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The tachycardia which presents with regularly irregular rhythm consists of a broad set of differential diagnoses. We present a case of cycle length alternans tachycardia in a patient, with Ebstein's anomaly and describe how a diagnosis was arrived at after careful analysis of electrocardiogram and EGMs.
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A 15-year-old boy with manifest preexcitation and recurrent palpitations had undergone an unsuccessful ablation procedure elsewhere and was subsequently referred to us. The ECG suggested a left free wall pathway but there was a pattern break in lead V2. This helped localise the accessory pathway to the summit region and achieve success.
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Electron-driven process helps the living organism in the generations of energy, biomass production and detoxification of synthetic compounds. Soluble quinone oxidoreductases (QORs) mediate the transfer of an electron from NADPH to various quinone and other compounds, helping in the detoxification of quinones. QORs play a crucial role in cellular metabolism and are thus potential targets for drug development. Here we report the crystal structure of the NADPH-dependent QOR from Leishmania donovani (LdQOR) at 2.05 Å. The enzyme exists as a homo-dimer, with each protomer consisting of two domains, responsible for binding NADPH cofactor and the substrate. Interestingly, the human QOR exists as a tetramer. Comparative analysis of the oligomeric interfaces of LdQOR with HsQOR shows no significant differences in the protomer/dimer assembly. The tetrameric interface of HsQOR is stabilized by salt bridges formed between Arg 169 and Glu 271 which is non-existent in LdQOR, with an Alanine replacing the glutamate. This distinct feature is conserved across other dimeric QORs, indicating the importance of this interaction for tetramer association. Among the homologs, the sequences of the loop region involved in the stabilization and binding of the adenine ring of the NADPH shows significant differences except for an Arginine & glycine residues. In dimer QORs, this Arginine acts as a gate to the co-factor, while the NADPH binding mode in the human homolog is distinct, stabilized by His 200 and Asn 229, which are not conserved in LdQOR. These distinct features have the potential to be utilized for therapeutic interventions.
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NAD(P)H Deshidrogenasa (Quinona) , Quinona Reductasas , Humanos , NADP/metabolismo , Subunidades de Proteína , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Quinona Reductasas/química , Quinona Reductasas/metabolismo , Quinonas , Arginina , Sitios de Unión , Cristalografía por Rayos XRESUMEN
Leishmaniasis is caused by â¼20 species of Leishmania that affects millions in endemic areas. Available therapies are not sufficient to effectively control the disease, cause severe side effects and eventually lead to drug resistance, making the discovery of novel therapeutic molecules an immediate need. Molecular target-based drug discovery, where the target is a defined molecular gene, protein or a mechanism, is a rationale driven approach for novel therapeutics. Humans obtain the essential amino acid such as threonine from dietary sources, while Leishmania synthesize it de-novo. Enzymes of the threonine biosynthesis pathway, including the rate limiting Homoserine kinase (HSK) which converts L-homoserine into ortho-phospho homoserine are thus attractive targets for rationale driven therapy. The absence of HSK in humans and its presence in Leishmania donovani enhances the opportunity to exploit HSK as a molecular target for anti-leishmanials therapeutic development. In this study, we utilize structure-based high throughput drug discovery (SBDD), followed by biochemical validation and identified two potential inhibitors (RH00038 and S02587) from Maybridge chemical library that targets L. donovani HSK. These two inhibitors effectively induced the mortality of Leishmania donovani in both amastigote and promastigote stages, with one of them being specific to parasite and twice as effective as the standard therapeutic molecule.
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The neglected tropical disease (NTD) leishmaniasis is the collective name given to a diverse group of illnesses caused by ~20 species belonging to the genus Leishmania, a majority of which are vector borne and associated with complex life cycles that cause immense health, social, and economic burdens locally, but individually are not a major global health priority. Therapeutic approaches against leishmaniasis have various inadequacies including drug resistance and a lack of effective control and eradication of the disease spread. Therefore, the development of a rationale-driven, target based approaches towards novel therapeutics against leishmaniasis is an emergent need. The utilization of Artificial Intelligence/Machine Learning methods, which have made significant advances in drug discovery applications, would benefit the discovery process. In this review, following a summary of the disease epidemiology and available therapies, we consider three important leishmanial metabolic pathways that can be attractive targets for a structure-based drug discovery approach towards the development of novel anti-leishmanials. The folate biosynthesis pathway is critical, as Leishmania is auxotrophic for folates that are essential in many metabolic pathways. Leishmania can not synthesize purines de novo, and salvage them from the host, making the purine salvage pathway an attractive target for novel therapeutics. Leishmania also possesses an organelle glycosome, evolutionarily related to peroxisomes of higher eukaryotes, which is essential for the survival of the parasite. Research towards therapeutics is underway against enzymes from the first two pathways, while the third is as yet unexplored.
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Xanthomonas oryzae pv. oryzae (Xoo) is a destructive pathogen that causes bacterial blight disease of rice worldwide. Xoo uses T3SS (type III secretion system) effectors to subvert rice innate immunity. However, the comprehensive knowledge of rice genes involved in T3SS effectors-mediated interaction remains unclear. In this study, the transcriptome profiles of rice infected with a virulent Xoo strain from North-eastern region of India relatives to its avirulent strain (that lacks functional T3SS) were analyzed at early (2-6 hpi) and late (16-24 hpi) hours of infection. Out of total 255 differentially expressed genes (DEGs), during early infection, 62 and 70 genes were upregulated and downregulated, respectively. At late infection, 70 and 53 genes were upregulated and downregulated, respectively. The transcriptomic data identified many differentially expressed resistant genes, transposons, transcription factors, serine/threonine protein kinase, cytochrome P450 and peroxidase genes that are involved in plant defense. Pathway analysis revealed that these DEGs are involved in hormone signaling, plant defense, cellular metabolism, growth and development processes. DEGs associated with plant defense were also validated through quantitative real-time PCR. Our study brings a comprehensive picture of the rice genes that are being differentially expressed during bacterial blight infection. Nevertheless, the DEG-associated pathways would provide sensible targets for developing resistance to bacterial blight. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03193-4.
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Cadherins play an essential role in the attachment of the blastocyst to the endometrium, a process known as endometrial receptivity. Loss of E-cadherin expression is essential during the process, while the expression level of the other cadherin, N-cadherin, has been reported to be altered in cases of infertility. Both E-cadherin and N-cadherin can be regulated by members of the PARP family. Specifically, PARP-2, which is under the epigenetic control of miR-149, has been observed to promote E-cadherin expression in other human cells. We investigated the roles of E-cadherin and N-cadherin in endometrial receptivity using mouse models for normal endometrial receptivity, pseudopregnancy, and LPS-induced endometrial receptivity failure. E-cadherin and phosphorylated E-cadherin were predominantly expressed during pre-receptive stages as well as in the implantation site of the receptive stage, which were observed reduced during the later stages of implantation in both implantation and non-implantation regions, while N-cadherin was detected only at pre-receptive stages. E-cadherin and N-cadherin were also seen in the uterus during pseudopregnancy, showing a downregulation trend during receptive and post-receptive stages. LPS-induced failed endometrial receptivity showed upregulation of E-cadherin and downregulation of N-cadherin. The E-cadherin expression promoter, GSK-3, was lost and its suppressor, SLUG was upregulated during normal course of endometrial receptivity in mouse model, while GSK-3 was increased during LPS-induced failed embryo implantation. In an in vitro model of embryo implantation, E-cadherin expression is promoted by PARP-2 and regulated by miR-149 epigenetically in human endometrium epithelial cells. In conclusion, E-cadherin is predominantly expressed during pre-receptive stage and promoted by PARP-2, which is regulated by miR-149 in the endometrial epithelial cells.
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Cadherinas/metabolismo , Endometrio/metabolismo , MicroARNs/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Animales , Implantación del Embrión/fisiología , Femenino , Ratones , Embarazo , Transducción de SeñalRESUMEN
The two-domain actin associated protein coronin interacts with filamentous (F-) actin, facilitating diverse biological processes including cell proliferation, motility, phagocytosis, host-parasite interaction and cargo binding. The conserved N-terminal ß-propeller domain is involved in protein: protein interactions, while the C-terminal coiled-coil domain mediates oligomerization, transducing conformational changes. The L. donovani coronin coiled-coil (LdCoroCC) domain exhibited a novel topology and oligomer association with an inherent asymmetry, caused primarily by three a residues of successive heptads. In the T.brucei homolog (TbrCoro), two of these 'a' residues are different (Val 493 & 507 replacing LdCoroCC Ile 486 and Met 500 respectively). The elucidated structure possesses a similar topology and assembly while comparative structural analysis shows that the T.brucei coronin coiled-coil domain (TbrCoroCC) too possesses the asymmetry though its magnitude is smaller. Analysis identifies that the asymmetric state is stabilized via cyclic salt bridges formed by Arg 497 and Glu 504. Co-localization studies (LdCoro, TbrCoro and corresponding mutant coiled coil constructs) with actin show that there are subtle differences in their binding patterns, with the double mutant V493I-V507M showing maximal effect. None of the constructs have an effect on F-actin length. Taken together with LdCoroCC, we therefore conclude that the inherent asymmetric structures are essential for kinetoplastids, and are of interest in understanding and exploiting actin dynamics.
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BACKGROUND: Day-of-surgery cancellation (DoSC) represents a substantial wastage of hospital resources and can cause significant inconvenience to patients and families. Cancellation is reported to impact between 2% and 20% of the 50 million procedures performed annually in American hospitals. Up to 85% of cancellations may be amenable to the modification of patients' and families' behaviors. However, the factors underlying DoSC and the barriers experienced by families are not well understood. OBJECTIVE: This study aims to conduct a geospatial analysis of patient-specific variables from electronic health records (EHRs) of Cincinnati Children's Hospital Medical Center (CCHMC) and of Texas Children's Hospital (TCH), as well as linked socioeconomic factors measured at the census tract level, to understand potential underlying contributors to disparities in DoSC rates across neighborhoods. METHODS: The study population included pediatric patients who underwent scheduled surgeries at CCHMC and TCH. A 5-year data set was extracted from the CCHMC EHR, and addresses were geocoded. An equivalent set of data >5.7 years was extracted from the TCH EHR. Case-based data related to patients' health care use were aggregated at the census tract level. Community-level variables were extracted from the American Community Survey as surrogates for patients' socioeconomic and minority status as well as markers of the surrounding context. Leveraging the selected variables, we built spatial models to understand the variation in DoSC rates across census tracts. The findings were compared to those of the nonspatial regression and deep learning models. Model performance was evaluated from the root mean squared error (RMSE) using nested 10-fold cross-validation. Feature importance was evaluated by computing the increment of the RMSE when a single variable was shuffled within the data set. RESULTS: Data collection yielded sets of 463 census tracts at CCHMC (DoSC rates 1.2%-12.5%) and 1024 census tracts at TCH (DoSC rates 3%-12.2%). For CCHMC, an L2-normalized generalized linear regression model achieved the best performance in predicting all-cause DoSC rate (RMSE 1.299%, 95% CI 1.21%-1.387%); however, its improvement over others was marginal. For TCH, an L2-normalized generalized linear regression model also performed best (RMSE 1.305%, 95% CI 1.257%-1.352%). All-cause DoSC rate at CCHMC was predicted most strongly by previous no show. As for community-level data, the proportion of African American inhabitants per census tract was consistently an important predictor. In the Texas area, the proportion of overcrowded households was salient to DoSC rate. CONCLUSIONS: Our findings suggest that geospatial analysis offers potential for use in targeting interventions for census tracts at a higher risk of cancellation. Our study also demonstrates the importance of home location, socioeconomic disadvantage, and racial minority status on the DoSC of children's surgery. The success of future efforts to reduce cancellation may benefit from taking social, economic, and cultural issues into account.
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Grupos Minoritarios , Características de la Residencia , Niño , Registros Electrónicos de Salud , Hospitales Pediátricos , Humanos , Factores SocioeconómicosRESUMEN
Nucleoside diphosphate kinases (NDK) are ubiquitous enzymes that catalyse the transfer of the γ phosphate from nucleoside triphosphates (NTPs) to nucleoside diphosphate (NDPs), to maintain appropriate NTP levels in cells. NDKs are associated with signal transduction, cell development, proliferation, differentiation, tumor metastasis, apoptosis and motility. The critical role of NDK in bacterial virulence renders it a potential drug target. The present manuscript reports crystal structure and functional characterization of Vibrio cholerae NDK (VNDK). The 16 kDa VNDK was crystallized in a solution containing 30% PEG 4000, 100 mM Tris-HCl pH 8.5 and 200 mM sodium acetate in orthorhombic space group P212121 with unit cell parameters a = 48.37, b = 71.21, c = 89.14 Å, α = ß = γ = 90° with 2 molecules in asymmetric unit. The crystal structure was solved by molecular replacement and refined to crystallographic Rfactor and Rfree values of 22.8% and 25.8% respectively. VNDK exists as both dimer and tetramer in solution as confirmed by size exclusion chromatography, glutaraldehyde crosslinking and small angle X-ray scattering while the crystal structure appears to be a dimer. The biophysical characterization states that VNDK has kinase and DNase activity with maximum stability at pH 8-9 and temperature up to 40 °C. VNDK shows elevated thermolability as compared to other NDK and shows preferential binding with GTP rationalized using computational studies.
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Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Nucleósido-Difosfato Quinasa/química , Nucleósido-Difosfato Quinasa/metabolismo , Vibrio cholerae/enzimología , Proteínas Bacterianas/aislamiento & purificación , Cristalografía por Rayos X , Desoxirribonucleasas/metabolismo , Estabilidad de Enzimas , Guanosina Trifosfato/metabolismo , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Nucleósido-Difosfato Quinasa/aislamiento & purificación , Multimerización de Proteína , Estructura Cuaternaria de Proteína , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Dispersión del Ángulo Pequeño , Alineación de Secuencia , Análisis de Secuencia de Proteína , Espectrometría de Fluorescencia , Temperatura , Vibrio cholerae/genéticaRESUMEN
The small mitochondrial genome (mtDNA) of the malaria parasite is known to transcribe its genes polycistonically, although promoter element(s) have not yet been identified. An unusually large Plasmodium falciparum candidate mitochondrial phage-like RNA polymerase (PfmtRNAP) with an extended N-terminal region is encoded by the parasite nuclear genome. Using specific antibodies against the enzyme, we established that PfmtRNAP was targeted exclusively to the mitochondrion and interacted with mtDNA. Phylogenetic analysis showed that it is part of a separate apicomplexan clade. A search for PfmtRNAP-associated transcription initiation factors using sequence homology and in silico protein-protein interaction network analysis identified PfKsgA1. PfKsgA1 is a dual cytosol- and mitochondrion-targeted protein that functions as a small subunit rRNA dimethyltransferase in ribosome biogenesis. Chromatin immunoprecipitation showed that PfKsgA1 interacts with mtDNA, and in vivo crosslinking and pull-down experiments confirmed PfmtRNAP-PfKsgA1 interaction. The ability of PfKsgA1 to serve as a transcription initiation factor was demonstrated by complementation of yeast mitochondrial transcription factor Mtf1 function in Rpo41-driven in vitro transcription. Pull-down experiments using PfKsgA1 and PfmtRNAP domains indicated that the N-terminal region of PfmtRNAP interacts primarily with the PfKsgA1 C-terminal domain with some contacts being made with the linker and N-terminal domain of PfKsgA1. In the absence of full-length recombinant PfmtRNAP, solution structures of yeast mitochondrial RNA polymerase Rpo41 complexes with Mtf1 or PfKsgA1 were determined by small-angle X-ray scattering. Protein interaction interfaces thus identified matched with those reported earlier for Rpo41-Mtf1 interaction and overlaid with the PfmtRNAP-interfacing region identified experimentally for PfKsgA1. Our results indicate that in addition to a role in mitochondrial ribosome biogenesis, PfKsgA1 has an independent function as a transcription initiation factor for PfmtRNAP.
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Proteínas Mitocondriales , Plasmodium falciparum , ARN Polimerasas Dirigidas por ADN/genética , Mitocondrias/genética , Proteínas Mitocondriales/genética , Factores de Iniciación de Péptidos , Filogenia , Plasmodium falciparum/genética , ARN Mitocondrial , Factores de Transcripción/genéticaRESUMEN
The glycosomal membrane-associated Leishmania donovani protein PEX14, which plays a crucial role in protein import from the cytosol to the glycosomal matrix, consists of three domains: an N-terminal domain where the signalling molecule binds, a transmembrane domain and an 84-residue coiled-coil domain (CC) that is responsible for oligomerization. CCs are versatile domains that participate in a variety of functions including supramolecular assembly, cellular signalling and transport. Recombinant PEX14 CC was cloned, overexpressed, affinity-purified with in-column thrombin cleavage and further purified by size-exclusion chromatography. Crystals that diffracted to 1.98â Å resolution were obtained from a condition consisting of 1.4â M sodium citrate tribasic dihydrate, 0.1â M HEPES buffer pH 7.5. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 143.98, b = 32.62, c = 95.62â Å, ß = 94.68°. Structure determination and characterization are in progress.
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Cristalografía por Rayos X/métodos , Leishmania donovani/metabolismo , Proteínas Protozoarias/química , Proteínas Protozoarias/metabolismo , Clonación Molecular , Unión Proteica , Conformación Proteica , Dominios Proteicos , Proteínas Protozoarias/genética , Proteínas Protozoarias/aislamiento & purificaciónRESUMEN
Purine bases, synthesized de novo or recycled through the salvage pathway, are precursors of nucleotide synthesis and are essential in a variety of physiological processes including cell division, growth, signaling, energy metabolism and synthesis of vitamins/co-factor. The protozoan kinetoplastid parasites including Leishmania cannot synthesize de novo and rely solely on the purine salvage pathway, recycling the degraded products of nucleic acid metabolism. Enzymes of this pathway are thus of therapeutic importance. The enzyme Hypoxanthine-guanine phosphoribosyl transferase (HGPRT) (EC 2.4.2.8) plays a central role in this pathway, converting the purine base to its monophosphate product. Towards the elucidation of its role, we have cloned, expressed, purified and determined the crystal structure of L. donovani HGPRT at 2.76 Å. Comparative structural analysis with the human homolog indicates differences in oligomer association. Comparative analyses identify insertions in the human homolog sequence in the tetramer interface. The results suggest that this difference can be exploited for therapeutic approaches.
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Hipoxantina Fosforribosiltransferasa/química , Leishmania donovani/enzimología , Proteínas Protozoarias/química , Clonación Molecular , Humanos , Hipoxantina Fosforribosiltransferasa/genética , Hipoxantina Fosforribosiltransferasa/aislamiento & purificación , Modelos Moleculares , Proteínas Protozoarias/genética , Proteínas Protozoarias/aislamiento & purificación , Homología Estructural de ProteínaRESUMEN
Protein-protein interactions of cellular importance are mediated by coiled coils (CCs), the ubiquitous structural motif formed by the association of two or more α-helices in a knobs into holes manner. Coronins, actin-associated multi-functional proteins that possess distinct cytoskeleton-dependent and independent functions, oligomerize through their C-terminal CC domain. The structure of the L. donovani coronin CC domain (LdCoroCC; PDB ID 5CX2) revealed, in addition to a novel topology and architecture, an inherent asymmetry, with one of the helices of the 4-helix bundle axially shifted (~2 turns). The structural analysis identified that steric hindrance by Ile 486, Leu 493 and Met 500 as the cause for this asymmetry. To experimentally validate this hypothesis and to better understand the sequence-structure relationship in CCs, these amino acids have been mutated (I486A, L493A, M500V and the double mutant I486A-L493A) and characterized. Thermal CD studies suggest that the I486A and M500V mutants have comparable Tm values to LdCoroCC, while the other mutants have lower melting temperatures. The mutant crystal structures (I486A, M500V and the double mutant) retain the 'ade' core packing as LdcoroCC. While the M500V structure is similar to LdCoroCC, the I486A and the I486A-L493A structures show an asymmetry to symmetry transition. This study reveals crucial role of residues at position 'a' in coiled-coil domain play an important role in stabilizing the asymmetry in LdCoroCC, which might be necessary pursue specific biological function(s) inside the Leishmania.
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Leishmania/metabolismo , Proteínas de Microfilamentos/química , Proteínas de Microfilamentos/metabolismo , Secuencia de Aminoácidos , Animales , Bovinos , Modelos Moleculares , Proteínas Mutantes/química , Dominios Proteicos , Multimerización de Proteína , Estructura Secundaria de ProteínaRESUMEN
BACKGROUND: Last-minute surgery cancellation represents a major wastage of resources and can cause significant inconvenience to patients. Our objectives in this study were: 1) To develop predictive models of last-minute surgery cancellation, utilizing machine learning technologies, from patient-specific and contextual data from two distinct pediatric surgical sites of a single institution; and 2) to identify specific key predictors that impact children's risk of day-of-surgery cancellation. METHODS AND FINDINGS: We extracted five-year datasets (2012-2017) from the Electronic Health Record at Cincinnati Children's Hospital Medical Center. By leveraging patient-specific information and contextual data, machine learning classifiers were developed to predict all patient-related cancellations and the most frequent four cancellation causes individually (patient illness, "no show," NPO violation and refusal to undergo surgery by either patient or family). Model performance was evaluated by the area under the receiver operating characteristic curve (AUC) using ten-fold cross-validation. The best performance for predicting all-cause surgery cancellation was generated by gradient-boosted logistic regression models, with AUC 0.781 (95% CI: [0.764,0.797]) and 0.740 (95% CI: [0.726,0.771]) for the two campuses. Of the four most frequent individual causes of cancellation, "no show" and NPO violation were predicted better than patient illness or patient/family refusal. Models showed good cross-campus generalizability (AUC: 0.725/0.735, when training on one site and testing on the other). To synthesize a human-oriented conceptualization of pediatric surgery cancellation, an iterative step-forward approach was applied to identify key predictors which may inform the design of future preventive interventions. CONCLUSIONS: Our study demonstrated the capacity of machine learning models for predicting pediatric patients at risk of last-minute surgery cancellation and providing useful insight into root causes of cancellation. The approach offers the promise of targeted interventions to significantly decrease both healthcare costs and also families' negative experiences.
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Minería de Datos , Cirugía General/estadística & datos numéricos , Aprendizaje Automático , Niño , Registros Electrónicos de Salud , Humanos , Modelos Logísticos , Curva ROCRESUMEN
Children with trisomy 18 are surviving longer and undergoing more aggressive life-sustaining therapy. This report describes two patients with trisomy 18 and hepatoblastoma (HB) successfully resected in the setting of significant pulmonary hypertension. Forty-four previously published cases of the association between HB and trisomy 18 are reviewed. With careful multidisciplinary preoperative planning, successful resection of HB in children with trisomy 18 who have significant pulmonary hypertension is feasible. Because HB and trisomy 18 are increasing in prevalence, the need for timely liver tumor resection in the setting of pulmonary hypertension will be more common.
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Anestésicos/administración & dosificación , Hepatectomía/métodos , Hepatoblastoma/cirugía , Hipertensión Pulmonar/cirugía , Neoplasias Hepáticas/cirugía , Síndrome de la Trisomía 18/cirugía , Femenino , Hepatoblastoma/complicaciones , Hepatoblastoma/tratamiento farmacológico , Hepatoblastoma/patología , Humanos , Hipertensión Pulmonar/complicaciones , Hipertensión Pulmonar/tratamiento farmacológico , Hipertensión Pulmonar/patología , Lactante , Neoplasias Hepáticas/complicaciones , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/patología , Pronóstico , Síndrome de la Trisomía 18/complicaciones , Síndrome de la Trisomía 18/tratamiento farmacológico , Síndrome de la Trisomía 18/patologíaRESUMEN
OBJECTIVES: To evaluate the effect of implementation of a comfort algorithm on infusion rates of opioids and benzodiazepines in postneonatal postoperative pediatric cardiac surgery patients. DESIGN: A quality improvement project, using statistical process control methodology. SETTING: Twenty-five-bed tertiary care pediatric cardiac ICU in an urban academic Children's hospital. PATIENTS: Postoperative pediatric cardiac surgery patients. INTERVENTIONS: Implementation of a guided comfort medication algorithm which consisted of key components; a low dose opioid continuous infusion, judicious use of frequent as needed opioids, initiation of dexmedetomidine infusion postoperatively, and minimal use of benzodiazepines. MEASUREMENTS AND MAIN RESULTS: Among the baseline group admitted over the 18 month period prior to comfort algorithm implementation, 58 of 116 intubated patients (50%) received a continuous opioid infusion, compared with 30 of 41 (73%) for the implementation group over the 9-month period following implementation. Following algorithm implementation, opioid infusion rates were decreased and benzodiazepine infusions were nearly eliminated. Dexmedetomidine use and infusion rates did not change. Although mean duration of sedative drug infusions did not change with implementation, the frequency of high outliers was diminished. Duration of mechanical ventilation, length of ICU stay (outcome measures), and the frequency of unplanned extubation (balancing measure) were not affected by implementation. CONCLUSIONS: Implementation of a pediatric comfort algorithm reduced opioid and benzodiazepine dosing, without compromising safety for postoperative pediatric cardiac surgical patients.
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Analgésicos Opioides/administración & dosificación , Benzodiazepinas/administración & dosificación , Hipnóticos y Sedantes/administración & dosificación , Unidades de Cuidado Intensivo Pediátrico/organización & administración , Dolor Postoperatorio/tratamiento farmacológico , Centros Médicos Académicos , Extubación Traqueal/estadística & datos numéricos , Algoritmos , Procedimientos Quirúrgicos Cardíacos/métodos , Unidades de Cuidados Coronarios/organización & administración , Cuidados Críticos/organización & administración , Dexmedetomidina/administración & dosificación , Utilización de Medicamentos , Femenino , Humanos , Unidades de Cuidado Intensivo Pediátrico/normas , Tiempo de Internación/estadística & datos numéricos , Masculino , Mejoramiento de la Calidad/organización & administración , Respiración Artificial/estadística & datos numéricosRESUMEN
The availability of complete genome sequence of Mycobacterium tuberculosis has provided an important tool to understand the mycobacterial biology with respect to host-pathogen interaction, which is an unmet need of the hour owing to continuous increasing drug resistance. Hypothetical proteins are often an overlooked pool though half the genome encodes for such proteins of unknown function that could potentially play vital roles in mycobacterial biology. In this context, we report the structural and functional characterization of the hypothetical protein Rv3272. Sequence analysis classifies Rv3272 as a Family III CoA transferase with the classical two domain structure and conserved Aspartate residue (D175). The crystal structure of the wild type protein (2.2â¯Å) demonstrated the associated inter-locked dimer while that of the D175A mutant co-crystallized with octanoyl-CoA demonstrated relative movement between the two domains. Isothermal titration calorimetry studies indicate that Rv3272 binds to fatty acyl-CoAs of varying carbon chain lengths, with palmitoyl-CoA (C16:0) exhibiting maximum affinity. To determine the functional relevance of Rv3272 in mycobacterial biology, we ectopically expressed Rv3272 in M. smegmatis and assessed that its expression encodes significant alteration in cell surface with marked differences in triacylglycerol accumulation. Additionally, Rv3272 expression protects mycobacteria from acidic, oxidative and antibiotic stress under in vitro conditions. Taken together, these studies indicate a significant role for Rv3272 in host-pathogen interaction.
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Proteínas Bacterianas/fisiología , Coenzima A Transferasas/fisiología , Mycobacterium tuberculosis/fisiología , Estrés Fisiológico/fisiología , Acilcoenzima A/química , Antibacterianos/farmacología , Proteínas Bacterianas/química , Pared Celular/química , Coenzima A Transferasas/química , Concentración de Iones de Hidrógeno , Ligandos , Metabolismo de los Lípidos , Simulación del Acoplamiento Molecular , Mycobacterium tuberculosis/efectos de los fármacos , Estrés OxidativoRESUMEN
The perennial wild rice Zizania latifolia is confined in the swampy habitat and wetland of the Indo-Burma biodiversity hotspot of India and infection by the biotrophic fungus Ustilago esculenta is hallmarked by swellings that develop to form localized smut-gall at the topmost internodal region. The cellular and proteomic events involved in the non-systemic colonization of Z. latifolia by U. esculenta leading to smut-gall formation is poorly understood. Proteins were extracted from the smut-gall region at the topmost internodal region below the apical meristematic tissue from the infected and uninfected parts of Z. latifolia. By combining transmission electron microscopy (TEM) and fluorescent microscopy (FM), we showed that U. esculenta hyphal morphological transitions and movement occurred both intercellularly and intracellularly while sporulation occurred intracellularly in selective cells. Following proteome profiling using two dimensional SDS-PAGE at different phenological phases of smut-gall development and U. esculenta infection, differentially expressed proteins bands and their relative abundance were detected and subjected to liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis. Importantly, the fungus explores at least 7 metabolic pathways and 5 major biological processes to subdue the host defense and thrive successfully on Z. latifolia. The fungus U. esculenta produces proteases and energy acquisition proteins those enhance it's defensive and survival mode in the host. The identified differentially regulated proteins shed-light into why inflorescence is being replaced by bulbous smut-gall at late stages of the disease, as well as the development of resistance in some Z. latifolia plants against U. esculenta infection.