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2.
Am J Surg ; 208(4): 556-62, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25241952

RESUMO

BACKGROUND: Limited data are available on outcome implications of clopidogrel use before kidney transplantation. METHODS: A novel dataset linking national transplant registry data with records from a large pharmacy claims clearinghouse (2005 to 2010) was examined to estimate risks of post-transplant death and graft failure associated with clopidogrel fills within 90 or more than 90 days before transplant. RESULTS: Clopidogrel fills within 90 days of transplant were associated with 61% of increased relative mortality risk and 23% of increased graft failure risk. Risks were higher in those whose last clopidogrel fill was more than 90 days before transplantation (111% for death, 59% for graft loss). Adverse prognostic associations persisted among recipients of live and deceased donor allografts, older recipients, and those with diabetes or reported cardiovascular disease. CONCLUSIONS: Clopidogrel use before kidney transplantation portends increased risks of post-transplant death and graft loss. Pharmacy claims may identify novel prognostic markers not currently captured in the transplant registry.


Assuntos
Rejeição de Enxerto/epidemiologia , Falência Renal Crônica/cirurgia , Transplante de Rim/mortalidade , Sistema de Registros , Medição de Risco/métodos , Ticlopidina/análogos & derivados , Doadores de Tecidos/estatística & dados numéricos , Doenças Cardiovasculares/complicações , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/epidemiologia , Clopidogrel , Relação Dose-Resposta a Droga , Feminino , Seguimentos , Rejeição de Enxerto/induzido quimicamente , Sobrevivência de Enxerto , Humanos , Falência Renal Crônica/complicações , Masculino , Pessoa de Meia-Idade , Inibidores da Agregação Plaquetária/administração & dosagem , Inibidores da Agregação Plaquetária/efeitos adversos , Período Pós-Operatório , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Taxa de Sobrevida/tendências , Ticlopidina/administração & dosagem , Ticlopidina/efeitos adversos , Transplante Homólogo , Estados Unidos/epidemiologia
3.
Curr Protoc Mol Biol ; 107: 19.9.1-19.9.36, 2014 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-24984850

RESUMO

Electronic data resources can enable molecular biologists to quickly get information from around the world that a decade ago would have been buried in papers scattered throughout the library. The ability to access, query, and display these data makes benchwork much more efficient and drives new discoveries. Increasingly, mastery of software resources and corresponding data repositories is required to fully explore the volume of data generated in biomedical and agricultural research, because only small amounts of data are actually found in traditional publications. The UCSC Genome Browser provides a wealth of data and tools that advance understanding of genomic context for many species, enable detailed analysis of data, and provide the ability to interrogate regions of interest across disparate data sets from a wide variety of sources. Researchers can also supplement the standard display with their own data to query and share this with others. Effective use of these resources has become crucial to biological research today, and this unit describes some practical applications of the UCSC Genome Browser.


Assuntos
Biologia Computacional/métodos , Bases de Dados de Ácidos Nucleicos , Genoma , Navegador
4.
PLoS One ; 7(9): e44694, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23049756

RESUMO

Poxviruses express highly active inhibitors, including serine proteinase inhibitors (serpins), designed to target host immune defense pathways. Recent work has demonstrated clinical efficacy for a secreted, myxomaviral serpin, Serp-1, which targets the thrombotic and thrombolytic proteases, suggesting that other viral serpins may have therapeutic application. Serp-2 and CrmA are intracellular cross-class poxviral serpins, with entirely distinct functions from the Serp-1 protein. Serp-2 and CrmA block the serine protease granzyme B (GzmB) and cysteine proteases, caspases 1 and 8, in apoptotic pathways, but have not been examined for extracellular anti-inflammatory activity. We examined the ability of these cross-class serpins to inhibit plaque growth after arterial damage or transplant and to reduce leukocyte apoptosis. We observed that purified Serp-2, but not CrmA, given as a systemic infusion after angioplasty, transplant, or cuff-compression injury markedly reduced plaque growth in mouse and rat models in vivo. Plaque growth was inhibited both locally at sites of surgical trauma, angioplasty or transplant, and systemically at non-injured sites in ApoE-deficient hyperlipidemic mice. With analysis in vitro of human cells in culture, Serp-2 selectively inhibited T cell caspase activity and blocked cytotoxic T cell (CTL) mediated killing of T lymphocytes (termed fratricide). Conversely, both Serp-2 and CrmA inhibited monocyte apoptosis. Serp-2 inhibitory activity was significantly compromised either in vitro with GzmB antibody or in vivo in ApoE/GzmB double knockout mice. Conclusions The viral cross-class serpin, Serp-2, that targets both apoptotic and inflammatory pathways, reduces vascular inflammation in a GzmB-dependent fashion in vivo, and inhibits human T cell apoptosis in vitro. These findings indicate that therapies targeting Granzyme B and/or T cell apoptosis may be used to inhibit T lymphocyte apoptosis and inflammation in response to arterial injury.


Assuntos
Aorta/efeitos dos fármacos , Estenose das Carótidas/tratamento farmacológico , Citotoxicidade Imunológica/efeitos dos fármacos , Inflamação/tratamento farmacológico , Serpinas/farmacologia , Linfócitos T/efeitos dos fármacos , Proteínas Virais/farmacologia , Angioplastia/efeitos adversos , Animais , Aorta/imunologia , Aorta/transplante , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Estenose das Carótidas/etiologia , Estenose das Carótidas/imunologia , Estenose das Carótidas/patologia , Caspase 1/metabolismo , Caspase 8/metabolismo , Linhagem Celular , Expressão Gênica/efeitos dos fármacos , Granzimas/antagonistas & inibidores , Granzimas/metabolismo , Humanos , Inflamação/etiologia , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos , Ratos , Ratos Sprague-Dawley , Serpinas/genética , Serpinas/isolamento & purificação , Linfócitos T/imunologia , Proteínas Virais/genética , Proteínas Virais/isolamento & purificação
5.
Artigo em Inglês | MEDLINE | ID: mdl-22684055

RESUMO

The role of ADAM-8 in cancer and inflammatory diseases such as allergy, arthritis and asthma makes it an attractive target for drug development. Therefore, the catalytic domain of human ADAM-8 was expressed, purified and crystallized in complex with a hydroxamic acid inhibitor, batimastat. The crystal structure of the enzyme-inhibitor complex was refined to 2.1 Å resolution. ADAM-8 has an overall fold similar to those of other ADAM members, including a central five-stranded ß-sheet and a catalytic Zn(2+) ion. However, unique differences within the S1' binding loop of ADAM-8 are observed which might be exploited to confer specificity and selectivity to ADAM-8 competitive inhibitors for the treatment of diseases involving this enzyme.


Assuntos
Proteínas ADAM/química , Domínio Catalítico , Proteínas de Membrana/química , Fenilalanina/análogos & derivados , Inibidores de Proteases/química , Tiofenos/química , Proteínas ADAM/metabolismo , Humanos , Ligantes , Proteínas de Membrana/metabolismo , Modelos Moleculares , Fenilalanina/química , Fenilalanina/metabolismo , Inibidores de Proteases/metabolismo , Ligação Proteica , Desdobramento de Proteína , Tiofenos/metabolismo
6.
Hepat Med ; 3: 89-98, 2011 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-21927552

RESUMO

BACKGROUND AND AIMS: Activation of the oval cell compartment occurs in the liver when hepatocytes are functionally compromised and/or unable to divide. Our goal was to investigate the systemic signals responsible for determining the efficiency of oval cell-mediated liver regeneration, focusing on the Notch signaling cascade. METHODS: The established oval cell induction protocol of 2-acetylaminofluorine (2-AAF) implantation followed by 70% surgical resection of the liver (partial hepatectomy, PH) was employed in a rat model. This oval cell induction model was further combined with injections of a γ-secretase inhibitor (GSI XX) to examine the effects of Notch inhibition on oval cell-aided regeneration of the liver. RESULTS: Notch signaling was found to be upregulated at the peak of oval cell induction during 2AAF-PH alone. Treatment with GSI XX led to interruption of the Notch signal, as shown by a decrease in expression of Hes1. While there was a robust oval cell response seen at day 11 post-PH, there was a measurable delay in differentiation when Notch was inhibited. This was confirmed morphologically as well as by immunohistochemistry for the oval cell markers, α-fetoprotein, OV-6, and CK19. The hepatocytes seen at day 22 demonstrated an enhanced hepatocellular mitoinhibition index (p21(Waf1)/Ki67), suggestive of dysregulated proliferation and cell cycle progression. Moreover, these hepatocytes exhibited decreased expression of hepatocyte functional markers, such as cytochrome P450 and glucose-6-phosphatase-α. CONCLUSIONS: Taken together, these results identify the Notch signaling pathway as a potent regulator of differentiation and proliferation in oval cells, which is necessary for functional for repair of the liver by oval cells.

7.
Bioorg Med Chem Lett ; 21(10): 2820-2, 2011 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-21507637

RESUMO

Seeking compounds preferentially potent and selective for MMP-13, we reported in the preceding Letter on a series of hydroxamic acids with a flexible benzamide tail groups.(1a) Here, we replace the amide moiety with non-hydrolyzable heterocycles in an effort to improve half-life. We identify a hydroxamate tetrazole 4e that spares MMP-1 and -14, shows >400-fold selectivity versus MMP-8 and >600-fold selectivity versus MMP-2, and has a 4.8 h half-life in rats. X-ray data (1.9 Å) for tetrazole 4c is presented.


Assuntos
Amidas/síntese química , Inibidores Enzimáticos/síntese química , Compostos Heterocíclicos/síntese química , Ácidos Hidroxâmicos/síntese química , Inibidores de Metaloproteinases de Matriz , Sulfonas/síntese química , Amidas/química , Animais , Cristalografia por Raios X , Inibidores Enzimáticos/química , Compostos Heterocíclicos/química , Ácidos Hidroxâmicos/química , Metaloproteinase 13 da Matriz/química , Modelos Moleculares , Ratos , Relação Estrutura-Atividade , Especificidade por Substrato , Sulfonas/química
8.
Protein Sci ; 20(4): 735-44, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21370305

RESUMO

A ((1S,2R)-2-hydroxy-2,3-dihydro-1H-inden-1-yl) succinamide derivative (here referred to as Compound 12) shows significant activity toward many matrix metalloproteinases (MMPs), including MMP-2, MMP-8, MMP-9, and MMP-13. Modeling studies had predicted that this compound would not bind to ADAMTS-5 (a disintegrin and metalloproteinase with thrombospondin motifs-5) due to its shallow S1' pocket. However, inhibition analysis revealed it to be a nanomolar inhibitor of both ADAMTS-4 and -5. The observed inconsistency was explained by analysis of crystallographic structures, which showed that Compound 12 in complex with the catalytic domain of ADAMTS-5 (cataTS5) exhibits an unusual conformation in the S1' pocket of the protein. This first demonstration that cataTS5 can undergo an induced conformational change in its active site pocket by a molecule like Compound 12 should enable the design of new aggrecanase inhibitors with better potency and selectivity profiles.


Assuntos
Proteínas ADAM/química , Amidas/química , Conformação Proteica , Proteína ADAMTS5 , Animais , Domínio Catalítico , Bovinos , Desenho de Fármacos , Humanos , Inibidores de Metaloproteinases de Matriz , Metaloproteinases da Matriz/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Succinatos
9.
J Med Chem ; 54(5): 1211-22, 2011 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-21302953

RESUMO

Structure-based virtual screening was applied to design combinatorial libraries to discover novel and potent soluble epoxide hydrolase (sEH) inhibitors. X-ray crystal structures revealed unique interactions for a benzoxazole template in addition to the conserved hydrogen bonds with the catalytic machinery of sEH. By exploitation of the favorable binding elements, two iterations of library design based on amide coupling were employed, guided principally by the docking results of the enumerated virtual products. Biological screening of the libraries demonstrated as high as 90% hit rate, of which over two dozen compounds were single digit nanomolar sEH inhibitors by IC(50) determination. In total the library design and synthesis produced more than 300 submicromolar sEH inhibitors. In cellular systems consistent activities were demonstrated with biochemical measurements. The SAR understanding of the benzoxazole template provides valuable insights into discovery of novel sEH inhibitors as therapeutic agents.


Assuntos
Benzoxazóis/química , Epóxido Hidrolases/antagonistas & inibidores , Epóxido Hidrolases/química , Modelos Moleculares , Relação Quantitativa Estrutura-Atividade , Bibliotecas de Moléculas Pequenas , Benzoxazóis/síntese química , Técnicas de Química Combinatória , Cristalografia por Raios X , Desenho de Fármacos , Ensaios Enzimáticos , Fluorometria , Ligação de Hidrogênio , Estrutura Molecular , Solubilidade
10.
Brief Bioinform ; 11(6): 598-609, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20798181

RESUMO

The amount of biological data is increasing rapidly, and will continue to increase as new rapid technologies are developed. Professionals in every area of bioscience will have data management needs that require publicly available bioinformatics resources. Not all scientists desire a formal bioinformatics education but would benefit from more informal educational sources of learning. Effective bioinformatics education formats will address a broad range of scientific needs, will be aimed at a variety of user skill levels, and will be delivered in a number of different formats to address different learning styles. Informal sources of bioinformatics education that are effective are available, and will be explored in this review.


Assuntos
Biologia Computacional/educação , Software , Bases de Dados Factuais , Ensino
11.
J Med Chem ; 53(18): 6653-80, 2010 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-20726512

RESUMO

α-Sulfone-α-piperidine and α-tetrahydropyranyl hydroxamates were explored that are potent inhibitors of MMP's-2, -9, and -13 that spare MMP-1, with oral efficacy in inhibiting tumor growth in mice and left-ventricular hypertrophy in rats and in the bovine cartilage degradation ex vivo explant system. α-Piperidine 19v (SC-78080/SD-2590) was selected for development toward the initial indication of cancer, while α-piperidine and α-tetrahydropyranyl hydroxamates 19w (SC-77964) and 9i (SC-77774), respectively, were identified as backup compounds.


Assuntos
Anti-Inflamatórios/síntese química , Antineoplásicos/síntese química , Fármacos Cardiovasculares/síntese química , Ácidos Hidroxâmicos/síntese química , Inibidores de Metaloproteinases de Matriz , Piperidinas/síntese química , Piranos/síntese química , Sulfonas/síntese química , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Antineoplásicos/química , Antineoplásicos/farmacologia , Disponibilidade Biológica , Fármacos Cardiovasculares/química , Fármacos Cardiovasculares/farmacologia , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/patologia , Bovinos , Cristalografia por Raios X , Humanos , Ácidos Hidroxâmicos/química , Ácidos Hidroxâmicos/farmacologia , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Macaca fascicularis , Camundongos , Camundongos Nus , Modelos Moleculares , Estrutura Molecular , Piperidinas/química , Piperidinas/farmacologia , Piranos/química , Piranos/farmacologia , Ratos , Relação Estrutura-Atividade , Sulfonas/química , Sulfonas/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
12.
J Mol Biol ; 400(3): 413-33, 2010 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-20478313

RESUMO

Janus kinases (JAKs) are critical regulators of cytokine pathways and attractive targets of therapeutic value in both inflammatory and myeloproliferative diseases. Although the crystal structures of active JAK1 and JAK2 kinase domains have been reported recently with the clinical compound CP-690550, the structures of both TYK2 and JAK3 with CP-690550 have remained outstanding. Here, we report the crystal structures of TYK2, a first in class structure, and JAK3 in complex with PAN-JAK inhibitors CP-690550 ((3R,4R)-3-[4-methyl-3-[N-methyl-N-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino]piperidin-1-yl]-3-oxopropionitrile) and CMP-6 (tetracyclic pyridone 2-t-butyl-9-fluoro-3,6-dihydro-7H-benz[h]-imidaz[4,5-f]isoquinoline-7-one), both of which bind in the ATP-binding cavities of both JAK isozymes in orientations similar to that observed in crystal structures of JAK1 and JAK2. Additionally, a complete thermodynamic characterization of JAK/CP-690550 complex formation was completed by isothermal titration calorimetry, indicating the critical role of the nitrile group from the CP-690550 compound. Finally, computational analysis using WaterMap further highlights the critical positioning of the CP-690550 nitrile group in the displacement of an unfavorable water molecule beneath the glycine-rich loop. Taken together, the data emphasize the outstanding properties of the kinome-selective JAK inhibitor CP-690550, as well as the challenges in obtaining JAK isozyme-selective inhibitors due to the overall structural and sequence similarities between the TYK2, JAK1, JAK2 and JAK3 isozymes. Nevertheless, subtle amino acid variations of residues lining the ligand-binding cavity of the JAK enzymes, as well as the global positioning of the glycine-rich loop, might provide the initial clues to obtaining JAK-isozyme selective inhibitors.


Assuntos
Benzimidazóis/metabolismo , Inibidores Enzimáticos/metabolismo , Janus Quinase 3/química , Piridonas/metabolismo , Pirimidinas/metabolismo , Pirróis/metabolismo , TYK2 Quinase/química , Sítios de Ligação , Calorimetria , Humanos , Janus Quinase 3/metabolismo , Cinética , Modelos Moleculares , Piperidinas , Ligação Proteica , Estrutura Terciária de Proteína , TYK2 Quinase/metabolismo
13.
Am J Pathol ; 176(6): 2732-42, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20413689

RESUMO

To date the molecular signals regulating activation, proliferation, and differentiation of hepatic oval cells are not fully understood. The Wnt family is essential in hepatic embryogenesis and implicated in hepatic carcinogenesis. This study elucidates novel findings implicating Wnt1 in directing oval cell differentiation during the rat 2-acetylaminofluorene (2AAF) and 2/3 partial hepatectomy (PHx) liver regeneration model. Proteins of Wnt family members were predominantly localized in pericentral hepatocytes during liver injury, oval cell activation, and hepatocyte regeneration. In addition, Wnt message increased coinciding with the rise in oval cell number, whereas protein levels peaked immediately after the height of oval cell proliferation. Immunohistochemical analysis demonstrated nuclear translocation of beta-catenin within oval cells throughout the 2AAF/PHx protocol. Furthermore, RNA interference was used in vivo to confirm the physiological requirement of Wnt1 during the oval cell induction. Ultimately, inhibition of Wnt1 resulted in failure of oval cells to differentiate into hepatocytes and alternatively induced atypical ductular hyperplasia. Taken together, these data indicate that in vivo exposure to Wnt1 shRNA inhibited rat oval cell liver regeneration. In the absence of Wnt1 signaling, oval cells failed to differentiate into hepatocytes and underwent atypical ductular hyperplasia, exhibiting epithelial metaplasia and mucin production. Furthermore, changes in Wnt1 levels are required for the efficient regeneration of the liver by oval cells during massive hepatic injury.


Assuntos
Diferenciação Celular/fisiologia , Regeneração Hepática/fisiologia , Fígado/citologia , Células-Tronco/fisiologia , Proteína Wnt1/metabolismo , 2-Acetilaminofluoreno/farmacologia , Animais , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/metabolismo , Carcinógenos/farmacologia , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Hepatectomia , Fígado/efeitos dos fármacos , Fígado/patologia , Fígado/cirurgia , Masculino , Tamanho do Órgão , Interferência de RNA , Ratos , Ratos Endogâmicos F344 , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neurotransmissores/genética , Receptores de Neurotransmissores/metabolismo , Transdução de Sinais/fisiologia , Células-Tronco/citologia , Proteína Wnt1/genética , alfa-Fetoproteínas/genética , alfa-Fetoproteínas/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
14.
Protein Expr Purif ; 69(1): 54-63, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19781647

RESUMO

Janus-associated kinases (JAKs) play critical roles in cytokine signaling, and have emerged as viable therapeutic targets in inflammation and oncology related diseases. To date, targeting JAK proteins with highly selective inhibitor compounds have remained elusive. We have expressed the active kinase domains for both JAK2 and JAK3 and devised purification protocols to resolve the non-, mono- (Y1007) and diphosphorylated (Y1007 and Y1008) states of JAK2 and non- and monophosphorylated states of JAK3 (Y980). An optimal purified protein yield of 20, 29 and 69mg per 20L cell culture was obtained for the three JAK2 forms, respectively, and 12.2 and 2.3mg per 10L fermentation for the two JAK3 forms allowing detailed biochemical and biophysical studies. To monitor the purification process we developed a novel HPLC activity assay where a sequential order of phosphorylation was observed whereby the first tyrosine residue was completely phosphorylated prior to phosphorylation of the tandem tyrosine residue. A Caliper-based microfluidics assay was used to determine the kinetic parameters (K(m) and k(cat)) for each phosphorylated state, showing that monophosphorylated (Y1007) JAK2 enzyme activity increased 9-fold over that of the nonphosphorylated species, and increased an additional 6-fold for the diphosphorylated (Y1007/Y1008) species, while phosphorylation of JAK3 resulted in a negligible increase in activity. Moreover, crystal structures have been generated for each isolated state of JAK2 and JAK3 with resolutions better than 2.4A. The generation of these reagents has enabled kinetic and structural characterization to inform the design of potent and selective inhibitors of the JAK family.


Assuntos
Janus Quinase 2/química , Janus Quinase 2/isolamento & purificação , Janus Quinase 3/química , Janus Quinase 3/isolamento & purificação , Sequência de Aminoácidos , Biocatálise , Cromatografia Líquida de Alta Pressão , Cristalização , Eletroforese em Gel de Poliacrilamida , Fermentação , Humanos , Cinética , Dados de Sequência Molecular , Fosforilação , Estrutura Terciária de Proteína
15.
Hepat Med ; 2010(2): 13-32, 2010 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-21852899

RESUMO

Oval cell-mediated liver regeneration is a highly complex process that involves the coordination of several signaling factors, chemokines and cytokines to allow for proper maintenance of the liver architecture. When hepatocyte proliferation is inhibited, an hepatic stem cell population, often referred to as "oval cells", is activated to aid in liver regeneration. The function of insulin-like growth factor binding protein-3 (IGFBP-3) during this process of oval cell activation is of particular interest because it is produced in liver and has been shown to induce migration and differentiation of other stem cell populations both in vitro and in vivo. Additionally, IGFBP-3 production has been linked to the transforming growth factor-ß (TGF-ß) superfamily, a pathway known to be induced during oval cell proliferation. In this study, we set out to determine whether IGFBP-3 plays a role in oval cell proliferation, migration and differentiation during this specific type of regeneration. Through activation of the oval cell-mediated liver regeneration in a rat model, we found that IGFBP-3 is elevated in the liver and serum of animals during peak days of oval cell activation and proliferation. Furthermore, in vitro assays found that WB-344 cells, a liver stem cell line similar to oval cells, were induced to migrate in the presence of IGFBP-3. When expression of IGFBP-3 was knocked down during oval cell activation in vivo, we found that oval cell proliferation was increased and observed the appearance of numerous atypical ductular structures, which were OV-6 and Ki67-positive. Finally, quantitative real-time PCR analysis of liver tissue from IGFBP-3 small interfering RNA (siRNA) treated animals determined that expression of TGFß family members, including TGF-ßRII and Smads 2-4, were significantly downregulated compared to animals at day 9 post-PHx alone or animals that received negative control siRNA. In conclusion, IGFBP-3 may function as a potent chemoattractant of oval cells during specific types of liver regeneration and may be involved in regulating oval cell proliferation and differentiation in vivo via the TGF-ß pathway.

16.
Curr Protoc Mol Biol ; Chapter 19: Unit19.9, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19816931

RESUMO

Electronic data resources can enable molecular biologists to query and display many useful features that make benchwork more efficient and drive new discoveries. The UCSC Genome Browser provides a wealth of data and tools that advance one's understanding of genomic context for many species, enable detailed understanding of data, and provide the ability to interrogate regions of interest. Researchers can also supplement the standard display with their own data to query and share with others. Effective use of these resources has become crucial to biological research today, and this unit describes some practical applications of the UCSC Genome Browser.


Assuntos
Biologia Computacional/métodos , Genômica/métodos , Software , Gráficos por Computador , Bases de Dados Genéticas , Interface Usuário-Computador
17.
J Biol Chem ; 284(36): 24185-91, 2009 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-19586907

RESUMO

Several inhibitors of a series of cis-1(S)2(R)-amino-2-indanol-based compounds were reported to be selective for the aggrecanases, ADAMTS-4 and -5 over other metalloproteases. To understand the nature of this selectivity for aggrecanases, the inhibitors, along with the broad spectrum metalloprotease inhibitor marimastat, were independently bound to the catalytic domain of ADAMTS-5, and the corresponding crystal structures were determined. By comparing the structures, it was determined that the specificity of the relative inhibitors for ADAMTS-5 was not driven by a specific interaction, such as zinc chelation, hydrogen bonding, or charge interactions, but rather by subtle and indirect factors, such as water bridging, ring rigidity, pocket size, and shape, as well as protein conformation flexibility.


Assuntos
Endopeptidases/química , Inibidores Enzimáticos/química , Proteínas ADAM/química , Proteína ADAMTS4 , Proteína ADAMTS5 , Animais , Bovinos , Humanos , Ligação de Hidrogênio , Pró-Colágeno N-Endopeptidase/química , Estrutura Terciária de Proteína , Homologia Estrutural de Proteína , Zinco/química
18.
Biochemistry ; 48(27): 6402-11, 2009 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-19496616

RESUMO

PH-797804 is a diarylpyridinone inhibitor of p38alpha mitogen-activated protein (MAP) kinase derived from a racemic mixture as the more potent atropisomer (aS), first proposed by molecular modeling and subsequently confirmed by experiments. On the basis of structural comparison with a different biaryl pyrazole template and supported by dozens of high-resolution crystal structures of p38alpha inhibitor complexes, PH-797804 is predicted to possess a high level of specificity across the broad human kinase genome. We used a structural bioinformatics approach to identify two selectivity elements encoded by the TXXXG sequence motif on the p38alpha kinase hinge: (i) Thr106 that serves as the gatekeeper to the buried hydrophobic pocket occupied by 2,4-difluorophenyl of PH-797804 and (ii) the bidentate hydrogen bonds formed by the pyridinone moiety with the kinase hinge requiring an induced 180 degrees rotation of the Met109-Gly110 peptide bond. The peptide flip occurs in p38alpha kinase due to the critical glycine residue marked by its conformational flexibility. Kinome-wide sequence mining revealed rare presentation of the selectivity motif. Corroboratively, PH-797804 exhibited exceptionally high specificity against MAP kinases and the related kinases. No cross-reactivity was observed in large panels of kinase screens (selectivity ratio of >500-fold). In cellular assays, PH-797804 demonstrated superior potency and selectivity consistent with the biochemical measurements. PH-797804 has met safety criteria in human phase I studies and is under clinical development for several inflammatory conditions. Understanding the rationale for selectivity at the molecular level helps elucidate the biological function and design of specific p38alpha kinase inhibitors.


Assuntos
Benzamidas/farmacologia , Biologia Computacional , Inibidores de Proteínas Quinases/farmacologia , Pironas/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Benzamidas/química , Cristalografia por Raios X , Humanos , Ligação de Hidrogênio , Modelos Moleculares , Estrutura Molecular , Fosforilação , Inibidores de Proteínas Quinases/química , Piridonas , Pironas/química , Especificidade por Substrato , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
Bioorg Med Chem Lett ; 19(15): 4092-6, 2009 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19539468
20.
Biotechnol Annu Rev ; 14: 63-108, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18606360

RESUMO

The volume and complexity of genomic sequence data, and the additional experimental data required for annotation of the genomic context, pose a major challenge for display and access for biomedical researchers. Genome browsers organize this data and make it available in various ways to extract useful information to advance research projects. The UCSC Genome Browser is one of these resources. The official sequence data for a given species forms the framework to display many other types of data such as expression, variation, cross-species comparisons, and more. Visual representations of the data are available for exploration. Data can be queried with sequences. Complex database queries are also easily achieved with the Table Browser interface. Associated tools permit additional query types or access to additional data sources such as images of in situ localizations. Support for solving researcher's issues is provided with active discussion mailing lists and by providing updated training materials. The UCSC Genome Browser provides a source of deep support for a wide range of biomedical molecular research (http://genome.ucsc.edu).


Assuntos
Mapeamento Cromossômico/métodos , Gráficos por Computador , Análise Mutacional de DNA/métodos , Análise de Sequência de DNA/métodos , Software , Interface Usuário-Computador , Sequência de Bases , California , Biologia Molecular/métodos , Dados de Sequência Molecular , Projetos de Pesquisa , Universidades
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