated with Losartan (G4) did not show these features. Although the G3 animals did not demonstrate the improvements detected in G4, mRNA expression results were similar. In mandibular tissue, EP promoted mRNA increases for ACE, AT1 receptor, and inflammatory mediators as well as decreases for antioxidant enzymes. However, Losartan treatments attenuated these responses in addition to promoting an increase in bone formation markers and transcription factors. CONCLUSION: AT1 receptor modulates EP progression.