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1.
Phys Chem Chem Phys ; 21(8): 4633-4640, 2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30747176

RESUMO

Using first principles density functional theory, we have studied the interaction mechanism of NO2 and SO2 gas molecules on an MoB2 monolayer, for gas sensing applications. The selectivity for a particular gas by the sensor has been analyzed through electronic structure calculations and adsorption studies. The calculations have been performed by considering the fact that the MoB2 monolayer as a sensing material encounters a change in its electrical properties, when gas molecules with different orientations get adsorbed on the surface. From the density of states study, we find better selectivity for NO2 as compared to SO2, as the latter leaves the electronic structure of the sensing material unaffected. Further, the adsorption curves support the above fact as the larger value of adsorption energy (Ead ∼ -1 eV) for NO2 indicates stronger adsorption. The chemisorptive nature for NO2, in contrast with the relatively weaker physisorption for SO2, additionally supports the fact that NO2 gas has a better perspective for MoB2 sensor application. Charge density plots for each case are in good agreement with the above conclusions. The faster recovery time attributes the MoB2 monolayer better as a sensor material for NO2 interaction.

3.
Microb Cell Fact ; 16(1): 133, 2017 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-28754100

RESUMO

BACKGROUND: A cellular stress response is triggered upon induction of recombinant protein expression which feedback inhibits both growth as well as protein synthesis. In order to separate these two effects, it was decided to study "quiescent cultures" which continue to be metabolically active and express recombinant proteins even after growth cessation. The idea was to identify and up-regulate genes which are responsible for protein synthesis in the absence of growth. This would ensure that, even if growth were adversely affected post induction, there would be no attendant reduction in the protein expression capability of the cells. This strategy allowed us to design host strains, which did not grow better post induction but had significantly higher levels of protein expression. RESULTS: A quiescent Escherichia coli culture, which is able to sustain recombinant protein expression in the absence of growth, was analyzed by transcriptomic and proteomic profiling. Many genes involved in carbon utilization, biosynthesis of building blocks and stress protection were found to be up-regulated in the quiescent phase. Analysis of the global regulators showed that fis, which tends to get down-regulated as the cells enter stationary phase, remained up-regulated throughout the non-growing quiescent phase. The downstream genes regulated by fis like carB, fadB, nrfA, narH and queA were also up-regulated in the quiescent phase which could be the reason behind the higher metabolic activity and protein expression ability of these non-growing cells. To test this hypothesis, we co-expressed fis in a control culture expressing recombinant L-asparaginase and observed a significantly higher buildup of L-asparaginase in the culture medium. CONCLUSIONS: This work represents an important breakthrough in the design of a superior host platform where a gene not directly associated with protein synthesis was used to generate a phenotype having higher protein expression capability. Many alternative gene targets were also identified which may have beneficial effects on expression ability.


Assuntos
Escherichia coli/metabolismo , Proteômica , Asparaginase/genética , Asparaginase/metabolismo , Ácido Aspártico/metabolismo , Regulação para Baixo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Perfilação da Expressão Gênica , Plasmídeos/genética , Plasmídeos/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Transcriptoma , Regulação para Cima
4.
Microb Cell Fact ; 11: 93, 2012 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-22759404

RESUMO

BACKGROUND: A useful goal for metabolic engineering would be to generate non-growing but metabolically active quiescent cells which would divert the metabolic fluxes towards product formation rather than growth. However, for products like recombinant proteins, which are intricately coupled to the growth process it is difficult to identify the genes that need to be knocked-out/knocked-in to get this desired phenotype. To circumvent this we adopted an inverse metabolic engineering strategy which would screen for the desired phenotype and thus help in the identification of genetic targets which need to be modified to get overproducers of recombinant protein. Such quiescent cells would obviate the need for high cell density cultures and increase the operational life span of bioprocesses. RESULTS: A novel strategy for generating a library, consisting of randomly down regulated metabolic pathways in E. coli was designed by cloning small genomic DNA fragments in expression vectors. Some of these DNA fragments got inserted in the reverse orientation thereby generating anti-sense RNA upon induction. These anti-sense fragments would hybridize to the sense mRNA of specific genes leading to gene 'silencing'. This library was first screened for slow growth phenotype and subsequently for enhanced over-expression ability. Using Green Fluorescent Protein (GFP) as a reporter protein on second plasmid, we were able to identify metabolic blocks which led to significant increase in expression levels. Thus down-regulating the ribB gene (3, 4 dihydroxy-2-butanone-4-phosphate synthase) led to a 7 fold increase in specific product yields while down regulating the gene kdpD (histidine kinase) led to 3.2 fold increase in specific yields. CONCLUSION: We have designed a high throughput screening approach which is a useful tool in the repertoire of reverse metabolic engineering strategies for the generation of improved hosts for recombinant protein expression.


Assuntos
Escherichia coli/metabolismo , Engenharia Metabólica , Proteínas Recombinantes/biossíntese , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Transferases Intramoleculares/genética , Transferases Intramoleculares/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , RNA Antissenso/genética , RNA Antissenso/metabolismo , RNA Mensageiro/metabolismo , Proteínas Recombinantes/genética
5.
J Biotechnol ; 351: 99-108, 2022 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-35500701

RESUMO

Over expression of recombinant proteins triggers a cellular stress response (CSR) that down-regulates numerous genes that have a key role in sustaining expression. Instead of trying to individually up-regulate these genes we hypothesized that a superior strategy would be to modulate the expression of global regulators that control the expression of many such downstream genes. Transcriptomic profiling of post induction cultures expressing recombinant asparaginase in Escherichia coli showed the down-regulation of several critical genes many of which were under the control of the global regulator lrp which is known to have a significant impact on both amino acid metabolism and protein translation. Therefore, to ameliorate the deleterious effects of the CSR we decided to supplement the activity of lrp using plasmid-based co-expression. We observed that the test culture containing an additional plasmid expressing lrp under the arabinose promoter gave a 50% higher yield of recombinant L-Asparaginase after 32 h in batch culture compared to the control, which had only one plasmid expressing the recombinant protein. This approach helped us design a better performing strain, which could sustain expression rates for a significantly longer time period. This work illustrates that modifying the expression of regulatory genes could serve as a better strategy to prevent the reprogramming of the cellular machinery which is the hallmark of the CSR and help in the design better hosts for recombinant protein expression.


Assuntos
Proteínas de Escherichia coli , Escherichia coli , Asparaginase/genética , Proteínas de Bactérias/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Leucina/metabolismo , Proteína Reguladora de Resposta a Leucina/genética , Proteína Reguladora de Resposta a Leucina/metabolismo , Proteínas Recombinantes/metabolismo
6.
J Phys Condens Matter ; 32(28): 285501, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32155599

RESUMO

In view of the recent experimental predictions of a weak structural transition in CoV2O4 we explore the possible orbital order states in its low temperature tetragonal phases from first principles density functional theory calculations. We observe that the tetragonal phase with I41/amd symmetry is associated with an orbital order involving complex orbitals with a reasonably large orbital moment at vanadium sites while in the phase with I41/a symmetry, the real orbitals with quenched orbital moment constitute the orbital order. Further, to study the competition between orbital order and electron itinerancy we considered Mn0.5Co0.5V2O4 as one of the parent compounds, CoV2O4, lies near itinerant limit while the other, MnV2O4, lies deep inside the orbitally ordered insulating regime. Orbital order and electron transport have been investigated using first principles density functional theory and Boltzmann transport theory in CoV2O4, MnV2O4 and Mn0.5Co0.5V2O4. Our results show that as we go from MnV2O4 to CoV2O4 there is enhancement in the electron's itinerancy while the nature of orbital order remains unchanged.

7.
Sci Rep ; 10(1): 10850, 2020 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-32616741

RESUMO

Isobutanol, in spite of its significant superiority over ethanol as a biofuel, remains commercially non-viable due to the non-availability of a suitable chassis which can handle the solvent toxicity associated with its production. To meet this challenge, we chose Lactococcus lactis which is known for its ability to handle environmental stress and carried out Adaptive laboratory evolution (ALE) in a continuous stirred tank reactor (CSTR) to evolve an isobutanol tolerant strain. The strain was grown for more than 60 days (> 250 generations) while gradually increasing the selection pressure, i.e. isobutanol concentration, in the feed. This led to the evolution of a strain that had an exceptionally high tolerance of up to 40 g/l of isobutanol even though a scanning electron microscope (SEM) study as well as analysis of membrane potential revealed only minor changes in cellular morphology. Whole genome sequencing which was done to confirm the strain integrity also showed comparatively few mutations in the evolved strain. However, the criticality of these mutations was reflected in major changes that occurred in the transcriptome, where gene expression levels from a wide range of categories that involved membrane transport, amino acid metabolism, sugar uptake and cell wall synthesis were significantly altered. Analysing the synergistic effect of these changes that lead to the complex phenotype of isobutanol tolerance can help in the construction of better host platforms for isobutanol production.


Assuntos
Proteínas de Bactérias/metabolismo , Butanóis/farmacologia , Tolerância a Medicamentos/genética , Genômica/métodos , Laboratórios/estatística & dados numéricos , Lactococcus lactis/genética , Transcriptoma/efeitos dos fármacos , Proteínas de Bactérias/genética , Lactococcus lactis/efeitos dos fármacos , Lactococcus lactis/crescimento & desenvolvimento
8.
Sci Rep ; 9(1): 11194, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31371785

RESUMO

A new multisensor (i.e. resistive and magnetic) CrI3-WTe2 heterostructure (HS) to detect the toxic gases BrF3 and COCl2 (Phosgene) has been theoretically studied in our present investigation. The HS has demonstrated sensitivity towards both the gases by varying its electronic and magnetic properties when gas molecule interacts with the HS. Fast recovery time (<0.14 fs) under UV radiation has been observed. We have considered two configurations of BrF3 adsorbed HS; (1) when F ion interacts with HS (C1) and (2) when Br ion interacts with HS (C2). In C1 case the adsorption energy Ead is observed to be -0.66 eV while in C2 it is -0.95 eV. On the other hand in case of COCl2 Ead is found to be -0.42 eV. Magnetic moments of atoms are also found to vary upon gas adsorption indicates the suitability of the HS as a magnetic gas sensor. Our observations suggest the suitability of CrI3-WTe2 HS to respond detection of the toxic gases like BrF3 and COCl2.

9.
J Clin Sleep Med ; 13(1): 95-104, 2017 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-27784416

RESUMO

STUDY OBJECTIVES: This preliminary study investigated the tolerability and efficacy of a novel mattress technology-the Sound-To-Sleep (STS) system-in the treatment of sleep problems in children with autism. METHODS: After screening, 45 children, ages 2.5 to 12.9 years, were randomized to order of mattress technology use (On-Off vs. Off-On). Treatment conditions (On vs. Off) lasted two weeks with immediate crossover. Tolerability, including study discontinuation and parent-report of mattress tolerance and ease of use, was tracked throughout the study. Efficacy assessments were obtained at baseline, prior to crossover, and end of study and included measures of autism traits, other psychopathology symptoms, sensory abnormalities, communication difficulties, quality of life, sleep diary parameters, and single-blinded actigraphy-derived sleep parameters. Statistical analyses evaluated differences in tolerability and efficacy when the STS system was on versus off. RESULTS: STS system use was well tolerated (n = 2, 4.4% dropout) and resulted in parent-reported sleep quality improvements (STS off mean = 4.3, 95% CI = 4.05-4.54 vs. on mean = 4.9, 95%CI = 4.67-5.14). The technology was described by parents as very easy to use and child tolerance was rated as good. Parent-diary outcomes indicated improvements in falling asleep and reduced daytime challenging behavior. Actigraphy-derived sleep parameters indicated improved sleep duration and sleep efficiency. Improvements in child and family quality of life were identified on parent questionnaires. CONCLUSIONS: A future large sample phase 2 trial of the STS system is warranted and would benefit from extended study duration, an objective primary efficacy outcome, and careful attention to methodological issues that promote compliance with the intervention and study procedures.


Assuntos
Estimulação Acústica/instrumentação , Transtorno Autístico/complicações , Leitos , Transtornos do Sono-Vigília/complicações , Transtornos do Sono-Vigília/terapia , Vibração/uso terapêutico , Estimulação Acústica/métodos , Actigrafia , Criança , Pré-Escolar , Estudos Cross-Over , Feminino , Humanos , Masculino , Estimulação Física/instrumentação , Estimulação Física/métodos , Resultado do Tratamento
10.
Mol Biotechnol ; 59(4-5): 168-178, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28374116

RESUMO

The phenylalanine pathway flux is controlled by two types of regulators, those that are specific to the pathway, as well as by global regulators. In order to demonstrate the importance of these global regulators, we first removed the pathway-specific regulators using all possible combinations of gene knockouts and knockins. We found that genes like aroG fbr performed best individually as well as in combination with other genes, while other genes like tyrA and tyrR worked only in combination with other modifications. Knocking in the tktA gene under a tyrR promoter and knocking out pykF further increased phenylalanine production demonstrating that the supply of precursor via PEP and E4P is also a rate-limiting step. Finally, we tested the role of global regulators on this deregulated pathway and found that Fis overexpression helps in both enhancing and sustaining the flux through this pathway. This work opens up the possibility of using global regulators in synergy with pathway-specific modifications to enhance product yields.


Assuntos
Escherichia coli/genética , Escherichia coli/metabolismo , Melhoramento Genético/métodos , Engenharia Metabólica/métodos , Modelos Biológicos , Fenilalanina/biossíntese , Simulação por Computador , Análise do Fluxo Metabólico/métodos , Redes e Vias Metabólicas/fisiologia , Fenilalanina/isolamento & purificação , Regulação para Cima/genética
11.
Chest ; 130(1): 143-8, 2006 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-16840394

RESUMO

BACKGROUND: Upper airway inflammation is now recognized in adults with obstructive sleep apnea (OSA) syndrome. However, the role played by eicosanoids such as leukotrienes and prostaglandins is unclear. OBJECTIVE: To investigate whether eicosanoids are measurable in exhaled breath condensate (EBC), and to determine whether differences in these inflammatory mediators emerge among children with and without sleep-disordered breathing (SDB). METHODS: EBC was collected from 50 consecutive snoring children undergoing overnight polysomnography for suspected SDB, and from 12 nonsnoring control subjects. Prostaglandin E2 (PGE2), leukotriene B4 (LTB4), and cysteinyl leukotrienes (cys-LTs: leukotriene C4 [LTC4]/leukotriene D4 [LTD4]/leukotriene E4 [LTE4]) EBC levels were analyzed using enzyme-linked immunosorbent assay. RESULTS: LTB4 levels were elevated in children with an apnea-hypopnea index (AHI) > 5/h (SDB; 97.6 +/- 6.3 pg/mL) compared to children with an AHI < 5/h (mild SDB; 66.4 +/- 19.1 pg/mL; p < 0.01) and control subjects (27.8 +/- 3.7 pg/mL; p < 0.01). Similarly, cys-LT (LTC4/LTD4/LTE4) concentrations were also increased in SDB (45.1 +/- 10.6 pg/mL in SDB vs 27.6 +/- 8.3 pg/mL in mild SDB, and 15.7 +/- 7.6 pg/mL in control subjects; p < 0.01). In contrast, PGE2 concentrations were similar among the three groups. CONCLUSIONS: Inflammatory mediators such as leukotrienes and prostaglandins can be readily quantified in EBC collected from the upper airway of children. Disease severity-dependent increases in leukotriene concentrations (LTB4 and LTC4/LTD4/LTE4) emerge among children and may serve as a noninvasive tool in the clinical assessment of these children.


Assuntos
Testes Respiratórios/métodos , Eicosanoides/classificação , Apneia Obstrutiva do Sono/classificação , Adolescente , Testes Respiratórios/instrumentação , Criança , Eicosanoides/isolamento & purificação , Eicosanoides/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Polissonografia , Índice de Gravidade de Doença , Apneia Obstrutiva do Sono/metabolismo , Ronco
12.
Sleep Med ; 7(3): 221-7, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16564219

RESUMO

BACKGROUND: Obstructive sleep apnea (OSA) is the most common form of sleep-disordered breathing, with almost 15 million Americans affected and many more at risk. Current diagnostic approach to OSA requires polysomnography, which is laborious, onerous, and time-consuming. There is ample evidence that inflammatory responses to the perturbations associated with OSA trigger a variety of genes and signaling cascades that ultimately lead to end-organ injury and changes in kidney function and protein expression. The aim of this study was therefore to analyze proteins in human urine and assess whether differential expression of particular proteins is associated with the presence of OSA. METHODS: Eleven OSA and 11 control children between the ages of three and 14 (males=17; females=5) underwent overnight sleep studies followed by a first-morning urine sample. Proteomic analysis of urine samples yielded a unique map of proteins, of which, five spots were selected for further analysis due to their significant intensity differences between OSA and control groups. RESULTS: Mass spectrometry followed by peptide mass fingerprinting conclusively identified four of the five spots as gelsolin, perlecan (a heparan sulfate proteoglycan), albumin, and immunoglobulin (P<0.05 and protein scores>67). CONCLUSIONS: Overall, increased expression of gelsolin and perlecan in the urinary proteome of children with OSA suggests that the episodic hypoxia associated with OSA may lead to changes in protein permeability or alternatively to increased catabolism of these proteins and their excretion in urine.


Assuntos
Gelsolina/genética , Gelsolina/urina , Proteoglicanas de Heparan Sulfato/genética , Proteoglicanas de Heparan Sulfato/urina , Proteômica/métodos , Síndromes da Apneia do Sono/genética , Síndromes da Apneia do Sono/urina , Índice de Massa Corporal , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Oximetria , Polissonografia , Síndromes da Apneia do Sono/diagnóstico , Apneia Obstrutiva do Sono/diagnóstico , Apneia Obstrutiva do Sono/genética , Apneia Obstrutiva do Sono/urina , Sono REM/fisiologia , Ronco/epidemiologia
13.
BMJ Case Rep ; 20162016 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-26917796

RESUMO

Advanced radiological imaging has largely replaced exploratory operations and has become an essential diagnostic tool clinicians routinely rely on. However, physicians are faced with a lot of radiological findings without histological proof, and assuming a more serious diagnosis may lead to unnecessary investigations and emotional stress for patients. We report an unusual presentation of chronic appendicitis with a synchronous peritoneal nodule on CT in a 76-year-old woman who presented with poor appetite, weight loss and a mass in the right iliac fossa. The coincidental finding of the nodule in addition to the suspicious appearance of the appendix raised concerns for primary appendiceal cancer with peritoneal metastasis. The case illustrates the patient's management and reflects on the learnt lessons with regard to careful use of invasive radiology-guided biopsies and interval imaging, as these could sometimes delay the diagnosis and management of a readily treatable disease.


Assuntos
Neoplasias do Apêndice/diagnóstico por imagem , Apendicite/patologia , Neoplasias Peritoneais/diagnóstico por imagem , Idoso , Neoplasias do Apêndice/cirurgia , Diagnóstico Diferencial , Feminino , Humanos , Neoplasias Peritoneais/cirurgia , Tomografia Computadorizada por Raios X
14.
J Clin Sleep Med ; 12(3): 311-7, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26518700

RESUMO

STUDY OBJECTIVES: While neck circumference (NC) is a useful predictor of obstructive sleep apnea (OSA) in adults, childhood OSA is difficult to predict clinically. We utilized the neck circumference-height ratio (NHR) to normalize NC in growing children. Our study aimed to determine if (1) NC is a reproducible clinical measurement; (2) NHR predicts OSA in children; (3) this metric translates to adults. METHODS: For this retrospective study, paired NC measurements (from clinic and sleep laboratory) in 100 consecutive adult subjects were used to confirm inter-observer reproducibility. Polysomnographic (PSG) and anthropometric data from children aged 5-18 years presenting consecutively between July 2007 and February 2012 was obtained. Children with genetic syndromes, severe neurological disorders, craniofacial abnormalities, tracheostomy, past adenotonsillectomy, in-hospital PSG or sleep efficiency < 80% were excluded. Data were analyzed using χ(2) test and logistic and linear regression models. These analyses were also applied to 99 adult patients with similar exclusion criteria. RESULTS: Adult NC measurement had inter-observer correlation of 0.85 (N = 100). Among children, after correcting for BMI-Z scores, NHR conferred additional predictive value, in both logistic regression and linear models, for both apnea-hypopnea index (AHI) > 2 and > 5 (N = 507). In children, for NHR > 0.25, the odds ratio of AHI > 2 was 3.47. In adults, for NHR > 0.25, the odds ratio of AHI > 5 was 18. CONCLUSIONS: NHR can be included as a simple screening tool for OSA in children and adults, which along with other predictors, may improve the ability of clinicians to triage children and adults at risk for OSA for further evaluation with PSG.


Assuntos
Estatura , Pescoço/anatomia & histologia , Síndromes da Apneia do Sono/diagnóstico , Adolescente , Pesos e Medidas Corporais/estatística & dados numéricos , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Polissonografia/estatística & dados numéricos , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Estudos Retrospectivos , Fatores de Risco , Índice de Gravidade de Doença
15.
Int J Pediatr Otorhinolaryngol ; 83: 25-30, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26968048

RESUMO

OBJECTIVE: There is a lack of consensus and a paucity of data regarding how to best treat pediatric patients with mild obstructive sleep apnea. The objective of our study was to compare outcomes following adenotonsillectomy vs. observation in children with mild obstructive sleep apnea based on polysomnography results. METHODS: A retrospective chart review was performed on children ages 9 months to 9 years with 2 or more polysomnograms completed at a tertiary care academic center. Children diagnosed with mild obstructive sleep apnea (obstructive apnea-hypopnea index 1-5) on polysomnography performed from 1999 to 2013 were included. Patients underwent adenotonsillectomy or watchful waiting for obstructive sleep apnea. The primary outcome was the change in apnea-hypopnea index. RESULTS: There were 62 patients who met inclusion criteria for the study; 19 of the 62 patients were obese, while 15 had a craniofacial syndrome or hypotonia. Eighteen patients underwent adenotonsillectomy for mild obstructive sleep apnea while 44 were observed. The mean apnea-hypopnea index of patients after adenotonsillectomy improved from 3.50 (95% Confidence Interval [CI] 2.97-4.03) to 2.69 (95% CI 1.48-3.90), while the mean apnea-hypopnea index of the observation group worsened from 3.09 (95% CI 2.76-3.42) to 5.18 (95% CI 2.46-7.90). Between-group analysis showed significant improvement in the surgery group (p=0.03), with a persistent improvement on multivariate analysis adjusting for baseline apnea-hypopnea index (p=0.05). This difference was seen mostly in non-obese, non-syndromic children (p=0.04). There was no significant difference between groups amongst obese (p=0.25) and syndromic (p=0.36) patients. CONCLUSIONS: Adenotonsillectomy leads to a significant improvement in apnea-hypopnea index on follow-up polysomnography over an observational approach, especially in non-obese, non-syndromic children. A prospective, randomized trial is necessary to help determine appropriate treatment strategies for pediatric mild obstructive sleep apnea.


Assuntos
Adenoidectomia/métodos , Polissonografia/métodos , Apneia Obstrutiva do Sono/terapia , Tonsilectomia/métodos , Conduta Expectante/métodos , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Estudos Retrospectivos , Apneia Obstrutiva do Sono/fisiopatologia
16.
Mol Biotechnol ; 30(2): 103-16, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15920280

RESUMO

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic growth factor, that has been used as a therapeutic agent in facilitating bone marrow and stem cell transplantation and in other clinical cases like neutropenia. Although biologically active recombinant GM-CSF has been successfully produced in Escherichia coli, the reported levels are extremely poor. In this study we looked into the possible reasons for poor expression and found that protein toxicity coupled with protease-based degradation was the principal reason for low productivity. To overcome this problem we attached a signal sequence, as well as an amino-terminal His-tag fusion to the GM-CSF gene. This combination had a dramatic effect on expression levels, which increased from 0.8 microg/mL in the control to 40 microg/mL. When a larger fusion partner, such as the Maltose-binding protein (MBP-tag), was used the expression levels increased further to 69.5 microg/mL, which along with the MBP-tag represented approx 12% of the total cellular protein.


Assuntos
Escherichia coli/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/biossíntese , Sinais Direcionadores de Proteínas , Proteínas de Transporte/genética , Códon/genética , Escherichia coli/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Microbiologia Industrial , Proteínas Ligantes de Maltose , Periplasma/química , Periplasma/metabolismo , Plasmídeos/genética , Estabilidade de RNA , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes
17.
J Clin Sleep Med ; 11(7): 741-6, 2015 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-25845895

RESUMO

STUDY OBJECTIVES: Caffeine, a commonly consumed psychoactive substance, can have significant effects on sleep. Caffeine intake among children is increasing, mainly in the form of sodas. However, adolescent caffeine consumers may lack knowledge about the caffeine content in common beverages. If true, this very fact may hamper the assessment of the effects of caffeine consumption on sleep in children if such assessments are a priori dependent on responders being able to reliably distinguish between caffeinated and noncaffeinated beverages. This preliminary study investigated adolescents' caffeine knowledge and intake at a Cleveland-area public middle school. METHODS: Seventh- and eighth-grade students were surveyed using: (1) the Caffeine Literacy and Sleep Study (CLASS), a 15-question pilot instrument designed to assess caffeine knowledge and intake by type, quantity and timing, as well as sleep habits; and (2) the Cleveland Adolescent Sleepiness Questionnaire (CASQ), a validated survey measuring excessive daytime sleepiness in adolescents. These questionnaires were distributed and collected during a specified class period. RESULTS: Of the 635 seventh- and eighth-grade students who attended school on the day of the study, 555 (87%) participated. Lack of knowledge about caffeine content of particular drinks was noted in seventh and eighth graders of both sexes with nearly 29% unaware that their favorite drinks contain caffeine and more than 50% unable to correctly identify the drinks with the most caffeine. A low percentage of students correctly identified light-colored sodas lacking caffeine: 7-Up (24.1%), Sierra Mist (38.9%), ginger ale (39.8%), Sprite (39.8%), and Fresca (53.7%). The percentages of students correctly identifying caffeinated light-colored beverages were: Arizona Green Tea (43.5%), Mello Yellow (50.9%), and A&W cream soda (67.6%). However, Mountain Dew was correctly identified by most (93.5%) as caffeinated. CONCLUSIONS: Students were not consistently able to identify caffeine content or lack thereof in some common beverages. The results of this pilot study show that caffeine literacy in adolescents warrants further investigation and educational intervention.


Assuntos
Bebidas/efeitos adversos , Cafeína/efeitos adversos , Conhecimentos, Atitudes e Prática em Saúde , Transtornos do Sono-Vigília/induzido quimicamente , Adolescente , Fatores Etários , Conscientização , Bebidas/estatística & dados numéricos , Cafeína/administração & dosagem , Criança , Estudos Transversais , Feminino , Humanos , Incidência , Masculino , Avaliação das Necessidades , Projetos Piloto , Medição de Risco , Fatores Sexuais , Transtornos do Sono-Vigília/epidemiologia , Estudantes/estatística & dados numéricos , Inquéritos e Questionários , Estados Unidos
18.
J Biosci Bioeng ; 115(3): 291-7, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23098681

RESUMO

The kinetics of recombinant human granulocyte-macrophage colony-stimulating factor (hGM-CSF) expression was studied under the strong T7 promoter in continuous culture of Escherichia coli using complex medium to design an optimum feeding strategy for high cell density cultivation. Continuous culture studies were done at different dilution rates and the growth and product formation profiles were monitored post-induction. Recombinant protein expression was in the form of inclusion bodies with a maximum specific product formation rate (q(p)) of 63.5 mg g(-1) DCW h(-1) at a dilution rate (D) of 0.3 h(-1). The maximum volumetric product concentration achieved at this dilution rate was 474 mg l(-1), which translated a ~1.4 and ~1.75 folds increase than the values obtained at dilution rates of 0.2 h(-1) and 0.4 h(-1) respectively. The specific product yield (Y(P/x)) peaked at 138 mg g(-1) DCW, demonstrating a ~1.6 folds increase in the values obtained at other dilution rates. A drop in q(p) was observed within 5-6 h of induction at all the dilution rates, possibly due to protein toxicity and metabolic stress associated with protein expression. The data from the continuous culture studies allowed us to design an optimal feeding strategy and induction time in fed-batch cultures which resulted in a maximum product concentration of 3.95 g l(-1) with a specific hGM-CSF yield (Y(P/x)) of 107 mg g(-1) DCW.


Assuntos
Escherichia coli/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/biossíntese , Técnicas de Cultura Celular por Lotes , Escherichia coli/genética , Escherichia coli/crescimento & desenvolvimento , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Corpos de Inclusão/metabolismo , Cinética , Modelos Biológicos , Proteínas Recombinantes/biossíntese
19.
J Clin Sleep Med ; 7(2): 158-62, 2011 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-21509330

RESUMO

INTRODUCTION: Children with Down syndrome (DS) commonly have obstructive sleep apnea syndrome (OSAS) and may assume a unique sleeping position not systematically described previously. We describe this sleep position in DS and explore its relationship with OSAS in comparison to control children (CC). METHODS: Overnight video-polysomnograms (PSG) of consecutive children with DS (age 2-18 y), referred to our center between April 2008 and October 2009, were retrospectively analyzed by a single scorer (ES). CC group comprised age and gender matched, non-syndromic, neurologically intact children referred to us for suspected OSAS over the same period. RESULTS: Each group had 17 subjects matched for age (median [IQR]; 6 [4-8]) and gender (65% female). DS group had higher BMI (median [IQR]; 18.8 [17.4-21.0]) than CC (17 [14.7 -18.8]; p = 0.03). There were however no significant differences (median [IQR]) between DS and CC with respect to sleep time in minutes (460 [425-499] vs 424[410-483]), sleep efficiency (%) (90.9 [87.4-92.4] vs 88.6 [79.9-93.1]), REM time (%) (17.1 [14.2-22.1] vs 19.2 [14.9-22.1]), supine time (%) (40.7 [24.8-56.0] vs 15.8 [0.40-44.5], p 0.06), mean oxygen saturation (%) (95 [94-96] vs 96 [95-97]), oxygen saturation nadir (89 [86-91] vs 89[94-92]), or total apnea-hypopnea index (4.3 [3-7.8] vs 5.1[1.9-9.6]). Despite these similarities between the groups, 9 (53%) DS children slept seated bent forward with head resting on bed for at least part of the total sleep time (%) (7.8 ± 10.9, range 0.8-35.7).This was absent in the CC group (p = 0.06). CONCLUSION: Some DS children assume a peculiar body position, sitting cross-legged flopped-forward with head resting on bed while asleep. This is absent in age- and gender-matched controls showing otherwise similar PSG characteristics. The reason for this posture is unclear from this study. However, this novel report of a unique sleeping position would provide us with a basis to conduct a prospective study involving a larger population to ascertain the contribution of this position for OSAS protection or to determine if it may be forme fruste parasomnia.


Assuntos
Síndrome de Down/complicações , Apneia Obstrutiva do Sono/etiologia , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Polissonografia , Postura/fisiologia , Decúbito Ventral/fisiologia , Sono/fisiologia , Decúbito Dorsal/fisiologia , Gravação em Vídeo
20.
Arch Otolaryngol Head Neck Surg ; 137(12): 1269-75, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22183910

RESUMO

OBJECTIVES: To identify the prevalence of clinically relevant findings during cardiac evaluations of pediatric patients with obstructive sleep apnea (OSA) undergoing adenotonsillectomy (TA), and to determine the association between cardiac findings and postoperative respiratory complications. DESIGN: Retrospective medical chart review. PATIENTS: Pediatric patients aged 10 months to 15 years who underwent both echocardiography and polysomnography (PSG) within 6 months prior to TA for OSA from April 2007 through April 2011. MAIN OUTCOME MEASURES: Two pediatric cardiologists independently reviewed echocardiographic studies for evidence of cardiovascular disease. Patients were stratified based on apnea-hypopnea index (AHI) severity (1-5, >5-10, and >10). These groups were compared according to demographic, electrocardiographic (ECG), and echocardiographic values, and postoperative respiratory complications. RESULTS: The medical charts of 57 of 900 patients identified were reviewed following exclusion of those with congenital cardiac abnormalities. The AHI groupings did not differ demographically. No clinically relevant abnormalities were identified on the echocardiogram of any patient. There was a statistically significant association between increased AHI and the appearance of postoperative respiratory complications (P < .05). Indicators of myocardial hypertrophy, such as left ventricular mass index, were not significantly related to AHI in contrast to previously published studies. No echocardiographic or ECG findings were identified that were associated with increased number of postoperative respiratory complications or OSA severity based on AHI. CONCLUSIONS: The lack of clinically relevant findings during preoperative cardiac evaluations suggests that aggressive cardiac workup in pediatric patients with OSA may not be indicated unless dictated by comorbidities. Consistent with results in prior studies, preoperative AHI can identify patients at risk for respiratory complications following TA.


Assuntos
Adenoidectomia , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/epidemiologia , Complicações Pós-Operatórias/epidemiologia , Complicações Pós-Operatórias/etiologia , Cuidados Pré-Operatórios , Apneia Obstrutiva do Sono/epidemiologia , Apneia Obstrutiva do Sono/cirurgia , Tonsilectomia , Adolescente , Criança , Pré-Escolar , Comorbidade , Comportamento Cooperativo , Ecocardiografia , Eletrocardiografia , Feminino , Humanos , Lactente , Comunicação Interdisciplinar , Modelos Logísticos , Masculino , Análise Multivariada , Equipe de Assistência ao Paciente , Polissonografia , Estudos Retrospectivos , Fatores de Risco , Procedimentos Desnecessários
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