Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 25
Filtrar
1.
ACS Med Chem Lett ; 12(6): 1005-1010, 2021 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-34141085

RESUMO

We report herein the discovery of quinazolindiones as potent and selective tankyrase inhibitors. Elucidation of the structure-activity relationship of the lead compound 1g led to truncated analogues that have good potency in cells, pharmacokinetic (PK) properties, and excellent selectivity. Compound 21 exhibited excellent potencies in cells and proliferation studies, good selectivity, in vitro activities, and an excellent PK profile. Compound 21 also inhibited H292 xenograft tumor growth in nude mice. The synthesis, biological, pharmacokinetic, in vivo efficacy studies, and safety profiles of compounds are presented.

2.
ACS Med Chem Lett ; 10(7): 1081-1085, 2019 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-31312412

RESUMO

Interleukin-1 Receptor-Associated Kinase 4 (IRAK4) is a key mediator of innate immunity. IRAK4 overactivation is linked with several autoimmune diseases. To date, many IRAK4 inhibitors have been developed to block the protein's kinase activity with the most advanced reaching Phase II clinical trials. Nevertheless, several reports suggest kinase activity is not disease-relevant in certain cell types, so removing scaffolding signaling in addition to IRAK4 kinase activity may offer a better therapeutic outcome. Herein, we describe the design and synthesis of an IRAK4 Proteolysis Targeted Chimera (PROTAC). We show that IRAK4 degradation induced by compound 9 leads to the inhibition of multiple cytokines in PBMCs. However, in IL-1ß stimulated human dermal fibroblasts, inhibition of IL-6 and TNF-α release was not observed despite IRAK4 degradation. Nonetheless, the possibility of targeting both IRAK4 kinase and scaffolding function could potentially lead to new therapeutic opportunities to treat autoimmune, inflammatory, and oncological diseases.

3.
J Med Chem ; 60(13): 5455-5471, 2017 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-28591512

RESUMO

The availability of high quality probes for specific protein targets is fundamental to the investigation of their function and their validation as therapeutic targets. We report the utilization of a dedicated chemoproteomic assay platform combining affinity enrichment technology with high-resolution protein mass spectrometry to the discovery of a novel nicotinamide isoster, the tetrazoloquinoxaline 41, a highly potent and selective tankyrase inhibitor. We also describe the use of 41 to investigate the biology of tankyrase, revealing the compound induced growth inhibition of a number of tumor derived cell lines, demonstrating the potential of tankyrase inhibitors in oncology.


Assuntos
Antineoplásicos/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Quinoxalinas/farmacologia , Tanquirases/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Ligantes , Modelos Moleculares , Estrutura Molecular , Quinoxalinas/síntese química , Quinoxalinas/química , Relação Estrutura-Atividade , Tanquirases/metabolismo
4.
Blood Adv ; 1(26): 2553-2562, 2017 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-29296907

RESUMO

Neutropenia is a common consequence of radiation and chemotherapy in cancer patients. The resulting immunocompromised patients become highly susceptible to potentially life-threatening infections. Granulocyte colony-stimulating factor (G-CSF) is known to stimulate neutrophil production and is widely used as a treatment of chemotherapy-induced neutropenia. A small-molecule G-CSF secretagogue without a requirement for refrigerated supply chain would offer a more convenient and cost-effective treatment of chemotherapy-induced neutropenia. Bacterial lipopeptides activate innate immune responses through Toll-like receptor 2 (TLR2) and induce the release of cytokines, including G-CSF, from macrophages, monocytes, and endothelial. Pam2CSK4 is a synthetic lipopeptide that effectively mimics bacterial lipoproteins known to activate TLR2 receptor signaling through the TLR2/6 heterodimer. Substrate-based drug design led to the discovery of GSK3277329, which stimulated the release of G-CSF in activated THP-1 cells, peripheral blood mononuclear cells, and human umbilical vein endothelial cells. When administered subcutaneously to cynomolgus monkeys (Macaca fascicularis), GSK3277329 caused systemic elevation of G-CSF and interleukin-6 (IL-6), but not IL-1ß or tumor necrosis factor α, indicating a selective cytokine-stimulation profile. Repeat daily injections of GSK3277329 in healthy monkeys also raised circulating neutrophils above the normal range over a 1-week treatment period. More importantly, repeated daily injections of GSK3277329 over a 2-week period restored neutrophil loss in monkeys given chemotherapy treatment (cyclophosphamide, Cytoxan). These data demonstrate preclinical in vivo proof of concept that TLR2 agonism can drive both G-CSF induction and subsequent neutrophil elevation in the cynomolgus monkey and could be a therapeutic strategy for the treatment of chemotherapy-induced neutropenia.

6.
PLoS One ; 10(5): e0127083, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25965667

RESUMO

CARMA-BCL10-MALT1 signalosomes play important roles in antigen receptor signaling and other pathways. Previous studies have suggested that as part of this complex, MALT1 functions as both a scaffolding protein to activate NF-κB through recruitment of ubiquitin ligases, and as a protease to cleave and inactivate downstream inhibitory signaling proteins. However, our understanding of the relative importance of these two distinct MALT1 activities has been hampered by a lack of selective MALT1 protease inhibitors with suitable pharmacologic properties. To fully investigate the role of MALT1 protease activity, we generated mice homozygous for a protease-dead mutation in MALT1. We found that some, but not all, MALT1 functions in immune cells were dependent upon its protease activity. Protease-dead mice had defects in the generation of splenic marginal zone and peritoneal B1 B cells. CD4+ and CD8+ T cells displayed decreased T cell receptor-stimulated proliferation and IL-2 production while B cell receptor-stimulated proliferation was partially dependent on protease activity. In dendritic cells, stimulation of cytokine production through the Dectin-1, Dectin-2, and Mincle C-type lectin receptors was also found to be partially dependent upon protease activity. In vivo, protease-dead mice had reduced basal immunoglobulin levels, and showed defective responses to immunization with T-dependent and T-independent antigens. Surprisingly, despite these decreased responses, MALT1 protease-dead mice, but not MALT1 null mice, developed mixed inflammatory cell infiltrates in multiple organs, suggesting MALT1 protease activity plays a role in immune homeostasis. These findings highlight the importance of MALT1 protease activity in multiple immune cell types, and in integrating immune responses in vivo.


Assuntos
Caspases/genética , Caspases/metabolismo , Imunidade Inata , Ativação Linfocitária , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Animais , Linfócitos B/imunologia , Células Dendríticas/imunologia , Técnicas de Introdução de Genes , Inflamação/genética , Inflamação/imunologia , Camundongos , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa , Mutação , Baço/imunologia , Linfócitos T/imunologia
7.
Antimicrob Agents Chemother ; 59(4): 2016-28, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25605356

RESUMO

New agents are urgently needed for the therapeutic treatment of Staphylococcus aureus infections. In that regard, S. aureus RNase RnpA may represent a promising novel dual-function antimicrobial target that participates in two essential cellular processes, RNA degradation and tRNA maturation. Accordingly, we previously used a high-throughput screen to identify small-molecule inhibitors of the RNA-degrading activity of the enzyme and showed that the RnpA inhibitor RNPA1000 is an attractive antimicrobial development candidate. In this study, we used a series of in vitro and cellular assays to characterize a second RnpA inhibitor, RNPA2000, which was identified in our initial screening campaign and is structurally distinct from RNPA1000. In doing so, it was found that S. aureus RnpA does indeed participate in 5'-precursor tRNA processing, as was previously hypothesized. Further, we show that RNPA2000 is a bactericidal agent that inhibits both RnpA-associated RNA degradation and tRNA maturation activities both in vitro and within S. aureus. The compound appears to display specificity for RnpA, as it did not significantly affect the in vitro activities of unrelated bacterial or eukaryotic ribonucleases and did not display measurable human cytotoxicity. Finally, we show that RNPA2000 exhibits antimicrobial activity and inhibits tRNA processing in efflux-deficient Gram-negative pathogens. Taken together, these data support the targeting of RnpA for antimicrobial development purposes, establish that small-molecule inhibitors of both of the functions of the enzyme can be identified, and lend evidence that RnpA inhibitors may have broad-spectrum antimicrobial activities.


Assuntos
Antibacterianos/farmacologia , RNA Bacteriano/efeitos dos fármacos , RNA de Transferência/efeitos dos fármacos , Ribonuclease P/antagonistas & inibidores , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Triagem em Larga Escala , Humanos , Hidrazinas/farmacologia , Testes de Sensibilidade Microbiana , Bibliotecas de Moléculas Pequenas , Tioureia/análogos & derivados , Tioureia/farmacologia , Transcrição Gênica/efeitos dos fármacos
8.
Environ Health ; 13: 95, 2014 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-25404430

RESUMO

BACKGROUND: Mercury vapor poses a known health risk with no clearly established safe level of exposure. Consequently there is debate over whether the level of prolonged exposure to mercury vapor from dental amalgam fillings, combining approximately 50% mercury with other metals, is sufficiently high to represent a risk to health. The objective of our study is to determine if mercury exposure from amalgam fillings is associated with risk of adverse health effects. METHODS: In a large longitudinal non-blind sample of participants from a preventative health program in Calgary, Canada we compared number of amalgam fillings, urine mercury measures and changes in 14 self-reported health symptoms, proposed to be mercury dependent sub-clinical measures of mental and physical health. The likelihood of change over one year in a sample of persons who had their fillings removed was compared to a sample of persons who had not had their fillings removed. We use non-parametric statistical tests to determine if differences in urine mercury were statistically significant between sample groups. Logistic regression models were used to estimate the likelihood of observing symptom improvement or worsening in the sample groups. RESULTS: At baseline, individuals with dental amalgam fillings have double the measured urine mercury compared to a control group of persons who have never had amalgam fillings. Removal of amalgam fillings decreases measured urine mercury to levels in persons without amalgam fillings. Although urine mercury levels in our sample are considered by Health Canada to be too low to pose health risks, removal of amalgam fillings reduced the likelihood of self-reported symptom deterioration and increased the likelihood of symptom improvement in comparison to people who retained their amalgam fillings. CONCLUSIONS: Our findings suggest that mercury exposure from amalgam fillings adversely impact health and therefore are a health risk. The use of safer alternative materials for dental fillings should be encouraged to avoid the increased risk of health deterioration associated with unnecessary exposure to mercury.


Assuntos
Amálgama Dentário/efeitos adversos , Mercúrio/toxicidade , Adulto , Canadá/epidemiologia , Monitoramento Ambiental , Feminino , Nível de Saúde , Humanos , Masculino , Mercúrio/urina , Pessoa de Meia-Idade , Medição de Risco , Autorrelato
9.
Cell ; 159(2): 281-94, 2014 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-25303525

RESUMO

Activity-dependent CREB phosphorylation and gene expression are critical for long-term neuronal plasticity. Local signaling at CaV1 channels triggers these events, but how information is relayed onward to the nucleus remains unclear. Here, we report a mechanism that mediates long-distance communication within cells: a shuttle that transports Ca(2+)/calmodulin from the surface membrane to the nucleus. We show that the shuttle protein is γCaMKII, its phosphorylation at Thr287 by ßCaMKII protects the Ca(2+)/CaM signal, and CaN triggers its nuclear translocation. Both ßCaMKII and CaN act in close proximity to CaV1 channels, supporting their dominance, whereas γCaMKII operates as a carrier, not as a kinase. Upon arrival within the nucleus, Ca(2+)/CaM activates CaMKK and its substrate CaMKIV, the CREB kinase. This mechanism resolves long-standing puzzles about CaM/CaMK-dependent signaling to the nucleus. The significance of the mechanism is emphasized by dysregulation of CaV1, γCaMKII, ßCaMKII, and CaN in multiple neuropsychiatric disorders.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Animais , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Calmodulina/metabolismo , Núcleo Celular/metabolismo , Neurônios/metabolismo , Fosforilação , Ratos Sprague-Dawley , Transcrição Gênica
10.
PLoS One ; 9(5): e96737, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24806487

RESUMO

NOD1 is an intracellular pattern recognition receptor that recognizes diaminopimelic acid (DAP), a peptidoglycan component in gram negative bacteria. Upon ligand binding, NOD1 assembles with receptor-interacting protein (RIP)-2 kinase and initiates a signaling cascade leading to the production of pro-inflammatory cytokines. Increased NOD1 signaling has been associated with a variety of inflammatory disorders suggesting that small-molecule inhibitors of this signaling complex may have therapeutic utility. We utilized a cell-based screening approach with extensive selectivity profiling to search for small molecule inhibitors of the NOD1 signaling pathway. Via this process we identified three distinct chemical series, xanthines (SB711), quinazolininones (GSK223) and aminobenzothiazoles (GSK966) that selectively inhibited iE-DAP-stimulated IL-8 release via the NOD1 signaling pathway. All three of the newly identified compound series failed to block IL-8 secretion in cells following stimulation with ligands for TNF receptor, TLR2 or NOD2 and, in addition, none of the compound series directly inhibited RIP2 kinase activity. Our initial exploration of the structure-activity relationship and physicochemical properties of the three series directed our focus to the quinazolininone biarylsulfonamides (GSK223). Further investigation allowed for the identification of significantly more potent analogs with the largest boost in activity achieved by fluoro to chloro replacement on the central aryl ring. These results indicate that the NOD1 signaling pathway, similarly to activation of NOD2, is amenable to modulation by small molecules that do not target RIP2 kinase. These compounds should prove useful tools to investigate the importance of NOD1 activation in various inflammatory processes and have potential clinical utility in diseases driven by hyperactive NOD1 signaling.


Assuntos
Benzotiazóis/farmacologia , Proteína Adaptadora de Sinalização NOD1/metabolismo , Quinazolinonas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Xantinas/farmacologia , Animais , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Fosforilação , Ligação Proteica , Relação Estrutura-Atividade
11.
Rev. argent. coloproctología ; 25(1): 34-40, mar. 2014. ilus
Artigo em Espanhol | LILACS | ID: lil-752830

RESUMO

En este artículo describimos la vida y obra de Florence Nightingale, que es considerada una de las pioneras en la práctica de la enfermería. Su mayor éxito fue su participación en la guerra de Crimea donde junto a otras 38 enfermeras voluntarias reformaron y limpiaron el hospital, e hicieron caer la tasa de mortalidad desde el 40% al 2%. Por las noches recorría los pabellones a la luz de una lámpara mientras visitaba a los enfermos lo cual le valió el nombre de “La señora de la lámpara”. La reina Victoria le otorgó la Cruz Roja Real y, en 1907, fue la primera mujer condecorada con la Orden al Mérito. Ella tuvo sólidos conocimientos en estadística y matemáticas lo cuales les fueron útiles para su labor de enfermera. A 114 años de su fallecimiento, vayan estas líneas como homenaje a la que se considera la precursora de la enfermería moderna de la Argentina y del mundo. Se comenta su influencia indirecta sobre la Escuela de Enfermería del Hospital Británico de Buenos Aires.


We describe Florence Nightingale’s life and works. She is considered a pioneer in nursing practice. Her participation in the Crimean War, together with 38 other voluntary nurses, was mainly in the cleaning and refurbishing of the hospital in Scutari, making the mortality rate to fall from 40% to 2%, this being her greatest success. She used to make her rounds at night in the wards under the light of a lamp, and therefore she was named “The Lady with the Lamp”. Queen Victoria bestowed on her the Royal Red Cross, and she was the first woman to be honored with the Order of Merit in 1907. She had solid knowledge on Statistics and Mathematics, which were useful for her nursing job. A hundred years after her death, let these words be a homage to the Nursing profession, and to who is considered a precursor of modern nursing in Argentina and in the whole world.


Assuntos
Escolas de Enfermagem/história , História da Enfermagem , Papel do Profissional de Enfermagem , Argentina , História da Medicina , Hospitais Privados
12.
PLoS One ; 8(8): e69619, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23936340

RESUMO

NOD2 is an intracellular pattern recognition receptor that assembles with receptor-interacting protein (RIP)-2 kinase in response to the presence of bacterial muramyl dipeptide (MDP) in the host cell cytoplasm, thereby inducing signals leading to the production of pro-inflammatory cytokines. The dysregulation of NOD2 signaling has been associated with various inflammatory disorders suggesting that small-molecule inhibitors of this signaling complex may have therapeutic utility. To identify inhibitors of the NOD2 signaling pathway, we utilized a cell-based screening approach and identified a benzimidazole diamide compound designated GSK669 that selectively inhibited an MDP-stimulated, NOD2-mediated IL-8 response without directly inhibiting RIP2 kinase activity. Moreover, GSK669 failed to inhibit cytokine production in response to the activation of Toll-like receptor (TLR)-2, tumor necrosis factor receptor (TNFR)-1 and closely related NOD1, all of which share common downstream components with the NOD2 signaling pathway. While the inhibitors blocked MDP-induced NOD2 responses, they failed to block signaling induced by NOD2 over-expression or single stranded RNA, suggesting specificity for the MDP-induced signaling complex and activator-dependent differences in NOD2 signaling. Investigation of structure-activity relationship allowed the identification of more potent analogs that maintained NOD2 selectivity. The largest boost in activity was achieved by N-methylation of the C2-ethyl amide group. These findings demonstrate that the NOD2 signaling pathway is amenable to modulation by small molecules that do not target RIP2 kinase activity. The compounds we identified should prove useful tools to investigate the importance of NOD2 in various inflammatory processes and may have potential clinical utility.


Assuntos
Amidas/química , Benzimidazóis/química , Benzimidazóis/farmacologia , Proteína Adaptadora de Sinalização NOD2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Citocinas/metabolismo , Células HEK293 , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , NF-kappa B/metabolismo , Relação Estrutura-Atividade , Receptor 2 Toll-Like/metabolismo
13.
FEBS Lett ; 587(6): 756-62, 2013 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-23395614

RESUMO

Recent studies suggest a link between mitochondria and proinflammatory cytokine generation. We previously demonstrated that overexpression of mitochondrial chaperone glucose-regulated protein75 (Grp75/mortalin) protects mitochondria. In this study we investigated the modulation of the lipopolisaccharide (LPS)-induced inflammatory response of microglial BV-2 cells by Grp75. We demonstrate that LPS-induced activation promotes significant metabolic changes suppressing mitochondrial function and increasing glycolysis. Overexpression of Grp75 attenuates the LPS-induced oxidative and metabolic responses, and suppresses proinflammatory activation, which depends on both NF-κB activation and lactate. Thus overexpression of Grp75 provides a novel strategy to modulate proinflammatory cytokine production of relevance to inflammation-associated pathologies.


Assuntos
Proteínas de Choque Térmico HSP70/genética , Proteínas de Membrana/genética , Microglia/metabolismo , Mitocôndrias/metabolismo , Chaperonas Moleculares/genética , Animais , Células Cultivadas , Citocinas/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Glicólise/genética , Proteínas de Choque Térmico HSP70/metabolismo , Inflamação/metabolismo , Inflamação/patologia , Ácido Láctico/metabolismo , Lipopolissacarídeos/farmacologia , Proteínas de Membrana/metabolismo , Camundongos , Microglia/efeitos dos fármacos , Microglia/patologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Chaperonas Moleculares/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Plasmídeos , Transfecção
14.
Thromb J ; 10(1): 7, 2012 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-22607090

RESUMO

BACKGROUND: DVT is the main cause of death in hospitalized patients and thromboprophylaxis is the only way to prevent these deaths. International recommendations suggested that active monitoring of DVT/PE prophylaxis can improve the efficacy in Hospitals. METHODS: We performed a cohort study in three consecutives periods to evaluate DVT prophylaxis in 388 adults hospitalized in a General Hospital. RESULTS: 85% of the population had high risk factors for DVT. Thromboprophylaxis was in accordance with local and International guidelines (ACCP 2008) in 72.7% and 86% of the patients respectively. No significant difference could be founded between clinical and surgical patients. One every 10 patients received higher prophylaxis than suggested by guidelines and two out of ten received deficient or no prophylaxis. The worst 2 groups of patients were those with moderate/low risk of DVT and the group with a contraindication to pharmacologic prophylaxis. We observed a progressive improvement of the DVT prophylaxis in the 3 periods of evaluation. CONCLUSIONS: Although the rate of recommended thromboprophylaxis is higher than many other reports in the region we still have some areas where we need to improve. Regular audits like these are very helpful to find out what specific areas of the hospital needs some careful attention in order to have a better quality of assistance.

15.
Rev Med Chil ; 140(9): 1218-27, 2012 Sep.
Artigo em Espanhol | MEDLINE | ID: mdl-23354647

RESUMO

BACKGROUND: William Osler was generally regarded as the greatest and most respected physician of his time. This paper describes Osler's life, his philosophy and views. He was an outstanding clinician who emphasized bedside teaching and observation. He possessed an extraordinary charm that inspired many others. As Professor of Medicine at four institutions in three countries, he was a great influence on medical education. He was a prolific writer, and his textbook became the most popular and widely read treatise on medicine in the world. He also was a medical historian, a classical scholar, and an avid bibliophile. He emphasized the value of hard work and ongoing education. His compassion and concern for patients and colleagues reflected his personality. We summarize Osler's descriptions, and some of his aphorisms. His wisdom is as relevant now, as it was in his time. Osler blended the art and science of Medicine perhaps better than anyone else, and remains a valuable role model for students and physicians more than ninety two years after his death.


Assuntos
Educação Médica/história , Aforismos e Provérbios como Assunto , Baltimore , Canadá , Inglaterra , História do Século XIX , História do Século XX , Philadelphia , Filatelia
16.
BMC Bioinformatics ; 12: 276, 2011 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-21729324

RESUMO

BACKGROUND: Activation of the NF-κB transcription factor and its associated gene expression in microglia is a key component in the response to brain injury. Its activation is dynamic and is part of a network of biochemical species with multiple feedback regulatory mechanisms. Mathematical modeling, which has been instrumental for understanding the NF-κB response in other cell types, offers a valuable tool to investigate the regulation of NF-κB activation in microglia at a systems level. RESULTS: We quantify the dynamic response of NF-κB activation and activation of the upstream kinase IKK using ELISA measurements of a microglial cell line following treatment with the pro-inflammatory cytokine TNFα. A new mathematical model is developed based on these data sets using a modular procedure that exploits the feedback structure of the network. We show that the new model requires previously unmodeled dynamics involved in the stimulus-induced degradation of the inhibitor IκBα in order to properly describe microglial NF-κB activation in a statistically consistent manner. This suggests a more prominent role for the ubiquitin-proteasome system in regulating the activation of NF-κB to inflammatory stimuli. We also find that the introduction of nonlinearities in the kinetics of IKK activation and inactivation is essential for proper characterization of transient IKK activity and corresponds to known biological mechanisms. Numerical analyses of the model highlight key regulators of the microglial NF-κB response, as well as those governing IKK activation. Results illustrate the dynamic regulatory mechanisms and the robust yet fragile nature of the negative feedback regulated network. CONCLUSIONS: We have developed a new mathematical model that incorporates previously unmodeled dynamics to characterize the dynamic response of the NF-κB signaling network in microglia. This model is the first of its kind for microglia and provides a tool for the quantitative, systems level study the dynamic cellular response to inflammatory stimuli.


Assuntos
Isquemia Encefálica/metabolismo , Microglia/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Animais , Isquemia Encefálica/patologia , Linhagem Celular , Citocinas/metabolismo , Regulação da Expressão Gênica , Quinase I-kappa B/metabolismo , Inflamação/metabolismo , Camundongos , NF-kappa B/antagonistas & inibidores , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
17.
Mitochondrion ; 11(2): 279-86, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21047562

RESUMO

Ca(2+) transfer from endoplasmic reticulum (ER) to mitochondria at contact sites between the organelles can induce mitochondrial dysfunction and programmed cell death after stress. The ER-localized chaperone glucose-regulated protein 78kDa (GRP78/BiP) protects neurons against excitotoxicity and apoptosis. Here we show that overexpressing GRP78 protects astrocytes against ischemic injury, reduces net flux of Ca(2+) from ER to mitochondria, increases Ca(2+) uptake capacity in isolated mitochondria, reduces free radical production, and preserves respiratory activity and mitochondrial membrane potential after stress. We conclude that GRP78 influences ER-mitochondrial Ca(2+) crosstalk to maintain mitochondrial function and protect astrocytes from ischemic injury.


Assuntos
Astrócitos/metabolismo , Cálcio/metabolismo , Proteínas de Choque Térmico/metabolismo , Isquemia/metabolismo , Mitocôndrias/metabolismo , Animais , Células Cultivadas , Retículo Endoplasmático/metabolismo , Chaperona BiP do Retículo Endoplasmático , Imuno-Histoquímica , Isquemia/fisiopatologia , Potenciais da Membrana , Camundongos , Mitocôndrias/fisiologia , Espécies Reativas de Oxigênio/metabolismo
18.
J Neurosci ; 30(37): 12242-51, 2010 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-20844120

RESUMO

The impairment of hippocampal neurogenesis has been linked to the pathogenesis of neurological disorders from chronic neurodegenerative disease to the progressive cognitive impairment of children who receive brain irradiation. Numerous studies provide evidence that inflammation downregulates neurogenesis, with multiple factors contributing to this impairment. Although mitochondria are one of the primary targets of inflammatory injury, the role of mitochondrial function in the modulation of neurogenesis remains relatively unstudied. In this study, we used neurosphere-derived cells to show that immature doublecortin (Dcx)-positive neurons are uniquely sensitive to mitochondrial inhibition, demonstrating rapid loss of mitochondrial potential and cell viability compared with glial cells and more mature neurons. Mitochondrial inhibition for 24 h produced no significant changes in astrocyte or oligodendrocyte viability, but reduced viability of mature neurons by 30%, and reduced survival of Dcx(+) cells by 60%. We demonstrate that protection of mitochondrial function with mitochondrial metabolites or the mitochondrial chaperone mtHsp75/mortalin partially reverses the inflammation-associated impairment of neurogenesis in vitro and in irradiated mice in vivo. Our findings highlight mitochondrial mechanisms involved in neurogenesis and indicate mitochondria as a potential target for protective strategies to prevent the impairment of neurogenesis by inflammation.


Assuntos
Metabolismo Energético/fisiologia , Mediadores da Inflamação/fisiologia , Mitocôndrias/patologia , Inibição Neural/fisiologia , Neurogênese/fisiologia , Animais , Diferenciação Celular/genética , Linhagem Celular , Células Cultivadas , Proteínas do Domínio Duplacortina , Proteína Duplacortina , Metabolismo Energético/genética , Proteínas de Choque Térmico HSP70/biossíntese , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/fisiologia , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Associadas aos Microtúbulos/antagonistas & inibidores , Proteínas Associadas aos Microtúbulos/biossíntese , Mitocôndrias/genética , Mitocôndrias/fisiologia , Inibição Neural/genética , Neurogênese/genética , Neurônios/metabolismo , Neurônios/patologia , Neuropeptídeos/antagonistas & inibidores , Neuropeptídeos/biossíntese , Tiamina/administração & dosagem
19.
Glia ; 58(9): 1042-9, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20235222

RESUMO

Brief forebrain ischemia is a model of the delayed hippocampal neuronal loss seen in patients following cardiac arrest and resuscitation. Previous studies demonstrated that selective dysfunction of hippocampal CA1 subregion astrocytes occurs hours to days before delayed neuronal death. In this study we tested the strategy of directing protection to astrocytes to protect neighboring neurons from forebrain ischemia. Two well-studied protective proteins, heat shock protein 72 (Hsp72) or superoxide dismutase 2 (SOD2), were genetically targeted for expression in astrocytes using the astrocyte-specific human glial fibrillary acidic protein (GFAP) promoter. The expression constructs were injected stereotacticly immediately above the hippocampal CA1 region on one side of the rat brain two days prior to forebrain ischemia. Cell type specific expression was confirmed by double label immunohistochemistry. When the expression constructs were injected two days before transient forebrain ischemia, the loss of CA1 hippocampal neurons observed seven days later was significantly reduced on the injected side compared with controls. This neuroprotection was associated with significantly better preservation of astrocyte glutamate transporter-1 immunoreactivity at 5-h reperfusion and reduced oxidative stress. Improving the resistance of astrocytes to ischemic stress by targeting either the cytosolic or mitochondrial compartment was thus associated with preservation of CA1 neurons following forebrain ischemia. Targeting astrocytes is a promising strategy for neuronal preservation following cardiac arrest and resuscitation.


Assuntos
Astrócitos/fisiologia , Isquemia Encefálica/fisiopatologia , Região CA1 Hipocampal/fisiopatologia , Proteínas de Choque Térmico HSP72/metabolismo , Neurônios/fisiologia , Superóxido Dismutase/metabolismo , Animais , Região CA1 Hipocampal/patologia , Hipóxia Celular/fisiologia , Células Cultivadas , Transportador 2 de Aminoácido Excitatório/metabolismo , Proteína Glial Fibrilar Ácida/genética , Proteínas de Choque Térmico HSP72/genética , Humanos , Masculino , Camundongos , Neurônios/patologia , Estresse Oxidativo/fisiologia , Regiões Promotoras Genéticas , Prosencéfalo/patologia , Prosencéfalo/fisiopatologia , Ratos , Ratos Sprague-Dawley , Superóxido Dismutase/genética , Transfecção
20.
J Cereb Blood Flow Metab ; 29(2): 365-74, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18985056

RESUMO

Mitochondria are known to be central to the cell's response to ischemia, because of their role in energy generation, in free radical generation, and in the regulation of apoptosis. Heat shock protein 75 (Hsp75/Grp75/mortalin/TRAP1) is a member of the HSP70 chaperone family, which is targeted to mitochondria. Overexpression of Hsp75 was achieved in rat brain by DNA transfection, and expression was observed in both astrocytes and neurons. Rats were subjected to 100 mins middle cerebral artery occlusion followed by assessment of infarct volume, neurological score, mitochondrial function, and levels of oxidative stress at 24 h reperfusion. Overexpression of Hsp75 reduced infarct area from 44.6%+/-21.1% to 25.7%+/-12.1% and improved neurological outcome significantly. This was associated with improved mitochondrial function as shown by protection of complex IV activity, marked reduction of free radical generation detected by hydroethidine fluorescence, reduction of lipid peroxidation detected by 4-hydroxy-2-nonenol immunoreactivity, and increased preservation of ATP levels. This suggests that targeting mitochondria for protection may be a useful strategy to reduce ischemic brain injury.


Assuntos
Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Mitocôndrias/metabolismo , Estresse Oxidativo , Trifosfato de Adenosina/metabolismo , Animais , Astrócitos/metabolismo , Isquemia Encefálica/genética , Transporte de Elétrons , Regulação da Expressão Gênica , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP90/genética , Metabolismo dos Lipídeos , Masculino , Neurônios/metabolismo , Ratos , Ratos Sprague-Dawley
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...