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CONTEXT: The percent of underrepresented minority (URM) students who apply to medical school has changed minimally in the past 40 years. Due to the lack of URM applicants, the consequent matriculation of URMs is grossly disproportionate from their percent representation of the US population. Increasing diversity among medical students and physicians has previously been identified as essential to decreasing healthcare disparities among US minorities. OBJECTIVES: The objective of our study was to recognize the barriers of applying to medical school among URMs in high school. METHODS: To identify and assess the prevalence of barriers, surveys were distributed to participants of Med-Achieve, a mini-medical school program of diverse high school students in New York City during the 2019-2020 academic year. RESULTS: Among students who will be first in their immediate family to attend college, 80.0% perceived a barrier to pursuing medical school. Specified barriers indicated include the cost of medical school (77%), a lack of guidance/role models (53.9%), and the predicted inability to do well in medical school classes (53.9%). At the end of the program, a statistically significant reduction in the barrier of lack of guidance/role models was seen. CONCLUSIONS: This study highlights the benefit of mini-medical school programs, especially programs with a mentoring component, to decrease the perceived barriers of applying to medical school among URMs. It also suggests the potential role of similar programs to increase diversity in medicine and to decrease healthcare disparities among minorities in the United States.
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Faculdades de Medicina , Estudantes de Medicina , Diversidade Cultural , Humanos , Mentores , Grupos Minoritários , Estados UnidosRESUMO
[This corrects the article DOI: 10.1371/journal.pone.0063083.].
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Global brain ischemia/reperfusion induces neuronal damage in vulnerable brain regions, leading to mitochondrial dysfunction and subsequent neuronal death. Induction of neuronal death is mediated by release of cytochrome c (cyt c) from the mitochondria though a well-characterized increase in outer mitochondrial membrane permeability. However, for cyt c to be released it is first necessary for cyt c to be liberated from the cristae junctions which are gated by Opa1 oligomers. Opa1 has two known functions: maintenance of the cristae junction and mitochondrial fusion. These roles suggest that Opa1 could play a central role in both controlling cyt c release and mitochondrial fusion/fission processes during ischemia/reperfusion. To investigate this concept, we first utilized in vitro real-time imaging to visualize dynamic changes in mitochondria. Oxygen-glucose deprivation (OGD) of neurons grown in culture induced a dual-phase mitochondrial fragmentation profile: (i) fragmentation during OGD with no apoptosis activation, followed by fusion of mitochondrial networks after reoxygenation and a (ii) subsequent extensive fragmentation and apoptosis activation that preceded cell death. We next evaluated changes in mitochondrial dynamic state during reperfusion in a rat model of global brain ischemia. Evaluation of mitochondrial morphology with confocal and electron microscopy revealed a similar induction of fragmentation following global brain ischemia. Mitochondrial fragmentation aligned temporally with specific apoptotic events, including cyt c release, caspase 3/7 activation, and interestingly, release of the fusion protein Opa1. Moreover, we uncovered evidence of loss of Opa1 complexes during the progression of reperfusion, and electron microscopy micrographs revealed a loss of cristae architecture following global brain ischemia. These data provide novel evidence implicating a temporal connection between Opa1 alterations and dysfunctional mitochondrial dynamics following global brain ischemia.
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Isquemia Encefálica/metabolismo , Dinâmica Mitocondrial , Traumatismo por Reperfusão/metabolismo , Animais , Apoptose , Região CA1 Hipocampal/irrigação sanguínea , Região CA1 Hipocampal/metabolismo , Hipóxia Celular , Linhagem Celular , Células Cultivadas , GTP Fosfo-Hidrolases/metabolismo , Masculino , Camundongos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Ratos , Ratos Sprague-DawleyRESUMO
Annulate lamellae are cytoplasmic organelles containing stacked sheets of membranes embedded with pore complexes. These cytoplasmic pore complexes at annulate lamellae are morphologically similar to nuclear pore complexes at the nuclear envelope. Although annulate lamellae has been observed in nearly all types of cells, their biological functions are still largely unknown. Here we show that SUMO1-modification of the Ran GTPase-activating protein RanGAP1 not only target RanGAP1 to its known sites at nuclear pore complexes but also to annulate lamellae pore complexes through interactions with the Ran-binding protein RanBP2 and the SUMO-conjugating enzyme Ubc9 in mammalian cells. Furthermore, upregulation of annulate lamellae, which decreases the number of nuclear pore complexes and concurrently increases that of annulate lamellae pore complexes, causes a redistribution of nuclear transport receptors including importin α/ß and the exportin CRM1 from nuclear pore complexes to annulate lamellae pore complexes and also reduces the rates of nuclear import and export. Moreover, our results reveal that importin α/ß-mediated import complexes initially accumulate at annulate lamellae pore complexes upon the activation of nuclear import and subsequently disassociate for nuclear import through nuclear pore complexes in cells with upregulation of annulate lamellae. Lastly, CRM1-mediated export complexes are concentrated at both nuclear pore complexes and annulate lamellae pore complexes when the disassembly of these export complexes is inhibited by transient expression of a Ran GTPase mutant arrested in its GTP-bound form, suggesting that RanGAP1/RanBP2-activated RanGTP hydrolysis at these pore complexes is required for the dissociation of the export complexes. Hence, our findings provide a foundation for further investigation of how upregulation of annulate lamellae decreases the rates of nuclear transport and also for elucidation of the biological significance of the interaction between annulate lamellae pore complexes and nuclear transport complexes in mammalian cells.
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Transporte Ativo do Núcleo Celular/fisiologia , Proteínas Ativadoras de GTPase/metabolismo , Chaperonas Moleculares/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Poro Nuclear/fisiologia , Proteína SUMO-1/metabolismo , Células 3T3 , Animais , Linhagem Celular Tumoral , Citoplasma/fisiologia , Proteínas Ativadoras de GTPase/genética , Células HEK293 , Células HeLa , Humanos , Carioferinas/metabolismo , Camundongos , Interferência de RNA , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley , Receptores Citoplasmáticos e Nucleares/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , alfa Carioferinas/metabolismo , beta Carioferinas/metabolismo , Proteína Exportina 1RESUMO
The 5-phosphoinositide phosphatase Sac3, in which loss-of-function mutations are linked to neurodegenerative disorders, forms a stable cytosolic complex with the scaffolding protein ArPIKfyve. The ArPIKfyve-Sac3 heterodimer interacts with the phosphoinositide 5-kinase PIKfyve in a ubiquitous ternary complex that couples PtdIns(3,5)P2 synthesis with turnover at endosomal membranes, thereby regulating the housekeeping endocytic transport in eukaryotes. Neuron-specific associations of the ArPIKfyve-Sac3 heterodimer, which may shed light on the neuropathological mechanisms triggered by Sac3 dysfunction, are unknown. Here we conducted mass spectrometry analysis for brain-derived interactors of ArPIKfyve-Sac3 and unraveled the α-synuclein-interacting protein Synphilin-1 (Sph1) as a new component of the ArPIKfyve-Sac3 complex. Sph1, a predominantly neuronal protein that facilitates aggregation of α-synuclein, is a major component of Lewy body inclusions in neurodegenerative α-synucleinopathies. Modulations in ArPIKfyve/Sac3 protein levels by RNA silencing or overexpression in several mammalian cell lines, including human neuronal SH-SY5Y or primary mouse cortical neurons, revealed that the ArPIKfyve-Sac3 complex specifically altered the aggregation properties of Sph1-GFP. This effect required an active Sac3 phosphatase and proceeded through mechanisms that involved increased Sph1-GFP partitioning into the cytosol and removal of Sph1-GFP aggregates by basal autophagy but not by the proteasomal system. If uncoupled from ArPIKfyve elevation, overexpressed Sac3 readily aggregated, markedly enhancing the aggregation potential of Sph1-GFP. These data identify a novel role of the ArPIKfyve-Sac3 complex in the mechanisms controlling aggregate formation of Sph1 and suggest that Sac3 protein deficiency or overproduction may facilitate aggregation of aggregation-prone proteins, thereby precipitating the onset of multiple neuronal disorders.
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Proteínas de Transporte/metabolismo , Flavoproteínas/metabolismo , Corpos de Lewy/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Doenças Neurodegenerativas/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Animais , Células COS , Chlorocebus aethiops , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Doenças Neurodegenerativas/enzimologia , Ligação ProteicaRESUMO
Penicillum janthinellum SDX7 was isolated from aged petroleum hydrocarbon-affected soil at the site of Anand, Gujarat, India, and was tested for different pH, temperature, agitation and concentrations for optimal growth of the isolate that was capable of degrading upto 95%, 63% and 58% of 1%, 3% and 5% kerosene, respectively, after a period of 16 days, at optimal growth conditions of pH 6.0, 30 °C and 180 rpm agitation. The GC/MS chromatograms revealed that then-alkane fractions are easily degraded; however, the rate might be lower for branched alkanes, n-alkylaromatics, cyclic alkanes and polynuclear aromatics. The test doses caused a concentration-dependent depletion of carbohydrates of P. janthinellum SDX7 by 3% to 80%, proteins by 4% to 81% and amino acids by 8% to 95% upto 16 days of treatment. The optimal concentration of 3% kerosene resulted in the least reduction of the metabolites of P. janthinellum such as carbohydrates, proteins and amino acids with optimal growth compared to 5% and 1% (v/v) kerosene doses on the 12(th) and 16(th) day of exposure. Phenols were found to be mounted by 43% to 66% at lower and higher concentrations during the experimental period. Fungal isolate P. janthinellum SDX7 was also tested for growth on various xenobiotic compounds.
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Querosene , Penicillium/crescimento & desenvolvimento , Penicillium/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Xenobióticos/metabolismo , Composição de Bases , Biotransformação , DNA Fúngico/química , DNA Fúngico/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Cromatografia Gasosa-Espectrometria de Massas , Genes de RNAr , Concentração de Íons de Hidrogênio , Índia , Dados de Sequência Molecular , Penicillium/genética , Penicillium/isolamento & purificação , RNA Fúngico/genética , RNA Ribossômico 18S/genética , Análise de Sequência de DNA , TemperaturaRESUMO
Penicillum janthinellum SDX7 was isolated from aged petroleum hydrocarbon-affected soil at the site of Anand, Gujarat, India, and was tested for different pH, temperature, agitation and concentrations for optimal growth of the isolate that was capable of degrading upto 95%, 63% and 58% of 1%, 3% and 5% kerosene, respectively, after a period of 16 days, at optimal growth conditions of pH 6.0, 30 °C and 180 rpm agitation. The GC/MS chromatograms revealed that then-alkane fractions are easily degraded; however, the rate might be lower for branched alkanes, n-alkylaromatics, cyclic alkanes and polynuclear aromatics. The test doses caused a concentration-dependent depletion of carbohydrates of P. janthinellum SDX7 by 3% to 80%, proteins by 4% to 81% and amino acids by 8% to 95% upto 16 days of treatment. The optimal concentration of 3% kerosene resulted in the least reduction of the metabolites of P. janthinellum such as carbohydrates, proteins and amino acids with optimal growth compared to 5% and 1% (v/v) kerosene doses on the 12th and 16th day of exposure. Phenols were found to be mounted by 43% to 66% at lower and higher concentrations during the experimental period. Fungal isolate P. janthinellum SDX7 was also tested for growth on various xenobiotic compounds.
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Querosene , Penicillium/crescimento & desenvolvimento , Penicillium/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Xenobióticos/metabolismo , Composição de Bases , Biotransformação , DNA Fúngico/química , DNA Fúngico/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Cromatografia Gasosa-Espectrometria de Massas , Genes de RNAr , Concentração de Íons de Hidrogênio , Índia , Dados de Sequência Molecular , Penicillium/genética , Penicillium/isolamento & purificação , RNA Fúngico/genética , /genética , Análise de Sequência de DNA , TemperaturaRESUMO
The endoplasmic reticulum (ER) is responsible for processing of proteins that are destined to be secreted, enclosed in a vesicle, or incorporated in the plasma membrane. Nascent peptides that enter the ER undergo a series of highly regulated processing steps to reach maturation as they transit the ER. Alterations in the intracellular environment that induce ER stress are thought to interrupt these processing steps, and result in unfolding of proteins in the ER. Accumulation of unfolded proteins concurrently activates three transmembrane stress sensors, IRE1, ATF6 and PERK, and is referred to as the Unfolded Protein Response (UPR). Our understanding of the mechanisms of UPR induction has been assembled primarily from experiments inducing ER stress with chemical and genetic manipulations. However, physiological stress often induces activation of ER stress sensors in a distinct manner from the canonical UPR. The unique activation profiles in vivo have prompted us to examine the mechanism of UPR activation in neurons following cerebral ischemia.
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Isquemia Encefálica/metabolismo , Estresse do Retículo Endoplasmático , Resposta a Proteínas não Dobradas , Fator 6 Ativador da Transcrição/metabolismo , Animais , Endorribonucleases/metabolismo , Humanos , Proteínas Serina-Treonina Quinases/metabolismo , eIF-2 Quinase/metabolismoRESUMO
Cellular mechanisms involved in multiple neurodegenerative diseases converge on mitochondria to induce overproduction of reactive oxygen species, damage to mitochondria, and subsequent cytochrome c release. Little is currently known regarding the contribution mitochondrial dynamics play in cytochrome c release following oxidative stress in neurodegenerative disease. Here we induced oxidative stress in the HT22 cell line with glutamate and investigated key mediators of mitochondrial dynamics to determine the role this process may play in oxidative stress induced neuronal death. We report that glutamate treatment in HT22 cells induces increase in reactive oxygen species (ROS), release of the mitochondrial fusion protein Opa1 into the cytosol, with concomitant release of cytochrome c. Furthermore, following the glutamate treatment alterations in cell signaling coincide with mitochondrial fragmentation which culminates in significant cell death in HT22 cells. Finally, we report that treatment with the antioxidant tocopherol attenuates glutamate induced-ROS increase, release of mitochondrial Opa1 and cytochrome c, and prevents cell death.
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GTP Fosfo-Hidrolases/metabolismo , Mitocôndrias/metabolismo , Estresse Oxidativo , Animais , Antioxidantes/farmacologia , Linhagem Celular , Camundongos , Mitocôndrias/efeitos dos fármacos , Tocoferóis/farmacologiaRESUMO
Present study reports the validation (oxidation) of computationally predicted oxidation of xenobiotic contaminants by commercially available pure laccase from Trametes versicolor. Selected contaminants were predicted as potential targets for laccase oxidation by using in-silico docking tool. The oxidation by laccase was measured by change in absorbance at specific λ max of each compound. Sinapic acid and tyrosine were taken as positive and negative controls, respectively. Oxidation was observed in m-chlorophenol, 2,4 di-chlorophenol, 2,4,6 tri-chlorophenol, captan, atrazine and thiodicarb, except malathion, which showed no activity. It could be speculated that the predicted substrates showing oxidation shared homology at structural and chemical level with positive control compounds. In case of malathion, structural non-homology with sinapic acid could be attributed to its inactivity towards laccase that required further structural analysis. Thus, a remediation tool proposing an advanced remediation approach combining the application of theoretical in-silico method and subsequent experimental validation using pure laccase could be proposed. As number and type of xenobiotics increase, the unfeasibility to screen them experimentally for bioremediation also rise. This approach would be useful to reduce the time and cost required in other screening methods.
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Recent advancements in isolation techniques for cytochrome c (Cytc) have allowed us to discover post-translational modifications of this protein. We previously identified two distinct tyrosine phosphorylated residues on Cytc in mammalian liver and heart that alter its electron transfer kinetics and the ability to induce apoptosis. Here we investigated the phosphorylation status of Cytc in ischemic brain and sought to determine if insulin-induced neuroprotection and inhibition of Cytc release was associated with phosphorylation of Cytc. Using an animal model of global brain ischemia, we found a â¼50% decrease in neuronal death in the CA1 hippocampal region with post-ischemic insulin administration. This insulin-mediated increase in neuronal survival was associated with inhibition of Cytc release at 24 hours of reperfusion. To investigate possible changes in the phosphorylation state of Cytc we first isolated the protein from ischemic pig brain and brain that was treated with insulin. Ischemic brains demonstrated no detectable tyrosine phosphorylation. In contrast Cytc isolated from brains treated with insulin showed robust phosphorylation of Cytc, and the phosphorylation site was unambiguously identified as Tyr97 by immobilized metal affinity chromatography/nano-liquid chromatography/electrospray ionization mass spectrometry. We next confirmed these results in rats by in vivo application of insulin in the absence or presence of global brain ischemia and determined that Cytc Tyr97-phosphorylation is strongly induced under both conditions but cannot be detected in untreated controls. These data suggest a mechanism whereby Cytc is targeted for phosphorylation by insulin signaling, which may prevent its release from the mitochondria and the induction of apoptosis.
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Isquemia Encefálica/metabolismo , Citocromos c/metabolismo , Insulina/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Processamento de Proteína Pós-Traducional , Tirosina/metabolismo , Animais , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/patologia , Região CA1 Hipocampal/efeitos dos fármacos , Região CA1 Hipocampal/metabolismo , Região CA1 Hipocampal/patologia , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/genética , Expressão Gênica , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Fosforilação/efeitos dos fármacos , Ratos , Transdução de Sinais/efeitos dos fármacos , Espectrometria de Massas por Ionização por Electrospray , SuínosRESUMO
A facultatively anaerobic, alkaliphilic, Gram-stain-positive, rod-shaped bacterium, designated strain 12/1(T), isolated from alkaline wastewater drained sludge of a beverage industry facility located near New Delhi, India, was subjected to a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain 12/1(T) belonged to the genus Exiguobacterium and was most closely related to Exiguobacterium aurantiacum DSM 6208(T) (99.46â%), E. aquaticum IMTB-3094(T) (99.18â%), E. mexicanum 8N(T) (99.06â%), E. profundum 10C(T) (98.17â%), E. aestuarii TF-16(T) (98.1â%) and E. marinum TF-80(T) (98.03â%). The DNA G+C content of strain 12/1(T) was 55.6 mol%, major respiratory isoprenoid quinone was MK-7, major polar lipids were phosphatidylglycerol, diphosphatidylglycerol and phosphatidylethanolamine and the cell-wall peptidoglycan was of the A3α l-Lys-Gly type, characteristics consistent with its affiliation to the genus Exiguobacterium. Strain 12/1(T) showed levels of DNA-DNA hybridization of less than 70â% with the closely related species of the genus Exiguobacterium. Overall, the phenotypic, chemotaxonomic and phylogenetic data presented in this study suggest that strain 12/1(T) represents a novel species of the genus Exiguobacterium, for which the name Exiguobacterium alkaliphilum sp. nov. is proposed. The type strain is 12/1(T) (â=âCCM 8459(T)â=âDSM 21148(T)).
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Bacillaceae/classificação , Bebidas , Filogenia , Águas Residuárias/microbiologia , Bacillaceae/genética , Bacillaceae/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Indústria Alimentícia , Índia , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Peptidoglicano/química , RNA Ribossômico 16S/genética , Esgotos/microbiologia , Vitamina K 2/análogos & derivados , Vitamina K 2/químicaRESUMO
Extremophiles are the microorganisms which can survive under extreme conditions of temperature, pressure, pH, salinity etc. They have gained much attention for their potential role in biotechnological and industrial applications. The large amount of experimental data in the literature is so diverse, that it becomes difficult and time consuming for the researcher to implement it in various areas of research. Therefore, a systematic arrangement of data and redirection in a similar fashion through web interface can assist researchers in analyzing the data as per their requirement. ExtremeDB is a freely available web based relational database which integrates general characteristics, genome-proteome information, industrial applications and recent scientific investigations of the seven major groups of 865 extremophillic microorganisms. The search options are user friendly and analyses tools such as Compare and Extreme BLAST have been incorporated for comparative analysis of two or more extremophiles and determining the sequence similarity of a given protein/nucleotide in relation to other extremophiles respectively. The effort put forth herein in the form of database, would open up new avenues on the potential utility of extremophiles in applied research. ExtremeDB is freely accessible via http://extrem.igib.res.in.
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Archaea/metabolismo , Bactérias/metabolismo , Bases de Dados Factuais , Internet , Archaea/genética , Bactérias/genética , Genoma Arqueal/genética , Genoma Bacteriano/genética , Proteoma/metabolismoRESUMO
OBJECTIVES: Hypothermia has a well-established neuroprotective effect and offers a foundation for combination therapy for brain ischemia. The authors evaluated the effect of combination therapy with insulin-like growth factor-1 (IGF-1) and hypothermia on brain structure and function in the setting of global brain ischemia and reperfusion in rats. METHODS: Male Sprague-Dawley rats were randomly assigned to groups by a registrar. Animals were subjected to 8 minutes of global brain ischemia using bilateral carotid occlusion and systemic hypotension, followed by 7 days (Stage I dose studies) or 28 days (Stage II outcome studies) of reperfusion. Sham controls were subjected to surgery, but not ischemia. Stage II animals were randomized to no treatment, IGF-1 at the dose determined in Stage I, hypothermia (32°C for 4 hours), or a combination of IGF-1 and hypothermia. Stage II animals underwent 21 days of spatial memory testing. At 7 days (Stage I) or 28 days (Stage II), brains were harvested for counting of CA1 neurons. The primary Stage II outcome was a neurologic outcome index computed as the ratio of viable CA1 neurons per 300-µm field to the number of days to reach success criteria on the memory task. RESULTS: Stage I experiments confirmed the neuroprotective effect of the hypothermia protocol and IGF-1 at a dose of 0.6 U/kg. Stage II studies suggested that early neuroprotection with hypothermia and IGF-1 was not well maintained to 28 days and that combination therapy was more beneficial than either IGF-1 or hypothermia alone. Median and interquartile ranges (IQRs) of viable neurons per 300-µm field were 114 (IQR = 99.5 to 136) for sham, three (IQR = 2 to 4.8) for untreated ischemia, four (IQR = 3 to 70.25) for ischemia treated with IGF-1 alone, 25 (IQR = 3 to 70) for ischemia treated with hypothermia alone, and 78 (IQR 47.3 to 97.5) for ischemia treated with combination therapy. Days to memory success criteria were 13.6 (IQR = 11.5 to 15.5 days) for sham, 23.5 (IQR = 20 to 25.5 days) for untreated ischemia, 17.5 (IQR = 15.5 to 25.5 days) for ischemia treated with IGF-1, 15 (IQR = 14.5 to 21 days) for ischemia treated with hypothermia, and 13.5 (IQR = 12.25 to 18.5 days) for ischemia treated with combination therapy. Neurologic outcome indices were 8.5 (IQR = 7.4 to 9.5) for sham, 0.14 (IQR = 0.08 to 0.2) for untreated ischemia, 0.18 (IQR = 0.17 to 4.6) for ischemia treated with IGF-1, 0.7 (IQR = 0.2 to 4.8) for ischemia treated with hypothermia, and 5.7 (IQR = 3.3 to 6.2) for ischemia treated with combination therapy. Statistically significant differences in neuron counts, days to memory test criteria, and outcome index were found between sham and untreated ischemic animals. Of the three treatment regimens, only combination therapy showed a statistically significant difference from the untreated ischemic group for neuronal salvage (p = 0.02), days to criteria (p = 0.043), and outcome index (p = 0.014). CONCLUSIONS: Combination therapy with IGF-1 (0.6 U/kg) and therapeutic hypothermia (32°C for 4 hours) at the onset of reperfusion synergistically preserves CA1 structure and function at 28 days after 8 minutes of global brain ischemia in healthy male rats.
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Hipotermia Induzida , Fator de Crescimento Insulin-Like I/uso terapêutico , Ataque Isquêmico Transitório/terapia , Animais , Terapia Combinada , Modelos Animais de Doenças , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Cardiomyocytes and neurons are highly susceptible to ischemia-reperfusion injury; accordingly, considerable effort has been devoted to elucidating the cellular mechanisms responsible for ischemia-reperfusion-induced cell death and developing novel strategies to minimize ischemia-reperfusion injury. Maintenance of mitochondrial integrity is, without question, a critical determinant of cell fate. However, there is emerging evidence of a novel and intriguing extension to this paradigm: mitochondrial dynamics (that is, changes in mitochondrial morphology achieved by fission and fusion) may play an important but as-yet poorly understood role as a determinant of cell viability. Focusing on heart and brain, our aims in this review are to provide a synopsis of the molecular mechanisms of fission and fusion, summarize our current understanding of the complex relationships between mitochondrial dynamics and the pathogenesis of ischemia-reperfusion injury, and speculate on the possibility that targeted manipulation of mitochondrial dynamics may be exploited for the design of novel therapeutic strategies for cardio- and neuroprotection.
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Mitocôndrias Cardíacas/fisiologia , Traumatismo por Reperfusão/fisiopatologia , HumanosRESUMO
Stroke and circulatory arrest cause interferences in blood flow to the brain that result in considerable tissue damage. The primary method to reduce or prevent neurologic damage to patients suffering from brain ischemia is prompt restoration of blood flow to the ischemic tissue. However, paradoxically, restoration of blood flow causes additional damage and exacerbates neurocognitive deficits among patients who suffer a brain ischemic event. Mitochondria play a critical role in reperfusion injury by producing excessive reactive oxygen species (ROS) thereby damaging cellular components, and initiating cell death. In this review, we summarize our current understanding of the mechanisms of mitochondrial ROS generation during reperfusion, and specifically, the role the mitochondrial membrane potential plays in the pathology of cerebral ischemia/reperfusion. Additionally, we propose a temporal model of ROS generation in which posttranslational modifications of key oxidative phosphorylation (OxPhos) proteins caused by ischemia induce a hyperactive state upon reintroduction of oxygen. Hyperactive OxPhos generates high mitochondrial membrane potentials, a condition known to generate excessive ROS. Such a state would lead to a "burst" of ROS upon reperfusion, thereby causing structural and functional damage to the mitochondria and inducing cell death signaling that eventually culminate in tissue damage. Finally, we propose that strategies aimed at modulating this maladaptive hyperpolarization of the mitochondrial membrane potential may be a novel therapeutic intervention and present specific studies demonstrating the cytoprotective effect of this treatment modality.
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Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Potencial da Membrana Mitocondrial , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Animais , Citoproteção , Humanos , Modelos BiológicosRESUMO
The objective of the study is to reveal the seasonal variations in the river water and sediment quality with respect to heavy metal contamination. To get the extent of trace metals contamination, water and sediment samples were collected from five different sites along the course of Sabarmati River and its tributary Kharicut canal in pre-monsoon, monsoon and post-monsoon seasons. The concentration of trace metals such as chromium, copper, lead, nickel and zinc was determined using inductively coupled plasma spectroscopy. The concentrations of heavy metals were found to be higher in the pre-monsoon season than in the monsoon and post-monsoon seasons in water samples. The pollution load index, contamination factor and degree of contamination (C(d)) in sediments were calculated to know the extent of anthropogenic pressures. The values of C(d) clearly indicated very high degree of contamination at Kharicut canal (S-4: 32.25 and S-5: 54.52) and considerable degree of contamination at three sites of Sabarmati river viz; S-1, S-2 and S-3 with values 14.30, 14.42 and 17.21, respectively. Lead and nickel could not be traced in any of the river water samples.
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Sedimentos Geológicos/química , Metais Pesados/análise , Rios/química , Poluentes Químicos da Água/análise , Monitoramento Ambiental , Índia , Estações do Ano , Poluição Química da Água/estatística & dados numéricosRESUMO
The group of antigen 85 proteins of Mycobacterium tuberculosis is responsible for converting trehalose monomycolate to trehalose dimycolate, which contributes to cell wall stability. Here, we have used a serial enrichment approach to identify new potential inhibitors by searching the libraries of compounds using both 2D atom pair descriptors and binary fingerprints followed by molecular docking. Three different docking softwares AutoDock, GOLD, and LigandFit were used for docking calculations. In addition, we applied the criteria of selecting compounds with binding efficiency close to the starting known inhibitor and showing potential to form hydrogen bonds with the active site amino acid residues. The starting inhibitor was ethyl-3-phenoxybenzyl-butylphosphonate, which had IC(50) value of 2.0 µM in mycolyltransferase inhibition assay. Our search from more than 34 million compounds from public libraries yielded 49 compounds. Subsequently, selection was restricted to compounds conforming to the Lipinski rule of five and exhibiting hydrogen bonding to any of the amino acid residues in the active site pocket of all three proteins of antigen 85A, 85B, and 85C. Finally, we selected those ligands which were ranked top in the table with other known decoys in all the docking results. The compound NIH415032 from tuberculosis antimicrobial acquisition and coordinating facility was further examined using molecular dynamics simulations for 10 ns. These results showed that the binding is stable, although some of the hydrogen bond atom pairs varied through the course of simulation. The NIH415032 has antitubercular properties with IC(90) at 20 µg/ml (53.023 µM). These results will be helpful to the medicinal chemists for developing new antitubercular molecules for testing.
Assuntos
Aciltransferases/química , Antígenos de Bactérias/química , Antituberculosos/química , Proteínas de Bactérias/química , Mycobacterium tuberculosis/enzimologia , Aciltransferases/metabolismo , Antígenos de Bactérias/metabolismo , Antituberculosos/metabolismo , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Desenho de Fármacos , Ligação de Hidrogênio , Ligantes , Simulação de Acoplamento MolecularRESUMO
BOD (Biochemical oxygen demand) is the pollution index of any water sample. One of the main factors influencing the estimation of BOD is the nature of microorganisms used as seeding material. In order to meet the variation in wastewater characteristics, one has to be specific in choosing the biological component that is the seeding material. The present study deals with the estimation of BOD of dairy wastewater using a specific microbial consortium and compares of the results with seeding material (BODSEED). Bacterial strains were isolated from 5 different sources and were screened by the conventional BOD method. The selected microbial seed comprises of Enterobacter sp., Pseudomonas sp. BOD : COD (Chemical oxygen demand) ratio using the formulated seed comes in the range of 0.7-0.8 whereas that using BODSEED comes in the ratio of 0.5-0.6. The ultimate BOD (UBOD) was also performed by exceeding the 3-day dilution BOD test. After 90 days, it has been observed that the ratio of BOD : COD increased in case of selected consortium 7 up to 0.91 in comparison to 0.74 by BODSEED. The results were analyzed statistically by t-test and it was observed that selected consortium was more significant than the BODSEED.
Assuntos
Análise da Demanda Biológica de Oxigênio/métodos , Resíduos Industriais/análise , Esgotos/análise , Microbiologia da Água , Animais , Biodegradação Ambiental , Meios de Cultura/metabolismo , Indústria de Laticínios , Enterobacter/isolamento & purificação , Enterobacter/metabolismo , Indústria Alimentícia/métodos , Consórcios Microbianos , Leite/metabolismo , Pseudomonas/isolamento & purificação , Pseudomonas/metabolismo , Reprodutibilidade dos Testes , Esgotos/microbiologiaRESUMO
INTRODUCTION AND RATIONALE: The detection of bioavailable phenol is a very important issue in environmental and human hazard assessment. Despite modest developments recently, there is a stern need for development of novel biosensors with high sensitivity for priority phenol pollutants. DmpR (Dimethyl phenol regulatory protein), an NtrC-like regulatory protein for the phenol degradation of Pseudomonas sp. strain CF600, represents an attractive biosensor regimen. Thus, we sought to design a novel biosensor by modifying the phenol detection capacity of DmpR by using mutagenic PCR. METHODS: Binding sites of 'A' domain of DmpR were predicted by LIGSITE, and molecular docking was performed by using GOLD to identify the regions where phenol may interact with DmpR. Total five point mutations, one single at position 42 (Phe-to-Leu), two double at 140 (Asp-to-Glu) and 143 (Gln-to-Leu), and two double at L113M (Leu-to- Met) and D116A (Asp-to- Ala) were created in DmpR by site-directed mutagenesis to construct the reporter plasmids pRLuc42R, pRLuc140p143R, and pRLuc113p116R, respectively. Luciferase assays were performed to measure the activity of luc gene in the presence of phenol and its derivatives, while RT-PCR was used to check the expression of luc gene in the presence of phenol. RESULTS: Only pRLuc42R and pRLuc113p116R showed positive responses to phenolic effectors. The lowest detectable concentration of phenol was 0.5 µM (0.047 mg/L), 0.1 µM for 2, 4-dimethylphenol and 2-nitrophenol, 10 µM for 2, 4, 6-trichlorophenol and 2-chlorophenol, 100 µM for 2, 4-dichlorophenol, 0.01 µM for 4-nitrophenol, and 1 µM for o-cresol. These concentrations were measured by modified luciferase assay within 3 hrs compared to 6-7 hrs in previous studies. Importantly, increased expression of luciferase gene of pRLuc42R was observed by RT-PCR. CONCLUSIONS: The present study offers an effective strategy to design a quick and sensitive biosensor for phenol by constructing recombinant bacteria having DmpR gene.