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1.
Med Sci Monit ; 25: 3181-3189, 2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31038126

RESUMO

BACKGROUND Modern medicine has suggested exercise therapy is one of the main treatments for postoperative rehabilitation of tumors. It can influence the recovery of cancer patients by changing the body's material metabolism and energy metabolism. However, studies on metabolic changes of exercise therapy on hepatocellular carcinoma (HCC) patients after surgery are limited. The aim of this study was to explore the effect of aerobic exercise on mice after orthotopic HCC surgery by serum metabolomics test and explore the related mechanism. MATERIAL AND METHODS A total of 60 C57Bl/6 mice were used to establish an orthotopic xenograft model of H22 mouse hepatoma cells. Mice were randomly divided into 6 groups and it was found that the metabolic products of the early postoperative exercise group and sedentary group mainly included L-tryptophan, citric acid, and other energy-related metabolites. RESULTS Energy metabolites, such as succinic acid of the high-intensity exercise group were increased after surgery, whereas phospholipid metabolites, including phosphatidylethanolamine (18: 0/0: 0), were decreased. In the moderate-intensity exercise group, the change tendency was consistent, and the level of various metabolites decreased. CONCLUSIONS Thus, it is likely that aerobic exercise reduced the degree of postoperative stress responses and improved energy metabolism in mice. The underlying mechanism involves improving the tricarboxylic acid cycle, intervening in energy metabolism, reorganization caused by the tumor, reducing the abnormal increase of phospholipase activity caused by the stress of liver cancer, reducing the level of hemolytic phospholipids, thereby inhibiting mitochondrial pathway-initiated apoptosis.


Assuntos
Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/cirurgia , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/cirurgia , Condicionamento Físico Animal/métodos , Alanina Transaminase/sangue , Animais , Apoptose/fisiologia , Aspartato Aminotransferases/sangue , Metabolismo Energético , Globinas/metabolismo , Xenoenxertos , Humanos , Neoplasias Hepáticas/patologia , Masculino , Metabolômica/métodos , Camundongos , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Albumina Sérica/metabolismo
2.
Int Arch Occup Environ Health ; 92(6): 873-881, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30955093

RESUMO

OBJECTIVES: Urinary excretion of 2,5-hexanedione is currently used to estimate the exposure levels of hexane occurring to an individual during the previous work shift. However, because hexane exposures and urinary 2,5-hexanedione levels can vary considerably from day to day, and subchronic to chronic exposures to hexane are required to produce neuropathy, this biomarker may not accurately reflect the risk of an individual for developing hexane neuropathy. This investigation examines the potential of hexane-derived pyrrole adducts produced on globin and plasma proteins as markers for integrating cumulative exposures. Because the pyrrole markers incorporate bioactivation of hexane to 2,5-hexandione and the initial step of protein adduction involved in hexane-induced neuropathy, they potentially can serve as biomarkers of effect through reflecting pathogenetic events within the nervous system. Additionally, pyrrole formation is an irreversible reaction suggesting that hexane-derived protein pyrroles can be used to assess cumulative exposures to provide a better characterization of individual susceptibilities. METHODS: To examine the utility of the proposed markers, blood samples were obtained from eleven workers who used hexane for granulating metal powders in a slurry to produce metal machining die tools and four non-exposed volunteers. Globin and plasma were isolated, and the proteins were digested using pepsin, reacted with Ehrlich's reagent and the level of pyrrole adducts were determined by absorbance at 530 nm. To determine the dose-response curve and dynamic range of the assay, erythrocytes were incubated with a range of 2,5-hexanedione concentrations and the net absorbance at 530 nm of isolated globin was measured. RESULTS: Pyrrole was detected in both the globin and plasma samples of the workers exposed to hexane and the levels of pyrroles in plasma were positively correlated with the levels of pyrroles in globin for most of the workers. CONCLUSIONS: This investigation demonstrates that detectable levels of hexane-derived protein pyrrole adducts are produced on peripheral proteins following occupational exposures to hexane and supports the utility of measuring pyrroles for integrating cumulative exposures to hexane.


Assuntos
Globinas/metabolismo , Hexanos/metabolismo , Plasma/química , Pirróis/sangue , Biomarcadores/sangue , Globinas/química , Humanos , Exposição Ocupacional/efeitos adversos , Pirróis/metabolismo
3.
Mol Biol Evol ; 36(6): 1134-1147, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30828717

RESUMO

As limits on O2 availability during submergence impose severe constraints on aerobic respiration, the oxygen binding globin proteins of marine mammals are expected to have evolved under strong evolutionary pressures during their land-to-sea transition. Here, we address this question for the order Sirenia by retrieving, annotating, and performing detailed selection analyses on the globin repertoire of the extinct Steller's sea cow (Hydrodamalis gigas), dugong (Dugong dugon), and Florida manatee (Trichechus manatus latirostris) in relation to their closest living terrestrial relatives (elephants and hyraxes). These analyses indicate most loci experienced elevated nucleotide substitution rates during their transition to a fully aquatic lifestyle. While most of these genes evolved under neutrality or strong purifying selection, the rate of nonsynonymous/synonymous replacements increased in two genes (Hbz-T1 and Hba-T1) that encode the α-type chains of hemoglobin (Hb) during each stage of life. Notably, the relaxed evolution of Hba-T1 is temporally coupled with the emergence of a chimeric pseudogene (Hba-T2/Hbq-ps) that contributed to the tandemly linked Hba-T1 of stem sirenians via interparalog gene conversion. Functional tests on recombinant Hb proteins from extant and ancestral sirenians further revealed that the molecular remodeling of Hba-T1 coincided with increased Hb-O2 affinity in early sirenians. Available evidence suggests that this trait evolved to maximize O2 extraction from finite lung stores and suppress tissue O2 offloading, thereby facilitating the low metabolic intensities of extant sirenians. In contrast, the derived reduction in Hb-O2 affinity in (sub)Arctic Steller's sea cows is consistent with fueling increased thermogenesis by these once colossal marine herbivores.


Assuntos
Adaptação Biológica , Evolução Molecular , Globinas/genética , Pseudogenes , Sirênios/genética , Animais , Conversão Gênica , Globinas/metabolismo , Masculino , Família Multigênica , Proteínas Mutantes Quiméricas , Oxigênio/metabolismo , Seleção Genética , Sirênios/metabolismo
4.
PLoS One ; 14(2): e0212492, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30768627

RESUMO

BACKGROUND: B-cell lymphoma/leukaemia 11A (BCL11A) is a C2H2-type zinc-finger transcription factor protein that is a critical modulator of haemoglobin switching and suppresses the production of foetal haemoglobin. Variation in the BCL11A gene ameliorates the severity of sickle cell disease (SCD) and ß-thalassemia (ß-thal). The BCL11A gene is located on chromosome 2p16.1 and encodes an 835-amino acid protein. METHOD: Using state-of-the-art in silico tools, this study examined the most pathogenic non-synonymous single nucleotide polymorphisms (nsSNPs) that disrupt the BCL11A protein and mediate foetal-to-adult globin switching. A total of 11,463 SNPs were retrieved from the Single Nucleotide Polymorphism database (dbSNP). These included 799 in the 5' untranslated region (UTR), 486 in the 3' UTR, and 266 non-synonymous, 189 coding synonymous, six nonsense, and six stop-gained SNPs. RESULTS AND DISCUSSION: In silico tools (SIFT, SNAP, PolyPhen-2, PANTHER, I-Mutant, PROVEAN, SNPs&GO, mCSM, and PhD-SNP) predicted the five most-deleterious nsSNPs: rs61742690, rs62142605, rs17028351, rs115666026, and rs74987258. Molecular dynamic simulation and homology modelling of the mutated proteins (S783N, D643N, G451S, K670R, and M313L) of the most deleterious nsSNPs revealed their functional and structural impact. nsSNP rs61742690 was predicted to be the most deleterious, as supported by eight of the nine in silico tools. CONCLUSIONS: Complete failure in the protein-protein interactions with functional partners (KLF1 and others) and significant changes (±100% variation) in the interface energy revealed that rs61742690 (S783N) in the zinc-finger domain is a suitable target for disrupting BCL11A-mediated foetal-to-adult globin switching.


Assuntos
Proteínas de Transporte/genética , Globinas/genética , Proteínas Nucleares/genética , Algoritmos , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Simulação por Computador , Bases de Dados de Ácidos Nucleicos , Hemoglobina Fetal/biossíntese , Hemoglobina Fetal/genética , Globinas/metabolismo , Humanos , Simulação de Dinâmica Molecular , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Polimorfismo de Nucleotídeo Único , Domínios e Motivos de Interação entre Proteínas/genética , Proteínas Repressoras , Homologia de Sequência de Aminoácidos , Dedos de Zinco/genética
5.
Mol Biol Rep ; 46(2): 2101-2110, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30729391

RESUMO

Truncated globins are 20-40 amino acids shorter than full length globins. Till date, globins have been characterized predominantly from bacteria involved in pathogenicity, nitrogen fixation and photosynthesis, where they are implicated in bacterial virulence within the host, protection of nitrogenase from oxygen inactivation and prevention of oxidative damage to the photosynthetic machinery respectively. Myxococcus xanthus, the model myxobacterium, is an obligate aerobe with a multicellular stage in its life cycle where cells encounter oxygen limitation. This work was undertaken to investigate the potential role of the truncated globin in M. xanthus. To examine the role of globins in this unique group of bacteria, the gene coding for a putative truncated globin (HbO) was identified in the genome of M. xanthus DK 1622. The sequence analysis by bioinformatics approaches revealed that HbO from M. xanthus (Mx-HbO) likely adopts a 2-on-2 alpha helical fold of the truncated globins. The gene coding for Mx-HbO was cloned and its expression in E. coli imparted reddish tinge to the cells. The spectral analysis confirmed it to be a functional globin. The expression of Mx-HbO in the heterologous host improved its growth, resulting in the attainment of higher cell density in culture. The transcript of Mx-hbO was induced threefold in the host cells when grown under low aeration condition as compared to the cells grown under high aeration condition. In M. xanthus, an obligate aerobe, where cell growth accompanies swarming, there is a higher density of cells in the middle of the swarm. Our results suggest that Mx-HbO is a functional globin and could facilitate the growth of cells facing oxygen deprivation, the condition prevailing in the middle of the swarm.


Assuntos
Globinas/genética , Myxococcus xanthus/genética , Hemoglobinas Truncadas/genética , Proteínas de Bactérias/metabolismo , Biologia Computacional/métodos , Simulação por Computador , Escherichia coli/genética , Globinas/metabolismo , Myxococcus xanthus/metabolismo , Transcrição Genética/genética , Hemoglobinas Truncadas/metabolismo
6.
Blood ; 133(12): 1358-1370, 2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30700418

RESUMO

Diamond-Blackfan anemia (DBA) is a congenital erythroblastopenia that is characterized by a blockade in erythroid differentiation related to impaired ribosome biogenesis. DBA phenotype and genotype are highly heterogeneous. We have previously identified 2 in vitro erythroid cell growth phenotypes for primary CD34+ cells from DBA patients and following short hairpin RNA knockdown of RPS19, RPL5, and RPL11 expression in normal human CD34+ cells. The haploinsufficient RPS19 in vitro phenotype is less severe than that of 2 other ribosomal protein (RP) mutant genes. We further documented that proteasomal degradation of HSP70, the chaperone of GATA1, is a major contributor to the defect in erythroid proliferation, delayed erythroid differentiation, increased apoptosis, and decreased globin expression, which are all features of the RPL5 or RPL11 DBA phenotype. In the present study, we explored the hypothesis that an imbalance between globin and heme synthesis may be involved in pure red cell aplasia of DBA. We identified disequilibrium between the globin chain and the heme synthesis in erythroid cells of DBA patients. This imbalance led to accumulation of excess free heme and increased reactive oxygen species production that was more pronounced in cells of the RPL5 or RPL11 phenotype. Strikingly, rescue experiments with wild-type HSP70 restored GATA1 expression levels, increased globin synthesis thereby reducing free heme excess and resulting in decreased apoptosis of DBA erythroid cells. These results demonstrate the involvement of heme in DBA pathophysiology and a major role of HSP70 in the control of balanced heme/globin synthesis.


Assuntos
Anemia de Diamond-Blackfan/patologia , Diferenciação Celular , Células Eritroides/patologia , Fator de Transcrição GATA1/metabolismo , Globinas/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Heme/metabolismo , Anemia de Diamond-Blackfan/metabolismo , Proliferação de Células , Células Cultivadas , Células Eritroides/metabolismo , Feminino , Seguimentos , Haploinsuficiência , Humanos , Lactente , Recém-Nascido , Masculino , Mutação , Fenótipo , Prognóstico , RNA Interferente Pequeno , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo
7.
J Chem Inf Model ; 59(1): 441-452, 2019 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-30516994

RESUMO

Proteins are sensitive to temperature, and abrupt changes in the normal temperature conditions can have a profound impact on both structure and function, leading to protein unfolding. However, the adaptation of certain organisms to extreme conditions raises questions about the structural features that permit the structure and function of proteins to be preserved under these adverse conditions. To gain insight into the molecular basis of protein thermostability in the globin family, we have examined three representative examples: human neuroglobin, horse heart myoglobin, and Drosophila hemoglobin, which differ in their melting temperatures and coordination states of the heme iron in the absence of external ligands. In order to elucidate the possible mechanisms that govern the thermostability of these proteins, microsecond-scale classical molecular dynamics simulations were performed at different temperatures. Structural fluctuations and essential dynamics were analyzed, indicating that the flexibility of the CD region, which includes the two short C and D helixes and the connecting CD loop, is directly related to the thermostability. We observed that a larger inherent flexibility of the protein produces higher thermostability, probably concentrating the thermal fluctuations observed at high temperature in flexible regions, preventing unfolding. Globally, the results of this work improve our understanding of thermostability in the globin family.


Assuntos
Globinas/química , Globinas/metabolismo , Heme , Simulação de Dinâmica Molecular , Temperatura Ambiente , Sequência de Aminoácidos , Animais , Conformação Proteica em alfa-Hélice , Dobramento de Proteína , Estabilidade Proteica
8.
PLoS One ; 13(12): e0208465, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30513111

RESUMO

A methodology to cluster proteins based on their dynamics' similarity is presented. For each pair of proteins from a dataset, the structures are superimposed, and the Anisotropic Network Model modes of motions are calculated. The twelve slowest modes from each protein are matched using a local mode alignment algorithm based on the local sequence alignment algorithm of Smith-Waterman. The dynamical similarity distance matrix is calculated based on the top scoring matches of each pair and the proteins are clustered using a hierarchical clustering algorithm. The utility of this method is exemplified on a dataset of protein chains from the globin family and a dataset of tetrameric hemoglobins. The results demonstrate the effect of the quaternary structure of globin members on their intrinsic dynamics and show good ability to distinguish between different states of hemoglobin, revealing the dynamical relations between them.


Assuntos
Globinas/química , Globinas/genética , Família Multigênica , Sequência de Aminoácidos , Animais , Archaea/classificação , Archaea/genética , Conjuntos de Dados como Assunto , Globinas/metabolismo , Hemoglobinas/química , Hemoglobinas/genética , Humanos , Methanosarcina/classificação , Methanosarcina/genética , Modelos Moleculares , Filogenia , Porcos-Espinhos , Conformação Proteica , Multimerização Proteica , Estrutura Quaternária de Proteína , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos
9.
Vascul Pharmacol ; 110: 7-15, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29969687

RESUMO

Cytoglobin is a widely expressed heme protein that binds oxygen, carbon monoxide and nitric oxide. Recent examination of cytoglobin in the vasculature indicates that it contributes to nitric oxide availability, which is central to normal blood vessel function through regulation of smooth muscle cell tone and physiological response. Given the potential implications of cytoglobin in vascular function, we examined how cytoglobin might be uniquely regulated in vascular smooth muscle cells. Our data demonstrate that endothelial cells can increase the expression of cytoglobin in vascular smooth muscle cells, and the induction of cytoglobin is cell contact-dependent. We show that Notch signaling is necessary for endothelial cell-induced cytoglobin expression and Notch2 and Notch3 are sufficient to drive its expression in aortic smooth muscle cells. We further reveal that in cytoglobin-depleted smooth muscle cells there is increased cellular nitric oxide. These data demonstrate that, in addition to being the main producer of vascular nitric oxide, endothelial cells facilitate the ability of smooth muscle cells to metabolize nitric oxide through upregulation of cytoglobin. Our results reveal a novel mechanism by which Notch signaling contributes to vascular function through regulation of a gene that controls nitric oxide levels.


Assuntos
Comunicação Celular , Células Endoteliais/metabolismo , Globinas/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Óxido Nítrico/metabolismo , Animais , Células Cultivadas , Técnicas de Cocultura , Citoglobina , Globinas/genética , Humanos , Camundongos Endogâmicos C57BL , Receptor Notch2/metabolismo , Receptor Notch3/metabolismo , Transdução de Sinais , Regulação para Cima
10.
Int J Mol Sci ; 19(7)2018 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-30041403

RESUMO

Neuroglobin is an endogenous neuroprotective protein, but the underlying neuroprotective mechanisms remain to be elucidated. Our previous yeast two-hybrid screening study identified that Dishevelled-1, a key hub protein of Wnt/ß-Catenin signaling, is an interaction partner of Neuroglobin. In this study, we further examined the role of Neuroglobin in regulating Dishevelled-1 and the downstream Wnt/ß-Catenin and NFκB signaling pathway. We found that Neuroglobin directly interacts with Dishevelled-1 by co-immunoprecipitation, and the two proteins are co-localized in both cytoplasma and nucleus of SK-N-SH cells. Moreover, the ectopic expression of Neuroglobin promotes the degradation of exogenous and endogenous Dishevelled-1 through the proteasomal degradation pathway. Furthermore, our results showed that Neuroglobin significantly inhibits the luciferase activity of Topflash reporter and the expression of ß-Catenin mediated by Dishevelled-1 in SK-N-SH cells. In addition, we also documented that Neuroglobin enhances TNF-α-induced NFκB activation via down-regulating Dishevelled-1. Finally, 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) assays showed that Neuroglobin is an important neuroprotectant that protects SK-N-SH cells from TNF-α-induced decrease in cell viability. Taken together, these findings demonstrated that Neuroglobin functions as an important modulator of the Wnt/ß-Catenin and NFκB signaling pathway through regulating Dishevelled-1.


Assuntos
Globinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Via de Sinalização Wnt , Linhagem Celular Tumoral , Proteínas Desgrenhadas/metabolismo , Globinas/genética , Humanos , NF-kappa B/metabolismo , Proteínas do Tecido Nervoso/genética , Neuroglobina , Ligação Proteica , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
11.
Life Sci ; 207: 50-60, 2018 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-29852187

RESUMO

AIM: The effects of diosmin (DS), pentoxifylline (PTX) and their combination on inflammatory response, oxidant/antioxidant balance, cytoglobin and cirrhotic reaction during bile duct ligation (BDL) were investigated and explored. MAIN METHODS: Fifty adult male Wistar albino rats were randomly allocated to five groups as following, sham: received vehicle only, BDL: subjected to common BDL without treatment, BDL plus DS: received 100 mg/kg/day orally, BDL plus PTX: received 50 mg/kg/day orally, BDL plus DS plus PTX: received DS and PTX in the same manner. The test period lasted 28 days, liver tissues and blood samples were collected to investigate biochemical markers (liver function biomarkers, oxidative stress markers, and antifibrotic markers), mRNA expression of Nrf-2, Keap-1, NF-κB-p65 and p38-MAPK by real-time PCR, protein expression of cytoglobin and NF-κB-p65 by western blot and iNOS and eNOS by immunohistochemistry. Histopathological study was performed to confirm our results. KEY FINDINGS: Chronic BDL induced a significant alteration in liver functions, oxidative stress and fibrotic markers. Furthermore, unfavorable effects on gene and protein expression were observed after BDL. Histopathological findings of this group showed parallel effects. DS, PTX and their combination treatment significantly ameliorated the disturbance that occurred due to BDL. Similar findings were observed in liver histopathology. SIGNIFICANCE: DS and PTX could mitigate liver cirrhosis through modulation of Keap-1/Nrf-2/GSH and NF-κB-p65/p38-MAPK signaling pathways. In addition, we demonstrated that the hepatoprotective effect of DS and PTX is mediated by up-regulation of cytoglobin with inhibition of fibrotic reaction.


Assuntos
Diosmina/farmacologia , Globinas/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Cirrose Hepática/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Pentoxifilina/farmacologia , Animais , Citoglobina , Progressão da Doença , Radicais Livres , Perfilação da Expressão Gênica , Inflamação , Fígado/metabolismo , Testes de Função Hepática , Masculino , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Oxidantes/metabolismo , Estresse Oxidativo , Ratos , Ratos Wistar , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo
12.
J Neurochem ; 145(6): 464-473, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29500821

RESUMO

Hemopexin (Hpx) binds heme with extraordinary affinity, and after haptoglobin may provide a second line of defense against the toxicity of extracellular hemoglobin (Hb). In this series of experiments, the hypothesis that Hpx protects neurons from Hb neurotoxicity was evaluated in murine primary cultures containing neurons and glial cells. Contrary to hypothesis, Hpx increased neuronal loss due to micromolar concentrations of Hb by 4- to 12-fold, as measured by LDH release assay; conversely, the neurotoxicity of hemin was completely prevented. The endogenous fluorescence of Hpx was quenched by Hb, consistent with transfer of Hb-bound heme to Hpx. This was associated with precipitation of globin chains, as detected by immunostaining and fluorescent Hb labeling. A portion of this precipitate attached firmly to cells and could not be removed by multiple washes. Concomitant treatment with haptoglobin (Hp) prevented globin precipitation and most of the increase in neuronal loss. Hpx weakly attenuated the increase in culture non-heme iron produced by Hb treatment, quantified by ferrozine assay. However, Hb-Hpx toxicity was iron-dependent, and was blocked by deferoxamine and ferrostatin-1. Up-regulation of cell ferritin expression, a primary cell defense against Hb toxicity, was not observed on western blots of culture lysates that had been concomitantly treated with Hpx. These results suggest that Hpx destabilizes Hb in the absence of haptoglobin, leading to globin precipitation and exacerbation of iron-dependent oxidative cell injury. Combined therapy with hemopexin plus haptoglobin may be preferable to hemopexin alone after CNS hemorrhage.


Assuntos
Haptoglobinas/metabolismo , Hemoglobinas/toxicidade , Hemopexina/toxicidade , Síndromes Neurotóxicas/fisiopatologia , Animais , Antídotos/farmacologia , Cicloexilaminas/farmacologia , Desferroxamina/farmacologia , Feminino , Ferritinas/metabolismo , Globinas/metabolismo , Heme Oxigenase-1/metabolismo , Hemina/toxicidade , Ferro/metabolismo , Masculino , Camundongos , Neuroglia/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ferroproteínas não Heme/metabolismo , Fenilenodiaminas/farmacologia , Gravidez , Cultura Primária de Células
13.
Exp Cell Res ; 366(1): 16-23, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29524391

RESUMO

Hemoglobins (Hbs) are evolutionarily conserved small globular proteins with characteristic 3-over-3 α-helical sandwich structure that is typically known as "globin fold". Hbs have been found to be involved in diverse biological functions and the characteristic property of oxygen transportation is relatively a recent adaptation. Drosophila genome possesses three globin genes (glob1, glob2, and glob3) and it was previously reported that adequate expression of glob1 is required for various aspects of development, and also to regulate the cellular level of reactive oxygen species (ROS). The present study illustrates the explicit role of glob1 gene in Drosophila development. We demonstrate a dynamic expression pattern of glob1 in larval tissues which largely concentrate around F-actin rich structures and also co-precipitate. Reduced expression of glob1 leads to developmental abnormalities which appeared to be largely mediated by inappropriately formed F-actin based cytoskeletal structures. Our subsequent analysis in FLP/FRT mediated somatic clones establishes specific role of Drosophila glob1 in maintenance of the integrity of F-actin based cytoskeleton during development. For the first time, we report interaction between Glob1 and actin, and propose a novel role of glob1 in maintenance of F-actin based cytoskeleton in Drosophila.


Assuntos
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Globinas/metabolismo , Animais , Proteínas de Drosophila/metabolismo , Hemoglobinas/metabolismo , Espécies Reativas de Oxigênio/metabolismo
14.
Exp Parasitol ; 185: 29-38, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29309785

RESUMO

Understanding the pathophysiology and associated host parasite interactions of the malaria infection is the prerequisite for developing effective prevention and treatment strategies. The exact mechanism underlying malaria associated ineffective and dyserythropoiesis is not yet fully understood. Being an important protein, haemoglobin serves as the main amino acid reservoir available to the intra-erythrocytic plasmodium. It is important to check the expression profiling of globin genes which may help us to understand host parasite interactions and its potential contribution to both infection and disease. Here, an in-vitro culture system was used to study the effect of different doses of Plasmodium falciparum on haematopoietic stem cell expansion, differentiation and expression of globin genes. Upon exposure to the different doses of P. falciparum parasites of strains 3D7, Dd2 and RKL9 (intact and lysed form) at different stages of erythroid development, cells demonstrated suppression in growth and differentiation. At almost all stages of erythroid development upon parasite exposure, the γ globin gene was found to be downregulated and the α/ß as well as α/non- α globin mRNA ratios in late stage erythroid cells were found to be reduced (p < .01) compared to the untreated controls. The imbalance in globin chain expression might be considered as one of the factors involved in malaria associated inappropriate erythropoietic responses.


Assuntos
Anemia/etiologia , Regulação da Expressão Gênica/genética , Globinas/genética , Células-Tronco Hematopoéticas/parasitologia , Malária Falciparum/genética , Anemia/genética , Anemia/metabolismo , Antígenos CD34/sangue , Biomarcadores/metabolismo , Células Cultivadas , Eritrócitos/parasitologia , Eritrócitos/patologia , Células Eritroides/imunologia , Sangue Fetal/citologia , Globinas/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Hemólise , Interações Hospedeiro-Parasita/genética , Humanos , Malária Falciparum/complicações , Reação em Cadeia da Polimerase em Tempo Real
15.
Brain Res ; 1678: 47-55, 2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-29038003

RESUMO

Retina is a critical component of the central nerve system that is responsible for the conversion of light stimulus into electrical spikes. Retinitis pigmentosa (RP) comprises a heterogeneous group of inherited retinal dystrophies leading to blindness. We examined retinal neuroglobin (Ngb) expression in a pharmacologically induced RP animal model, the N-Methyl-N-nitrosourea (MNU) administered mice. The retinal Ngb expression in MNU administered mice attenuated following a time dependent manner, suggesting Ngb was involved in the photoreceptor degeneration. Conversely, the intravenous delivery of Hemin, a Ngb up-regulator, enhanced the Ngb expressions in the retinas of MNU administered mice. Optokinetic behavioral tests and Electroretinogram (ERG) examination suggested that the Hemin treatment could improve the visual function of MNU administered mice. The retinal morphology of the Hemin treated group was much more intact than the MNU group as evidenced by retinal sections and optical coherence tomography (OCT) examinations. Moreover, immunostaining experiments showed the cone photoreceptors in the MNU administered mice were also rescued by Hemin treatment. Furthermore, mechanism studies suggested the Hemin treatment not only alleviated the oxidative stress, but also rectified the apoptotic changes in the retinas of MNU administered mice. In conclusion, the intraperitoneally delivery of Hemin can enhance the Ngb expressions in the MNU administered retinas, thereby ameliorating the photoreceptor degeneration and associated visual impairments. These findings would shed light on the opportunity to develop Ngb into a therapeutic molecular against RP.


Assuntos
Globinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Células Fotorreceptoras/efeitos dos fármacos , Degeneração Retiniana/prevenção & controle , Animais , Modelos Animais de Doenças , Eletrorretinografia/métodos , Globinas/efeitos dos fármacos , Hemina/metabolismo , Metilnitrosoureia/efeitos adversos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/efeitos dos fármacos , Neuroglobina , Estresse Oxidativo , Células Fotorreceptoras/metabolismo , Células Fotorreceptoras de Vertebrados/metabolismo , Retina/metabolismo , Degeneração Retiniana/fisiopatologia
16.
Mol Cell Biochem ; 437(1-2): 133-142, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28620820

RESUMO

Cell proliferation and migration are crucial in many physiological processes including development, cancer, tissue repair, and wound healing. Cell migration is regulated by several signaling molecules. Identification of genes related to cell migration is required to understand molecular mechanism of non-healing chronic wounds which is a major concern in clinics. In the current study, the role of cytoglobin (CYGB) gene in fibroblast cell migration and proliferation was described. L929 mouse fibroblast cells were transduced with lentiviral particles for CYGB and GFP, and analyzed for cell proliferation and migration ability. Fibroblast cells overexpressing CYGB displayed decreased cell proliferation, colony formation capacity, and cell migration. Phosphorylation levels of mTOR and two downstream effectors S6 and 4E-BP1 which take part in PI3K/AKT/mTOR signaling declined in CYGB-overexpressing cells. Microarray analysis indicated that CYGB overexpression leads to downregulation of cell proliferation, migration, and tumor growth associated genes in L929 cell line. This study demonstrated the role of CYGB in fibroblast cell motility and proliferation. CYGB could be a promising candidate for further studies as a potential target for diseases related to cell migration such as cancer and chronic wound treatment.


Assuntos
Movimento Celular , Proliferação de Células , Fibroblastos/metabolismo , Globinas/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Citoglobina , Fibroblastos/citologia , Globinas/genética , Camundongos , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
18.
Neurol Sci ; 39(2): 275-285, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29101592

RESUMO

Neuroglobin (Ngb) is expressed in the central and peripheral nervous system, cerebrospinal fluid, retina, and endocrine tissues where it is involved in binding O2 and other gasotransmitters. Several studies have highlighted its endogenous neuroprotective function. Huntington's disease (HD), a dominant hereditary disease, is characterized by the gradual loss of neurons in discrete areas of the central nervous system. We analyzed the expression of Ngb in the brain tissue of a mouse model of HD, in order to define the role of Ngb with respect to individual cell type vulnerability in HD and to gender and age of mice. Our results showed different expressions of Ngb among neurons of a specific region and between different brain regions. We evidenced a decreased intensity of Ngb at 13 weeks of age, compared to 7 weeks of age. The double immunofluorescence and fluorescence resonance energy transfer (FRET) experiments showed that the co-localization between Ngb and huntingtin at the subcellular level was not close enough to account for a direct interaction. We also observed a different expression of Ngb in the striatum, depending on the sex and age of animals. These findings provide the first experimental evidence for an adaptive response of Ngb in HD, suggesting that Ngb may exert neuroprotective effects in HD beyond its role in reducing sensitivity to oxidative stress.


Assuntos
Corpo Estriado/metabolismo , Regulação da Expressão Gênica/genética , Globinas/metabolismo , Doença de Huntington/patologia , Proteínas do Tecido Nervoso/metabolismo , Fatores de Ribosilação do ADP , Animais , Toxinas Bacterianas , Linhagem Celular Tumoral , Colinesterases/metabolismo , Corpo Estriado/patologia , Modelos Animais de Doenças , Feminino , Transferência Ressonante de Energia de Fluorescência , Proteína Huntingtina/genética , Doença de Huntington/genética , Masculino , Camundongos , Camundongos Transgênicos , Mutação/genética , Neuroglobina , Neurônios/metabolismo , Parvalbuminas/metabolismo , Fatores Sexuais , Fatores de Tempo
19.
Nitric Oxide ; 73: 39-51, 2018 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-29275194

RESUMO

Despite the large number of globins recently discovered in bacteria, our knowledge of their physiological functions is restricted to only a few examples. In the microbial world, globins appear to perform multiple roles in addition to the reversible binding of oxygen; all these functions are attributable to the heme pocket that dominates functional properties. Resistance to nitrosative stress and involvement in oxygen chemistry seem to be the most prevalent functions for bacterial globins, although the number of globins for which functional roles have been studied via mutation and genetic complementation is very limited. The acquisition of structural information has considerably outpaced the physiological and molecular characterisation of these proteins. The genome of the Antarctic cold-adapted bacterium Pseudoalteromonas haloplanktis TAC125 (PhTAC125) contains genes encoding three distinct single-chain 2/2 globins, supporting the hypothesis of their crucial involvement in a number of functions, including protection against oxidative and nitrosative stress in the cold and O2-rich environment. In the genome of PhTAC125, the genes encoding 2/2 globins are constitutively transcribed, thus suggesting that these globins are not functionally redundant in their physiological function in PhTAC125. In the present study, the physiological role of one of the 2/2 globins, Ph-2/2HbO-2217, was investigated by integrating in vivo and in vitro results. This role includes the involvement in the detoxification of reactive nitrogen and O2 species including NO by developing two in vivo and in vitro models to highlight the protective role of Ph-2/2HbO-2217 against reactive nitrogen species. The PSHAa2217 gene was cloned and over-expressed in the flavohemoglobin-deficient mutant of Escherichia coli and the growth properties and O2 uptake in the presence of NO of the mutant carrying the PSHAa2217 gene were analysed. The ferric form of Ph-2/2HbO-2217 is able to catalyse peroxynitrite isomerisation in vitro, indicating its potential role in the scavenging of reactive nitrogen species. Here we present in vitro evidence for the detoxification of NO by Ph-2/2HbO-2217.


Assuntos
Proteínas de Bactérias/genética , Globinas/genética , Estresse Nitrosativo/genética , Pseudoalteromonas/genética , Regiões Antárticas , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Clonagem Molecular , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Genoma Bacteriano , Globinas/química , Globinas/metabolismo , Heme/química , Heme/metabolismo , Inativação Metabólica/genética , Isomerismo , Óxido Nítrico/metabolismo , Óxido Nítrico/toxicidade , Ácido Peroxinitroso/metabolismo , Pseudoalteromonas/fisiologia , S-Nitrosoglutationa/farmacologia
20.
PLoS One ; 12(12): e0189179, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29216269

RESUMO

Environmental factors or adverse growth conditions that may reduce cell function or viability are considered stress. The cell ability to sense and respond to environmental stresses determine its function and survival destiny. We recently defined Neuroglobin (NGB), a heme-protein, as a compensatory protein in the 17ß-Estradiol (E2) anti-apoptotic activity and as a sensor of oxidative stress in both neurons and breast cancer cells. Here, the possibility that NGB levels could represent a pivotal regulator of integrated response of cancer cells to stress has been evaluated. Data obtained in neuroblastoma and in breast cancer cell lines evidence that nutrient deprivation significantly up-regulated NGB levels at different time points. However, the analysis of autophagy activation led to exclude any possible role of stress- or E2-induced NGB in the upstream regulation of general autophagy. However, the over-expression of Flag-NGB in ERα stable transfected HEK-293 cells completely affects nutrient deprivation-induced decrease in cell number. In addition, reported results indicate that modulation of the anti-apoptotic Bcl-2 level may play a key role in the protective NGB function against energetic stress. Overall, these data define a role of NGB as compensatory protein in the cell machinery activated in response to stress and as general stress adaptation marker of cancer cells susceptible to oxidative stress, oxygen and, as demonstrated here for the first time, even to nutrient willingness. Despite the lacking of any direct NGB role on autophagic flux activated by energetic stress, NGB upregulation appears functional in delaying stress-related cell death allowing an appropriate cell response and adaptation to the changing extracellular conditions.


Assuntos
Neoplasias da Mama/patologia , Globinas/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Neuroblastoma/patologia , Neurônios/patologia , Autofagia , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Meios de Cultura , Globinas/metabolismo , Células HEK293 , Humanos , Proteínas do Tecido Nervoso/metabolismo , Neuroblastoma/metabolismo , Neuroglobina , Neurônios/metabolismo , Estresse Oxidativo
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