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4.
Viruses ; 12(2)2020 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-32059509

RESUMO

Human immunodeficiency virus (HIV)-infected individuals treated with anti-retroviral therapy often develop chronic non-infectious lung disease. To determine the mechanism of HIV-1-associated lung disease we evaluated the dynamics of lung leukocytes in HIV-1 transgenic (Tg) mice with integrated HIV-1 provirus. In HIV-Tg mice, lipopolysacharide (LPS) induced significantly higher levels of neutrophil infiltration in the lungs compared to wild-type (WT) mice. In WT mice, the initial neutrophil infiltration was followed by macrophage infiltration and fast resolution of leukocytes infiltration. In HIV-Tg mice, resolution of lung infiltration by both neutrophils and macrophages was significantly delayed, with macrophages accumulating in the lumen of lung capillaries resulting in a 45% higher rate of mortality. Trans-endothelial migration of HIV-Tg macrophages was significantly reduced in vitro and this reduction correlated with lower HIV-1 gene expression. HIV-1 transcription inhibitor, 1E7-03, enhanced trans-endothelial migration of HIV-Tg macrophages in vitro, decreased lung neutrophil infiltration in vivo, and increased lung macrophage levels in HIV-Tg mice. Moreover, 1E7-03 reduced levels of inflammatory IL-6 cytokine, improved bleeding score and decreased lung injury. Together this indicates that inhibitors of HIV-1 transcription can correct abnormal dynamics of leukocyte infiltration in HIV-Tg, pointing to the utility of transcription inhibition in the treatment of HIV-1 associated chronic lung disease.


Assuntos
Infecções por HIV/fisiopatologia , Indóis/farmacologia , Pulmão/patologia , Infiltração de Neutrófilos/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Ureia/análogos & derivados , Animais , Citocinas/imunologia , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/genética , Inflamação , Lipopolissacarídeos , Pulmão/imunologia , Pulmão/virologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Transgênicos , Organismos Livres de Patógenos Específicos , Ureia/farmacologia
5.
Retrovirology ; 15(1): 39, 2018 05 23.
Artigo em Inglês | MEDLINE | ID: mdl-29792216

RESUMO

BACKGROUND: HIV-1 transcription activator protein Tat is phosphorylated in vitro by CDK2 and DNA-PK on Ser-16 residue and by PKR on Tat Ser-46 residue. Here we analyzed Tat phosphorylation in cultured cells and its functionality. RESULTS: Mass spectrometry analysis showed primarily Tat Ser-16 phosphorylation in cultured cells. In vitro, CDK2/cyclin E predominantly phosphorylated Tat Ser-16 and PKR-Tat Ser-46. Alanine mutations of either Ser-16 or Ser-46 decreased overall Tat phosphorylation. Phosphorylation of Tat Ser-16 was reduced in cultured cells treated by a small molecule inhibitor of CDK2 and, to a lesser extent, an inhibitor of DNA-PK. Conditional knock-downs of CDK2 and PKR inhibited and induced one round HIV-1 replication respectively. HIV-1 proviral transcription was inhibited by Tat alanine mutants and partially restored by S16E mutation. Pseudotyped HIV-1 with Tat S16E mutation replicated well, and HIV-1 Tat S46E-poorly, but no live viruses were obtained with Tat S16A or Tat S46A mutations. TAR RNA binding was affected by Tat Ser-16 alanine mutation. Binding to cyclin T1 showed decreased binding of all Ser-16 and Ser-46 Tat mutants with S16D and Tat S46D mutationts showing the strongest effect. Molecular modelling and molecular dynamic analysis revealed significant structural changes in Tat/CDK9/cyclin T1 complex with phosphorylated Ser-16 residue, but not with phosphorylated Ser-46 residue. CONCLUSION: Phosphorylation of Tat Ser-16 induces HIV-1 transcription, facilitates binding to TAR RNA and rearranges CDK9/cyclin T1/Tat complex. Thus, phosphorylation of Tat Ser-16 regulates HIV-1 transcription and may serve as target for HIV-1 therapeutics.


Assuntos
Regulação Viral da Expressão Gênica , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/fisiologia , Serina/metabolismo , Transcrição Gênica , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo , Sequência de Aminoácidos , Células Cultivadas , Ciclina T/química , Ciclina T/genética , Ciclina T/metabolismo , Quinase 2 Dependente de Ciclina/química , Quinase 2 Dependente de Ciclina/genética , Quinase 2 Dependente de Ciclina/metabolismo , Quinase 9 Dependente de Ciclina/química , Quinase 9 Dependente de Ciclina/metabolismo , Técnicas de Silenciamento de Genes , Infecções por HIV/genética , Interações Hospedeiro-Patógeno , Humanos , Modelos Biológicos , Modelos Moleculares , Mutação , Fosforilação , Ligação Proteica , Conformação Proteica , RNA Viral , Ubiquitinação , Replicação Viral , eIF-2 Quinase/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/química , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética
7.
Biology (Basel) ; 5(4)2016 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-27918433

RESUMO

Protein phosphatase 1 (PP1), a cellular serine/threonine phosphatase, is targeted to cellular promoters by its major regulatory subunits, PP1 nuclear targeting subunit, nuclear inhibitor of PP1 (NIPP1) and RepoMan. PP1 is also targeted to RNA polymerase II (RNAPII) by NIPP1 where it can dephosphorylate RNAPII and cycle-dependent kinase 9 (CDK9). Here, we show that treatment of cells with a small molecule activator of PP1 increases the abundance of a neuregulin-1 (NRG-1)-derived peptide. NRG-1 mRNA and protein levels were increased in the cells stably or transiently expressing mutant NIPP1 (mNIPP1) that does not bind PP1, but not in the cells expressing NIPP1. Expression of mNIPP1 also activated the NRG-1 promoter in an NF-κB-dependent manner. Analysis of extracts from mNIPP1 expressing cells by glycerol gradient centrifugation showed a redistribution of PP1 and CDK9 between large and small molecular weight complexes, and increased CDK9 Thr-186 phosphorylation. This correlated with the increased CDK9 activity. Further, RNAPII co-precipitated with mNIPP1, and phosphorylation of RNAPII C-terminal domain (CTD) Ser-2 residues was greater in cells expressing mNIPP1. In mNIPP1 expressing cells, okadaic acid, a cell-permeable inhibitor of PP1, did not increase Ser-2 CTD phosphorylation inhibited by flavopiridol, in contrast to the NIPP1 expressing cells, suggesting that PP1 was no longer involved in RNAPII dephosphorylation. Finally, media conditioned with mNIPP1 cells induced the proliferation of wild type 84-31 cells, consistent with a role of neuregulin-1 as a growth promoting factor. Our study indicates that deregulation of PP1/NIPP1 holoenzyme activates NRG-1 expression through RNAPII and CDK9 phosphorylation in a NF-κB dependent manner.

8.
Hum Mol Genet ; 25(20): 4601-4609, 2016 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28173069

RESUMO

Blood erythropoietin (EPO) increases primarily to hypoxia. In sickle cell anaemia (homozygous HBBE6V; HbSS), plasma EPO is elevated due to hemolytic anaemia-related hypoxia. Hydroxyurea treatment reduces haemolysis and anaemia by increasing foetal haemoglobin, which leads to lower hypoxic transcriptional responses in blood mononuclear cells but paradoxically further increases EPO. To investigate this apparent hypoxia-independent EPO regulation, we assessed two sickle cell disease (SCD) cohorts for genetic associations with plasma EPO, by prioritizing 237,079 quantitative trait loci for expression level and/or transcript isoform variations of 12,727 genes derived from SCD blood mononuclear cells. We found an association between the T allele of SNP rs60684937 and increased plasma EPO (n = 567, combined P = 5.5 × 10 − 8 adjusted for haemoglobin and hydroxyurea) and validated it in independent SCD patients (n = 183, P = 0.018). The T allele of rs60684937 was associated with a relatively increased expression of a non-coding transcript of PRKAR1A (cAMP-dependent protein kinase type I-alpha regulatory subunit) in 58 SCD patients (P = 7.9 × 10 − 7) and 58 HapMap Yoruba samples (P = 0.0011). In conclusion, we demonstrate that plasma EPO elevation with hydroxyurea in SCD is independent of hypoxic responses and that genetic variation at SNP rs60684937 may contribute to EPO regulation through a cAMP-dependent protein kinase A pathway.


Assuntos
Anemia Falciforme/metabolismo , Subunidade RIalfa da Proteína Quinase Dependente de AMP Cíclico/genética , Eritropoetina/genética , Regulação da Expressão Gênica , Hidroxiureia/farmacologia , Variantes Farmacogenômicos , Polimorfismo de Nucleotídeo Único , Anemia Falciforme/sangue , Anemia Falciforme/tratamento farmacológico , Anemia Falciforme/genética , Antidrepanocíticos/farmacologia , Antidrepanocíticos/uso terapêutico , Eritropoetina/sangue , Feminino , Perfilação da Expressão Gênica , Estudos de Associação Genética , Humanos , Hidroxiureia/uso terapêutico , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Masculino
9.
Blood Adv ; 1(3): 170-183, 2016 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-28203649

RESUMO

The low incidence of HIV-1 infection in patients with sickle cell disease (SCD) and inhibition of HIV-1 replication in vitro under the conditions of low intracellular iron or heme treatment suggests a potential restriction of HIV-1 infection in SCD. We investigated HIV-1 ex vivo infection of SCD peripheral blood mononuclear cells (PBMCs) and found that HIV-1 replication was inhibited at the level of reverse transcription (RT) and transcription. We observed increased expression of heme and iron-regulated genes, previously shown to inhibit HIV-1, including ferroportin, IKBα, HO-1, p21, and SAM domain and HD domain-containing protein 1 (SAMHD1). HIV-1 inhibition was less pronounced in hepcidin-treated SCD PBMCs and more pronounced in the iron or iron chelators treated, suggesting a key role of iron metabolism. In SCD PBMCs, labile iron levels were reduced and protein levels of ferroportin, HIF-1α, IKBα, and HO-1 were increased. Hemin treatment induced ferroportin expression and inhibited HIV-1 in THP-1 cells, mimicking the HIV-1 inhibition in SCD PBMCs, especially as hepcidin similarly prevented HIV-1 inhibition. In THP-1 cells with knocked down ferroportin, IKBα, or HO-1 genes but not HIF-1α or p21, HIV-1 was not inhibited by hemin. Activity of SAMHD1-regulatory CDK2 was decreased, and SAMHD1 phosphorylation was reduced in SCD PBMCs and hemin-treated THP-1 cells, suggesting SAMHD1-mediated HIV-1 restriction in SCD. Our findings point to ferroportin as a trigger of HIV-1 restriction in SCD settings, linking reduced intracellular iron levels to the inhibition of CDK2 activity, reduction of SAMHD1 phosphorylation, increased IKBα expression, and inhibition of HIV-1 RT and transcription.

10.
Sci Pharm ; 83(3): 535-48, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26839837

RESUMO

Complete eradication of HIV-1 infection is impeded by the existence of latent HIV-1 reservoirs in which the integrated HIV-1 provirus is transcriptionally inactive. Activation of HIV-1 transcription requires the viral Tat protein and host cell factors, including protein phosphatase-1 (PP1). We previously developed a library of small compounds that targeted PP1 and identified a compound, SMAPP1, which induced HIV-1 transcription. However, this compound has a limited bioavailability in vivo and may not be able to reach HIV-1-infected cells and induce HIV-1 transcription in patients. We packaged SMAPP1 in polymeric polyethylene glycol polymethyl methacrylate nanoparticles and analyzed its release and the effect on HIV-1 transcription in a cell culture. SMAPP1 was efficiently packaged in the nanoparticles and released during a 120-hr period. Treatment of the HIV-1-infected cells with the SMAPP1-loaded nanoparticles induced HIV-1 transcription. Thus, nanoparticles loaded with HIV-1-targeting compounds might be useful for future anti-HIV-1 therapeutics.

11.
Antimicrob Agents Chemother ; 58(11): 6558-71, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25155598

RESUMO

HIV-1 transcription is activated by the Tat protein, which recruits CDK9/cyclin T1 to the HIV-1 promoter. CDK9 is phosphorylated by CDK2, which facilitates formation of the high-molecular-weight positive transcription elongation factor b (P-TEFb) complex. We previously showed that chelation of intracellular iron inhibits CDK2 and CDK9 activities and suppresses HIV-1 transcription, but the mechanism of the inhibition was not understood. In the present study, we tested a set of novel iron chelators for the ability to inhibit HIV-1 transcription and elucidated their mechanism of action. Novel phenyl-1-pyridin-2yl-ethanone (PPY)-based iron chelators were synthesized and examined for their effects on cellular iron, HIV-1 inhibition, and cytotoxicity. Activities of CDK2 and CDK9, expression of CDK9-dependent and CDK2-inhibitory mRNAs, NF-κB expression, and HIV-1- and NF-κB-dependent transcription were determined. PPY-based iron chelators significantly inhibited HIV-1, with minimal cytotoxicity, in cultured and primary cells chronically or acutely infected with HIV-1 subtype B, but they had less of an effect on HIV-1 subtype C. Iron chelators upregulated the expression of IκB-α, with increased accumulation of cytoplasmic NF-κB. The iron chelators inhibited CDK2 activity and reduced the amount of CDK9/cyclin T1 in the large P-TEFb complex. Iron chelators reduced HIV-1 Gag and Env mRNA synthesis but had no effect on HIV-1 reverse transcription. In addition, iron chelators moderately inhibited basal HIV-1 transcription, equally affecting HIV-1 and Sp1- or NF-κB-driven transcription. By virtue of their involvement in targeting several key steps in HIV-1 transcription, these novel iron chelators have the potential for the development of new therapeutics for the treatment of HIV-1 infection.


Assuntos
Quinase 2 Dependente de Ciclina/metabolismo , Quinase 9 Dependente de Ciclina/metabolismo , HIV-1/genética , Quinase I-kappa B/biossíntese , Quelantes de Ferro/farmacologia , Linhagem Celular , Sobrevivência Celular , Ciclina A/biossíntese , Ciclina A/genética , Ciclina E/biossíntese , Ciclina E/genética , Ciclina T/biossíntese , Quinase 2 Dependente de Ciclina/antagonistas & inibidores , Quinase 9 Dependente de Ciclina/antagonistas & inibidores , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Inibidor de Quinase Dependente de Ciclina p21/genética , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Células HEK293 , HIV-1/efeitos dos fármacos , Humanos , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , RNA Mensageiro/biossíntese , Transcrição Reversa/efeitos dos fármacos , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Fator de Transcrição RelA/metabolismo , Transcrição Gênica/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Produtos do Gene env do Vírus da Imunodeficiência Humana/biossíntese , Produtos do Gene gag do Vírus da Imunodeficiência Humana/biossíntese
13.
Haematologica ; 98(4): 560-7, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23403324

RESUMO

Mutations of VHL (a negative regulator of hypoxia-inducible factors) have position-dependent distinct cancer phenotypes. Only two known inherited homozygous VHL mutations exist and they cause polycythemia: Chuvash R200W and Croatian H191D. We report a second polycythemic Croatian H191D homozygote distantly related to the first propositus. Three generations of both families were genotyped for analysis of shared ancestry. Biochemical and molecular tests were performed to better define their phenotypes, with an emphasis on a comparison with Chuvash polycythemia. The VHL H191D mutation did not segregate in the family defined by the known common ancestors of the two subjects, suggesting a high prevalence in Croatians, but haplotype analysis indicated an undocumented common ancestor ∼six generations ago as the founder of this mutation. We show that erythropoietin levels in homozygous VHL H191D individuals are higher than in VHL R200W patients of similar ages, and their native erythroid progenitors, unlike Chuvash R200W, are not hypersensitive to erythropoietin. This observation contrasts with a report suggesting that polycythemia in VHL R200W and H191D homozygotes is due to the loss of JAK2 regulation from VHL R200W and H191D binding to SOCS1. In conclusion, our studies further define the hematologic phenotype of VHL H191D and provide additional evidence for phenotypic heterogeneity associated with the positional effects of VHL mutations.


Assuntos
Mutação de Sentido Incorreto , Policitemia/genética , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Sequência de Bases , Proliferação de Células/efeitos dos fármacos , Croácia , Análise Mutacional de DNA , Relação Dose-Resposta a Droga , Células Precursoras Eritroides/citologia , Células Precursoras Eritroides/efeitos dos fármacos , Células Precursoras Eritroides/metabolismo , Eritropoetina/sangue , Eritropoetina/farmacologia , Saúde da Família , Feminino , Expressão Gênica/efeitos dos fármacos , Genótipo , Granulócitos/citologia , Granulócitos/efeitos dos fármacos , Granulócitos/metabolismo , Haplótipos , Homozigoto , Humanos , Masculino , Linhagem , Fenótipo , Policitemia/sangue , Policitemia/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Federação Russa
14.
J Mol Med (Berl) ; 91(1): 59-67, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23015148

RESUMO

In Chuvash polycythemia, a homozygous 598C>T mutation in the von Hippel-Lindau gene (VHL) leads to an R200W substitution in VHL protein, impaired degradation of α-subunits of hypoxia-inducible factor (HIF)-1 and HIF-2, and augmented hypoxic responses during normoxia. Chronic hypoxia of high altitude is associated with decreased serum glucose and insulin concentrations. Other investigators reported that HIF-1 promotes cellular glucose uptake by increased expression of GLUT1 and increased glycolysis by increased expression of enzymes such as PDK. On the other hand, inactivation of Vhl in murine liver leads to hypoglycemia associated with a HIF-2-related decrease in the expression of the gluconeogenic enzyme genes Pepck, G6pc, and Glut2. We therefore hypothesized that glucose concentrations are decreased in individuals with Chuvash polycythemia. We found that 88 Chuvash VHL ( R200W ) homozygotes had lower random glucose and glycosylated hemoglobin A1c levels than 52 Chuvash subjects with wild-type VHL alleles. Serum metabolomics revealed higher glycerol and citrate levels in the VHL ( R200W ) homozygotes. We expanded these observations in VHL ( R200W ) homozygote mice and found that they had lower fasting glucose values and lower glucose excursions than wild-type control mice but no change in fasting insulin concentrations. Hepatic expression of Glut2 and G6pc, but not Pdk2, was decreased, and skeletal muscle expression of Glut1, Pdk1, and Pdk4 was increased. These results suggest that both decreased hepatic gluconeogenesis and increased skeletal uptake and glycolysis contribute to the decreased glucose concentrations. Further study is needed to determine whether pharmacologically manipulating HIF expression might be beneficial for treatment of diabetic patients.


Assuntos
Glicemia/metabolismo , Hemoglobinas Glicadas/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Hipóxia/sangue , Policitemia/sangue , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Adulto , Alelos , Animais , Feminino , Regulação da Expressão Gênica , Genótipo , Proteínas Facilitadoras de Transporte de Glucose/genética , Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Glucose-6-Fosfatase/genética , Glucose-6-Fosfatase/metabolismo , Homozigoto , Humanos , Hipóxia/genética , Hipóxia/fisiopatologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Insulina/sangue , Fígado/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Músculo Esquelético/metabolismo , Mutação , Policitemia/genética , Policitemia/fisiopatologia , Complexo Piruvato Desidrogenase/genética , Complexo Piruvato Desidrogenase/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor Von Hippel-Lindau/genética
15.
Biology (Basel) ; 1(2): 175-195, 2012 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-22934150

RESUMO

A major challenge in studies of human diseases involving macrophages is low yield and heterogeneity of the primary cells and limited ability of these cells for transfections and genetic manipulations. To address this issue, we developed a simple and efficient three steps method for somatic 293T cells reprogramming into monocytes and macrophage-like cells. First, 293T cells were reprogrammed into induced pluripotent stem cells (iPSCs) through a transfection-mediated expression of two factors, Oct-4 and Sox2, resulting in a high yield of iPSC. Second, the obtained iPSC were differentiated into monocytes using IL-3 and M-CSF treatment. And third, monocytes were differentiated into macrophage-like cells in the presence of M-CSF. As an example, we developed HIV-1-resistant macrophage-like cells from 293T cells with knockdown of CDK2, a factor critical for HIV-1 transcription. Our study provides a proof-of-principle approach that can be used to study the role of host cell factors in HIV-1 infection of human macrophages.

16.
Haematologica ; 97(2): 193-200, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21993671

RESUMO

BACKGROUND: Patients with Chuvash polycythemia, (homozygosity for the R200W mutation in the von Hippel Lindau gene (VHL)), have elevated levels of hypoxia inducible factors HIF-1 and HIF-2, often become iron-deficient secondary to phlebotomy, and have elevated estimated pulmonary artery pressure by echocardiography. The objectives of this study were to provide a comprehensive echocardiographic assessment of cardiovascular physiology and to identify clinical, hematologic and cardiovascular risk factors for elevation of tricuspid regurgitation velocity in children and adults with Chuvash polycythemia. DESIGN AND METHODS: This cross-sectional observational study of 120 adult and pediatric VHL(R200W) homozygotes and 31 controls at outpatient facilities in Chuvashia, Russian Federation included echocardiography assessment of pulmonary artery pressure (tricuspid regurgitation velocity), cardiac volume, and systolic and diastolic function, as well as hematologic and clinical parameters. We determined the prevalence and risk factors for elevation of tricuspid regurgitation velocity in this population and its relationship to phlebotomy. RESULTS: The age-adjusted mean ± SE tricuspid regurgitation velocity was higher in VHL(R200W) homozygotes than controls with normal VHL alleles (2.5±0.03 vs. 2.3±0.05 m/sec, P=0.005). The age-adjusted left ventricular diastolic diameter (4.8±0.05 vs. 4.5±0.09 cm, P=0.005) and left atrial diameter (3.4±0.04 vs. 3.2±0.08 cm, P=0.011) were also greater in the VHL(R200W) homozygotes, consistent with increased blood volume, but the elevation in tricuspid regurgitation velocity persisted after adjustment for these variables. Among VHL(R200W) homozygotes, phlebotomy therapy was associated with lower serum ferritin concentration, and low ferritin independently predicted higher tricuspid regurgitation velocity (standardized beta=0.29; P=0.009). CONCLUSIONS: Children and adults with Chuvash polycythemia have higher estimated right ventricular systolic pressure, even after adjustment for echocardiography estimates of blood volume. Lower ferritin concentration, which is associated with phlebotomy, independently predicts higher tricuspid regurgitation velocity (www.clinicaltrials.gov identifier NCT00495638).


Assuntos
Anemia Ferropriva/genética , Hipóxia/genética , Policitemia/genética , Pressão Propulsora Pulmonar/fisiologia , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adolescente , Adulto , Anemia Ferropriva/epidemiologia , Anemia Ferropriva/metabolismo , Estudos de Casos e Controles , Criança , Estudos Transversais , Feminino , Homozigoto , Humanos , Hipóxia/epidemiologia , Hipóxia/metabolismo , Masculino , Pessoa de Meia-Idade , Mutação , Policitemia/epidemiologia , Policitemia/metabolismo , Federação Russa/epidemiologia , Insuficiência da Valva Tricúspide/epidemiologia , Insuficiência da Valva Tricúspide/genética , Insuficiência da Valva Tricúspide/metabolismo , Regulação para Cima/fisiologia , Função Ventricular Esquerda/fisiologia
17.
Blood ; 118(19): 5278-82, 2011 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-21876117

RESUMO

Hypoxia is known to reduce the expression of hepcidin, the master regulator of iron metabolism. However, it is not clear whether this response is primarily related to increased erythropoiesis driven by hypoxically stimulated erythropoietin or to a more direct effect of hypoxia on hepcidin expression. The germline loss-of-function VHL(R200W) mutation is common in Chuvashia, Russia, and also occurs elsewhere. VHL(R200W) homozygotes have elevated hypoxia-inducible factor 1α (HIF-1α) and HIF-2α levels, increased red cell mass, propensity to thrombosis, and early mortality. Ninety VHL(R200W) homozygotes and 52 controls with normal VHL alleles from Chuvashia, Russia, were studied under basal circumstances. In univariate analyses, serum hepcidin concentration was correlated positively with serum ferritin concentration and negatively with homozygosity for VHL(R200W). After adjustment for serum erythropoietin and ferritin concentrations by multiple linear regression, the geometric mean (95% confidence interval of mean) hepcidin concentration was 8.1 (6.3-10.5) ng/mL in VHL(R200W) homozygotes versus 26.9 (18.6-38.0) ng/mL in controls (P < .001). In contrast, a significant independent relationship of serum erythropoietin, hemoglobin, or RBC count with hepcidin was not observed. In conclusion, up-regulation of the hypoxic response leads to decreased expression of hepcidin that may be independent of increased erythropoietin levels and increased RBC counts.


Assuntos
Peptídeos Catiônicos Antimicrobianos/sangue , Peptídeos Catiônicos Antimicrobianos/genética , Mutação em Linhagem Germinativa , Policitemia/sangue , Policitemia/genética , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adulto , Estudos de Casos e Controles , Regulação para Baixo , Eritropoetina/sangue , Feminino , Ferritinas/sangue , Hepcidinas , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Federação Russa
18.
Zhen Ci Yan Jiu ; 36(2): 90-4, 2011 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-21717774

RESUMO

OBJECTIVE: To explore the effect of "Xiusanzhen" [electroacupuncture (EA) of bilateral "Yingxiang" (LI 20) and "Yintang" (EX-HN 3)] on activities of hippocampal muscarinic cholinergic receptor (mAChR) and the involved neural path in Alzheimer Disease(AD)rats. METHODS: Forty Sprague Dawley rats were randomly divided into normal control, model, olfactory nerve severing (ONS)-EA of "Xiusanzhen" (ONS-EA) and EA of "Xiusanzhen" (EA) groups. AD model was established by intra-hippocampal injection (AP: 3.5 mm, L:2 mm, H: -2.8 mm) of Abeta(1-40) starch-like peptide (10 microg/2 microL) under the aid of a microsyringe installed in a brain stereotaxis instrument. For rats of the ONS-EA group, the olfactory nerve was severed by using a surgical knife after drilling a hole on the skull (5 mm anterior to the bregma, L, R: 2 mm). The mAChR density, and its maximum binding capacity (Bmax) and dissociation constant (Kd) of the hippocampus tissue were measured by using radio-ligand binding analysis and Lowry's microamount protein assay. RESULTS: In comparison with the normal control group, the hippocampal mAChR density and its Bmax in the model group were decreased remarkably (P < 0.05), while the Kd of M-receptor in the model group was increased remarkably (P < 0.05). In comparison with the model group, hippocampal mAChR density and its Bmax in the EA group were up-regulated obviously (P < 0.05), while the Kd of mAChR in the EA group was down-regulated significantly (P < 0.05). No significant differences were found between the model and ONS-EA groups in mAChR density and its Bmax and Kd (P > 0.05). CONCLUSION: "Xiusanzhen"-EA can effectively up-regulate hippocampal mAChR density and Bmax and down-regulate Kd of M-receptor of hippocampus in AD rats, which is dependent on the intact olfactory nerve pathway.


Assuntos
Pontos de Acupuntura , Doença de Alzheimer/terapia , Eletroacupuntura , Hipocampo/metabolismo , Receptores Muscarínicos/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Modelos Animais de Doenças , Expressão Gênica , Humanos , Masculino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Receptores Muscarínicos/genética
19.
Haematologica ; 96(9): 1371-4, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21606165

RESUMO

The germ-line loss-of-function VHL(R200W) mutation is common in Chuvashia, Russia and occurs in other parts of the world. VHL(R200W) homozygotes have elevated hypoxia inducible factor (HIF)-1 and HIF-2 levels, increased hemoglobin concentration, propensity to thrombosis and early mortality. Because the mutation persists from an ancient origin, we hypothesized that there is a heterozygote advantage. Thirty-four VHL(R200W) heterozygotes and 44 controls over 35 years of age from Chuvashia, Russia were studied. Anemia was defined as hemoglobin less than 130 g/L in men and less than 120 g/L in women. Mild anemia was present in 15% of VHL(R200W) heterozygotes and 34% of controls without a mutated VHL allele. By multivariate logistic regression, the odds of anemia were reduced an estimated 5.6-fold in the VHL(R200W) heterozygotes compared to controls (95% confidence interval 1.4-22.7; P=0.017). In conclusion, heterozygosity for VHL(R200W) may provide protection from anemia; such protection could explain the persistence of this mutation.


Assuntos
Anemia/genética , Heterozigoto , Mutação/genética , Policitemia/genética , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adulto , Idoso , Anemia/epidemiologia , Feminino , Regulação da Expressão Gênica , Humanos , Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Pessoa de Meia-Idade , Policitemia/metabolismo , Prevalência
20.
Haematologica ; 96(8): 1092-8, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21546502

RESUMO

BACKGROUND: Bone changes are common in sickle cell disease, but the pathogenesis is not fully understood. Tartrate-resistant acid phosphatase (TRACP) type 5b is produced by bone-resorbing osteoclasts. In other forms of hemolytic anemia, increased iron stores are associated with osteoporosis. We hypothesized that transfusional iron overload would be associated with increased osteoclast activity in patients with sickle cell disease. DESIGN AND METHODS: We examined tartrate-resistant acid phosphatase 5b concentrations in patients with sickle cell disease and normal controls of similar age and sex distribution at steady state. Serum tartrate-resistant acid phosphatase 5b concentration was measured using an immunocapture enzyme assay and plasma concentrations of other cytokines were assayed using the Bio-Plex suspension array system. Tricuspid regurgitation velocity, an indirect measure of systolic pulmonary artery pressure, was determined by echocardiography. RESULTS: Tartrate-resistant acid phosphatase 5b concentrations were higher in 58 adults with sickle cell disease than in 22 controls (medians of 4.4 versus 2.4 U/L, respectively; P=0.0001). Among the patients with sickle cell disease, tartrate-resistant acid phosphatase 5b independently correlated with blood urea nitrogen (standardized beta=0.40, P=0.003), interleukin-8 (standardized beta=0.30, P=0.020), and chemokine C-C motif ligand 5 (standardized beta=-0.28, P=0.031) concentrations, but not with serum ferritin concentration. Frequent blood transfusions (>10 units in life time) were not associated with higher tartrate-resistant acid phosphatase 5b levels in multivariate analysis. There were strong correlations among tartrate-resistant acid phosphatase 5b, alkaline phosphatase and tricuspid regurgitation velocity (r>0.35, P<0.001). CONCLUSIONS: Patients with sickle cell disease have increased osteoclast activity as reflected by serum tartrate-resistant acid phosphatase 5b concentrations. Our results may support a potential role of inflammation rather than increased iron stores in stimulating osteoclast activity in sickle cell disease. The positive relationships among tartrate-resistant acid phosphatase 5b, alkaline phosphatase and tricuspid regurgitation velocity raise the possibility of a common pathway in the pulmonary and bone complications of sickle cell disease.


Assuntos
Anemia Falciforme/metabolismo , Osteoclastos/metabolismo , Fosfatase Ácida/sangue , Adulto , Biomarcadores/sangue , Estudos de Casos e Controles , Feminino , Humanos , Isoenzimas/sangue , Masculino , Pessoa de Meia-Idade , Valores de Referência , Fosfatase Ácida Resistente a Tartarato
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