RESUMO
The purpose of this study is to assess the differences in VFA diagnostic accuracy when using bilateral decubitus views and whether diagnostic accuracy is affected by scoliosis. Our findings show that the current practice of performing only one side is valid; however, bilateral views can improve specificity in scoliosis. INTRODUCTION: The diagnostic accuracy of vertebral fracture assessment (VFA) can be influenced by poor patient position and scoliosis. This study aims to assess the differences in VFA diagnostic accuracy for right and left lateral decubitus views and the effect of scoliosis. METHODS: One hundred fourteen postmenopausal women received right and left lateral thoracolumbar spine dual-energy VFA and radiography. Cobb angles were measured from the posteroanterior absorptiometry image, and lumbar spine radiography was the standard reference for vertebral fracture and also provides the levels investigated. McNemar's test was used to compare accuracy between the two decubitus position and Fisher's exact test was used for patients with and without scoliosis. RESULTS: Forty-two vertebral fractures (VFs) were identified. There was no significant difference in sensitivity (p = 0.125) or specificity (p = 0.866) between the left lateral decubitus (64.3, 97.2%) and right lateral decubitus (76.2, 91.1%), respectively, views. Scoliotic patients had a significantly worse specificity (92.7 vs 98.1%, p = 0.003) than patients without scoliosis; however, a combination of both decubitus positions significantly improved specificity (p < 0.001). CONCLUSION: Right and left side lateral decubitus views have excellent agreement with radiography and similar diagnostic accuracy in the detection of VFs. Thus, the current practice of performing only one side is valid. With scoliosis, bilateral decubitus views can improve the specificity of detecting VF; however, this would increase radiation dose.
Assuntos
Fraturas por Compressão/diagnóstico por imagem , Fraturas por Osteoporose/diagnóstico por imagem , Fraturas da Coluna Vertebral/diagnóstico por imagem , Absorciometria de Fóton/métodos , Idoso , Idoso de 80 Anos ou mais , Feminino , Fraturas por Compressão/complicações , Humanos , Vértebras Lombares/diagnóstico por imagem , Vértebras Lombares/lesões , Pessoa de Meia-Idade , Osteoporose Pós-Menopausa/complicações , Osteoporose Pós-Menopausa/diagnóstico por imagem , Fraturas por Osteoporose/complicações , Radiografia/métodos , Escoliose/complicações , Escoliose/diagnóstico por imagem , Sensibilidade e Especificidade , Fraturas da Coluna Vertebral/complicaçõesRESUMO
OBJECTIVES: The purpose of this study was to identify delayed auditory maturation and the fate of premature infants who failed the newborn hearing screening (NHS) in neonatal intensive care unit. MATERIALS AND METHODS: A total of 1375 neonates underwent NHS using the transient evoked otoacoustic emission (TEOAE) in a tertiary hospital between 2007 and 2010 according to the Joint Committee on Infant Hearing guidelines. In addition, a structured telephone survey was given to caregivers of infants who were lost to follow-up NHS. Auditory steady-state response (ASSR) threshold and the threshold change in diagnostic test failures were analysed. RESULT: Among the 1375 NICU babies, 344 (25.0%) babies, 111 (9.7%) babies and 64 (4.6%) babies failed to pass the first TEOAE, second TEOAE and diagnostic ASSR, respectively. However, at the age of about 5 years, 12 (0.9%) infants showed permanent hearing loss (PHL). The ASSR threshold improved from 69.0 ± 19.7 dB to 52.9 ± 21.6 dB in <4 months (P < 0.001). Premature infants of <29 weeks of gestational age at birth showed higher referral (P = 0.003) rate at the first OAE test compared to the others, and the difference continued until the last follow-up. The odds ratio for the initial ASSR threshold >67.5 dB for PHL was 9.00 (95% confidence interval, 1.7-46.7). CONCLUSION: Most of first TEOAE screening failures (91.3%) showed normal hearing and speech development. Hearing levels in premature infants can improve over time, particularly in neonates with initial ASSR threshold <67.5 dB.
Assuntos
Limiar Auditivo/fisiologia , Potenciais Evocados Auditivos do Tronco Encefálico/fisiologia , Audição/fisiologia , Doenças do Prematuro/diagnóstico , Recém-Nascido Prematuro , Triagem Neonatal/métodos , Emissões Otoacústicas Espontâneas/fisiologia , Surdez/diagnóstico , Surdez/epidemiologia , Surdez/fisiopatologia , Feminino , Idade Gestacional , Testes Auditivos , Humanos , Incidência , Recém-Nascido , Doenças do Prematuro/epidemiologia , Doenças do Prematuro/fisiopatologia , Masculino , República da Coreia/epidemiologia , Estudos RetrospectivosRESUMO
The coordinated gold compound, 2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranosato-S-triethyl phosphine gold (auranofin; Ridaura), was evaluated for antitumor activity in a variety of mouse tumor models. Of the 15 tumor models evaluated, auranofin was found to be active only against i.p. P388 leukemia. A number of dose schedules was used to measure activity against P388 with optimal activity observed at 12 mg/kg given daily, i.p., on Days 1 to 5. Auranofin was active against i.p. P388 leukemia only when administered i.p.; the drug was completely inactive when administered i.v., s.c., or p.o. on Days 1 to 5. Evaluation of the effects of auranofin in vitro demonstrated that survival curves for B16 melanoma cells as measured by the clongenic and dye exclusion assays were exponential and monophasic; cell cycle distribution was not altered, and auranofin displayed no preferential cytotoxicity to logarithmic or plateau growth phase cell populations; auranofin inhibited DNA, RNA, and protein synthesis at cytotoxic concentrations but showed no selective effect; the cytotoxic activity and cellular association of gold from auranofin were dose, time, and temperature dependent; and binding of auranofin gold to serum proteins markedly decreased cellular uptake of gold and cytotoxicity of auranofin in vitro.
Assuntos
Antineoplásicos/uso terapêutico , Aurotioglucose/análogos & derivados , Ouro/análogos & derivados , Leucemia P388/tratamento farmacológico , Leucemia Experimental/tratamento farmacológico , Melanoma/tratamento farmacológico , Animais , Auranofina , Aurotioglucose/uso terapêutico , Aurotioglucose/toxicidade , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Cinética , Leucemia P388/patologia , Melanoma/patologia , Camundongos , Camundongos Endogâmicos DBA , Camundongos Endogâmicos , Ensaio Tumoral de Célula-TroncoRESUMO
S49 cyc- lymphoma cells contain a mutation resulting in loss of a functional guanine nucleotide regulatory protein rendering their adenylate cyclase refractory to most stimuli. S49 wild-type and cyc- clones were used in the present study to investigate the possible association of altered cAMP metabolism with tumorigenicity and metastatic potential. The S49 clones were implanted i.v., i.p., and intracerebrally in both athymic nude mice and syngeneic, immunocompetent BALB/c mice. Both S49 clones gave rise to tumors when inoculated into athymic mice, and no differences were observed in the tumorigenicity or metastatic potential of S49 wild-type and cyc- cells. Implantation of S49 clones in syngeneic BALB/c mice gave rise to few tumors except when administered intracerebrally, where wild-type cells were more tumorigenic than cyc- cells. This raises the possibility of differences in immunogenicity between the S49 clones. Analysis of cell lines derived from tumors grown in athymic mice showed that they retained the phenotype of the S49 clones used for inoculations. The results indicate that, despite differences in adenylate cyclase responsiveness, S49 wild-type and cyc- cells are both highly tumorigenic and metastatic.
Assuntos
Adenilil Ciclases/fisiologia , AMP Cíclico/fisiologia , Proteínas de Ligação ao GTP/fisiologia , Linfoma/enzimologia , Animais , Membrana Celular/enzimologia , Ciclofosfamida/farmacologia , Linfoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Metástase NeoplásicaRESUMO
The use of the human tumor cloning assay as a predictor of clinical response of human tumors to drugs is predicated on the hypothesis that the in vivo response of a tumor to a drug can be correlated with the in vitro response of cells derived from the tumor. To test this hypothesis, we utilized a murine tumor model in which the in vivo and in vitro responses of a tumor can be accurately and reproducibly compared. Drug activity was assessed in P388 leukemia with the standard in vivo antitumor assay (i.p. tumor/i.p. drug administration) and an in vitro assay wherein the ascites tumor cells are removed from mice, treated with a drug, and directly cloned in soft agar to measure clonogenic capacity. The response of P388 cells to analogues within four separate classes of antitumor agents, anthracyclines, anthraquinones, platinum(II) coordination complexes, and phosphinogold(I) complexes was evaluated. The clonogenic assay failed to discriminate between highly active in vivo antitumor agents and analogues with only marginal in vivo efficacy (i.e., doxorubicin and daunorubicin versus rhodomycins A and B, ametantrone versus NSC 276740, cisplatin versus transplatin, [Au(dppe)2]Cl versus [Au(depe)2]PF6. Furthermore, the in vitro clonogenic assay failed to detect carboplatin which was a highly active agent in vivo. The basis for these discrepancies was explored by a more detailed comparison of doxorubicin and rhodomycin B. In vivo or in vitro drug exposure with subsequent measurement of cell kill by the in vitro clonogenic and in vivo tumorigenic assay demonstrated that the in vitro assay overestimated the cytotoxic potency of the drugs relative to the tumorigenic assay. Treatment of tumors in vivo with doxorubicin at doses below the maximally tolerated dose in mice resulted in multiple log cell kill as measured in vitro or in vivo, whereas rhodomycin B was cytotoxic only at dose levels exceeding its maximally tolerated dose. The results indicate that a subset of tumor stem cells capable of forming colonies in soft agar are significantly more sensitive to the cytotoxic effects of anthracyclines than are in vivo tumorigenic stem cells. Cytotoxic potency as measured by an in vitro soft agar clonogenic assay is not an accurate predictor of in vivo antitumor efficacy even in a model in which ascites tumor cells are directly exposed to i.p. drug. The in vitro cytotoxicity assay is useful only as a nonselective prescreen and must be used in combination with other indicators of tumor cell selectivity and dose-limiting organ toxicity.
Assuntos
Ensaio de Unidades Formadoras de Colônias/métodos , Ensaio Tumoral de Célula-Tronco/métodos , Animais , Antraciclinas , Antraquinonas/farmacologia , Carboplatina , Cisplatino/farmacologia , Daunorrubicina/farmacologia , Doxorrubicina/farmacologia , Humanos , Leucemia P388/patologia , Camundongos , Naftacenos/farmacologia , Compostos Organoplatínicos/farmacologiaRESUMO
We have previously reported the cytotoxicity and antitumor activity of bis(diphenylphosphino)ethane (DPPE) and a variety of its transition metal complexes. During studies of the chemistry of a gold complex of this group [(AuCl)2(DPPE)], it was observed that this complex readily underwent ring closure on reaction with DPPE to form the tetrahedral complex [Au(DPPE)2]+. Various counterion forms (e.g., Cl-) of this cation were isolated and were found to exhibit a remarkably high stability in solution. Evaluation of [Au(DPPE)2]Cl in mice bearing i.p. P388 leukemia demonstrated that the compound produced an average of 87% increase in life span at its maximally tolerated dose (2-3 mumol/kg/day for 5 days). Activity was also seen in i.p. M5076 reticulum cell sarcoma (60% increase in life span) and s.c. mammary adenocarcinoma 16/c. Modest activity was evident in i.p. B16 melanoma and L1210 leukemia. A subline of P388 leukemia resistant to cisplatin was not cross-resistant to [Au(DPPE)2]Cl. In addition, combination therapy of [Au(DPPE)2]Cl and cisplatin against i.p. P388 demonstrated an advantage over single-agent therapy. In vitro studies of [Au(DPPE)2]Cl showed that the compound: is cytotoxic to tumor cell lines; is only minimally inhibited in its cytotoxic activity by the presence of serum; produces DNA protein cross-links and DNA strand breaks in cells; and inhibits macromolecular synthesis with a preferential inhibitory effect on protein synthesis relative to DNA and RNA synthesis. 31P nuclear magnetic resonance spectroscopy indicated that the compound is stable in the presence of serum proteins, thiols, or disulfides and that it reacts with Cu(II) resulting in the formation of a Cu(I)DPPE complex. The results of these in vivo and in vitro experiments suggest that the contrasting pharmacological profile of [Au(DPPE)2]Cl with respect to other gold(I) phosphine complexes may be related to both the kinetic stability of the complex and its stability in the presence of thiols.
Assuntos
Antineoplásicos , Ouro/uso terapêutico , Leucemia P388/tratamento farmacológico , Leucemia Experimental/tratamento farmacológico , Compostos Organometálicos , Compostos Organofosforados/uso terapêutico , Sarcoma Experimental/tratamento farmacológico , Animais , Sobrevivência Celular/efeitos dos fármacos , Fenômenos Químicos , Química , Cisplatino/administração & dosagem , Cobre , Sulfato de Cobre , DNA/efeitos dos fármacos , Leucemia L1210/tratamento farmacológico , Espectroscopia de Ressonância Magnética , Melanoma Experimental/tratamento farmacológico , Camundongos , Ácidos Nucleicos/biossíntese , Compostos Organoáuricos , Biossíntese de ProteínasRESUMO
Bis(diphenylphosphine)ethane (DPPE) and its bis[chlorogold(I)] [DPPE(Au2Cl2)], and bis[trichlorogold(III)] [DPPE(Au2Cl6)], complexes have in vivo antitumor activity. To determine if interaction with metals in situ can play a role in the antitumor activity of DPPE, we have studied the effects of DPPE, DPPE(Au2Cl2), DPPE(Au2Cl6) and mixtures of DPPE with metal salts on in vitro and in vivo biological systems. The in vitro cytotoxic potencies of the two DPPE-gold complexes were approximately 10-fold greater than that of DPPE. In addition, the cytotoxic potency of DPPE was increased when incubated with cells in the presence of Au(III) and Cu(II) salts, whereas Mg(II), Zn(II), Mn(II), Fe(II), Co(II), and Cd(II) had no effect. The effects of DPPE, DPPE(Au2Cl2) and mixtures of DPPE and metal salts on the activity of a model enzyme system, DNA polymerase alpha were measured. While DPPE did not inhibit the activity of DNA polymerase alpha, the DPPE(Au2Cl2) complex and mixtures of DPPE and Cu(II) salts inhibited the activity of the enzyme. Consistent with the effects observed in vitro, coadministration of Cu(II) or Au(III) increased the in vivo potency of DPPE in mice bearing i.p. P388 leukemia. Fifteen other DPPE analogues were evaluated for in vivo antitumor activity and for the effect of Cu(II) on their in vitro cytotoxic potency; there was a relationship between the ability of Cu(II) to potentiate the cytotoxic activities of DPPE analogues and their having in vivo antitumor activity.
Assuntos
Antineoplásicos/farmacologia , Metais/farmacologia , Compostos Organofosforados/farmacologia , Animais , Antineoplásicos/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Cobre/farmacologia , Dano ao DNA , DNA Polimerase II/antagonistas & inibidores , Ouro/farmacologia , Leucemia P388/tratamento farmacológico , Camundongos , Camundongos Endogâmicos , Compostos Organofosforados/metabolismoRESUMO
CoA-independent transacylase (CoA-IT) appears to play a critical role in lipid mediator generation by rapidly moving arachidonate (AA) between phospholipid pools during cell activation. Tumor necrosis factor-alpha (TNF) pretreatment of human neutrophils increases agonist-induced production of inflammatory mediators. The current study tested if the TNF-induced increase in lipid mediator production may be, in part, due to altered CoA-IT activity. Neutrophils were treated with TNF (250 U/ml, 30 min), homogenates prepared, and CoA-IT activity measured by the ability of these homogenates to acylate 1-[3H]alkyl-2-lyso-sn-glycero-3-phosphocholine (GPC). There was an increased CoA-IT activity, from 9.1 +/- 1.1 to 13.7 +/- 1.4 pmol/mg per min in control vs. TNF-treated samples, respectively. Varying the concentration of 1-alkyl-2-lyso-GPC revealed an increased CoA-IT activity in microsomes that was due to an increased Vmax, from 26 to 54 pmol/mg per min. The ability of TNF to increase CoA-IT activity was concentration-dependent, with maximal response observed at 25 U/ml. This effect on CoA-IT appears to be specific, in that TNF treatment of neutrophils had no effect on CoA-dependent acylation of 1-acyl-2-lyso-sn-glycero-3-phosphocholine, using either AA-CoA or linolenoyl-CoA as substrates. In the intact cell, the movement of [3H]AA from other phospholipids into PE in fMLP-stimulated neutrophils was greatly enhanced after TNF treatment, demonstrating a functional consequence of increased CoA-IT activity. In addition, TNF treatment doubled platelet-activating factor production in response to the chemotactic peptide fMLP, as measured by [3H]acetate incorporation, while the response to A23187 remained unchanged. Taken together, these results provide the first evidence of modulation of CoA-IT activity by a proinflammatory cytokine and suggest that one mechanism for augmented lipid mediator formation is through increases in CoA-IT activity.
Assuntos
Aciltransferases/metabolismo , Neutrófilos/enzimologia , Fator de Necrose Tumoral alfa/farmacologia , Ácido Araquidônico/metabolismo , Humanos , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/efeitos dos fármacos , Fosfolipídeos/metabolismo , Regulação para CimaRESUMO
Coenzyme A-independent transacylase (CoA-IT) mediates the transfer of polyunsaturated fatty acids from the sn-2 position of a donor phospholipid to the sn-2 position of an acceptor lyso-phospholipid. We have characterized this activity in U937 cells, a human monocytic cell line. The microsomes of these cells contained CoA-IT activity which demonstrated a fatty acid preference for transferring arachidonic acid into exogenously added 1-alkyl-2-lyso-GPC. This enzymatic activity was optimum between pH 6.5 and 9, was heat labile and displayed an apparent Km for 1-alkyl-2-lyso-GPC of 0.4 microM. This activity was not dependent on Ca2+, Mg2+, CoA or ATP, was not inhibited by 2-mercaptoethanol nor by addition of product, 1-alkyl-2-acyl-GPC. The activity of this enzyme was not altered by differentiation of U937 cells towards the macrophage with Me2SO. Treatment of U937 cells with dexamethasone had no effect on transacylase activity. The activity of this enzyme was decreased by the serine esterase inhibitors phenylmethyl-sulfonyl fluoride and N-tosyl-L-phenylalanine chloromethyl ketone and by the histidine modifier diethyl pyrocarbonate, suggesting that CoA-IT may belong to a family of acyltransferase enzymes typified by LCAT. CoA-IT activity was not affected by compounds that affect PLA2 activity, such as quinacrine, aristolochic acid and arachidonic acid, suggesting a mechanism of action for CoA-IT different from classical, low molecular weight PLA2 enzymes. In conclusion, U937 cells contain CoA-IT activity and this study extends our previous knowledge of this enzyme by demonstrating the differences between CoA-IT and PLA2 enzymes and suggesting similarities between CoA-IT and LCAT.
Assuntos
Aciltransferases/metabolismo , Ácidos Aristolóquicos , Microssomos/enzimologia , Aciltransferases/antagonistas & inibidores , Ácido Araquidônico , Ácidos Araquidônicos/metabolismo , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Dexametasona/farmacologia , Dietil Pirocarbonato/farmacologia , Ácido Edético/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Fenantrenos/farmacologia , Quinacrina/farmacologiaRESUMO
Platelet-activating factor (PAF) production is carefully controlled in inflammatory cells. The specific removal of arachidonate (AA) from 1-O-alkyl-2-arachidonoyl-sn-glycero-3-phosphocholine (GPC), thought to be mediated by CoA-independent transacylase (CoA-IT), is required to generate the PAF precursor 1-O-alkyl-2-lyso-GPC in human neutrophils. Exposure of A23187-stimulated human monocytes to the CoA-IT inhibitors SK&F 98625 and SK&F 45905 inhibited PAF formation (IC50s of 10 and 12 microM, respectively), indicating that these cells also need CoA-IT activity for PAF production. Because CoA-IT activity transfers arachidonate to a 2-lyso phospholipid substrate, its activity is obligated to an sn-2 acyl hydrolase to form the 2-lyso phospholipid substrate. SB 203347, an inhibitor of 14 kDa phospholipase A2 (PLA2), and AACOCF3, an inhibitor of 85 kDa PLA2, both inhibited AA release from A23187-stimulated human monocytes. However, AACOCF3 had no effect on A23187-induced PAF formation at concentrations as high as 3 microM. Further, depletion of 85 kDa PLA2 using antisense (SB 7111, 1 microM) had no effect on PAF production, indicating a lack of a role of 85 kDa PLA2 in PAF biosynthesis. Both SB 203347 and the 14 kDa PLA2 inhibitor scalaradial blocked PAF synthesis in monocytes (IC50s of 2 and 0.5 microM, respectively), suggesting a key role of 14 kDa PLA2 in this process. Further, A23187-stimulated monocytes produced two forms of PAF: 80% 1-O-alkyl-2-acetyl-GPC and 20% 1-acyl-2-acetyl-GPC, which were both equally inhibited by SB 203347. In contrast, inhibition of CoA-IT using SK&F 45905 (20 microM) had a greater effect on the production of 1-O-alkyl (-80%) than of 1-acyl (-14%) acetylated material. Finally, treatment of U937 cell membranes with exogenous human recombinant (rh) type II 14 kDa PLA2, but not rh 85 kDa PLA2, induced PAF production. Elimination of membrane CoA-IT activity by heat treatment impaired the ability of 14 kDa PLA2 to induce PAF formation. Taken together, these results suggest that a 14 kDa PLA2-like activity, and not 85 kDa PLA2, is coupled to monocyte CoA-IT-induced PAF production.
Assuntos
Aciltransferases/metabolismo , Monócitos/metabolismo , Fosfolipases A/metabolismo , Fator de Ativação de Plaquetas/biossíntese , Anti-Inflamatórios/farmacologia , Ácido Araquidônico/metabolismo , Ácidos Araquidônicos/farmacologia , Benzenossulfonatos/farmacologia , Calcimicina/farmacologia , Inibidores Enzimáticos/farmacologia , Homosteroides/farmacologia , Humanos , Monócitos/efeitos dos fármacos , Monócitos/enzimologia , Neutrófilos/efeitos dos fármacos , Fosfolipases A/antagonistas & inibidores , Fosfolipases A2 , Proteínas Recombinantes/metabolismo , Sesterterpenos , Sulfonamidas/farmacologia , Terpenos/farmacologia , Ureia/análogos & derivados , Ureia/farmacologiaRESUMO
A series of gold(I) coordination complexes including analogues of the antiarthritic agent auranofin 1 were evaluated for in vitro cytotoxic potency against both B16 melanoma cells and P388 leukemia cells and in vivo antitumor activity against P388 leukemia in mice. A number of the complexes showed potent cytotoxic activity in vitro and antitumor activity in vivo, with the phosphine-coordinated gold(I) thiosugar complexes demonstrating the greatest in vitro and in vivo activity. The data compiled for 63 complexes of the general structural formula LAuX provide the basis for the following observations: potent in vitro cytotoxic activity is observed for substituted (phosphine) gold complexes, lack of potency in vitro correlates well with lack of antitumor activity, potent cytotoxicity in vitro is not necessarily predictive of activity in vivo, in vivo antitumor activity is generally optimized by ligation of Au(I) with a substituted phosphine and a thiosugar.
Assuntos
Antineoplásicos/farmacologia , Ouro/farmacologia , Animais , Auranofina , Aurotioglucose/análogos & derivados , Aurotioglucose/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Leucemia P388/tratamento farmacológico , Melanoma/patologia , Camundongos , Solubilidade , Relação Estrutura-AtividadeRESUMO
5-Dialkylaminosulfonylisatins have been identified as potent, nonpeptide inhibitors of caspases 3 and 7. The most active compound within this series (34) inhibited caspases 3 and 7 in the 2-6 nM range and exhibited approximately 1000-fold selectivity for caspases 3 and 7 versus a panel of five other caspases (1, 2, 4, 6, and 8) and was at least 20-fold more selective versus caspase 9. Sequence alignments of the active site residues of the caspases strongly suggest that the basis of this selectivity is due to binding in the S2 subsite comprised of residues Tyr204, Trp206, and Phe256 which are unique to caspases 3 and 7. These compounds inhibit apoptosis in three cell-based models: human Jurkat T cells, human chondrocytes, and mouse bone marrow neutrophils.
Assuntos
Inibidores de Caspase , Inibidores de Cisteína Proteinase/síntese química , Isatina/análogos & derivados , Isatina/síntese química , Sulfonamidas/síntese química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Caspase 3 , Caspase 7 , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Condrócitos/citologia , Condrócitos/efeitos dos fármacos , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/farmacologia , Desenho de Fármacos , Humanos , Isatina/química , Isatina/farmacologia , Células Jurkat , Cinética , Camundongos , Modelos Moleculares , Conformação Molecular , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/fisiologia , Proteínas Recombinantes/antagonistas & inibidores , Relação Estrutura-Atividade , Sulfonamidas/química , Sulfonamidas/farmacologiaRESUMO
The ability of gold coordination complexes to bind to DNA and produce inter-strand cross-links in DNA was assessed in an assay system based on the fluorescence properties of the DNA intercalative dye, ethidium bromide. Results from these studies using a variety of gold(I) and gold(III) complexes suggest that the ability of gold complexes to bind to and produce inter-strand cross-links in DNA is not dependent on the oxidation state of gold in the complex but is influenced by the nature of the coordinating ligands. Those complexes in which the gold was ligated through one or more weakly coordinating ligands showed evidence for DNA binding. However, only those complexes with two or more of these relatively weak coordinating ligands produced inter-strand cross-links. Both the amount of binding to and cross-linking of DNA by these compounds were decreased by treatment of the gold-DNA complex with 2-mercaptoethanol and other thiol containing agents. As shown by agarose gel electrophoresis, 2-mercaptoethanol caused a dissociation of the gold-DNA complexes and a regeneration of closed circular superhelical pBR322 DNA. DNA strand breakage also resulted from treatment of a number of gold-DNA complexes with 2-mercaptoethanol; this was observed with the gold compounds which were shown to produce inter-strand cross-links in DNA. The amount of DNA strand breakage produced by treatment of gold-DNA complexes with 2-mercaptoethanol was influenced by the initial conformation of the DNA; gold-DNA complexes which resulted from the binding of gold compounds to covalently closed superhelical DNA were more sensitive to the breakage induced by 2-mercaptoethanol treatment than those complexes in which closed circular, relaxed DNA was used as substrate. The DNA breakage was not reduced in partially anaerobic conditions or by free-radical scavengers, suggesting that it is not mediated by oxygen. The results are discussed with respect to the potential for the interaction of gold complexes with intracellular DNA and chromatin and their biological implications.
Assuntos
DNA/metabolismo , Ouro/farmacologia , Anaerobiose , Animais , Bovinos , Cisplatino/farmacologia , Reagentes de Ligações Cruzadas/farmacologia , DNA Super-Helicoidal/metabolismo , Eletroforese em Gel de Ágar , Etídio , Fluorescência , Radicais Livres , Ouro/metabolismo , Mercaptoetanol/farmacologia , Conformação de Ácido Nucleico , Oxigênio/metabolismo , PlasmídeosRESUMO
Recent studies suggest that the first step in platelet-activating factor (PAF) biosynthesis, 1-alkyl-2-lyso-GPC (lyso PAF) formation, may be initiated by the selective transfer of arachidonate from 1-alkyl-2-arachidonoyl-GPC to an acceptor lyso phospholipid by a CoA-independent transacylase activity (CoA-IT). The present study was designed to determine whether the formation of 1-alkyl-2-lyso-GPC and the release of arachidonic acid can occur by different mechanisms. These experiments examined both the formation of 1-[3H]alkyl-2-lyso-GPC from 1-[3H]alkyl-2-acyl-GPC and the release of arachidonic acid from membrane phospholipids as determined by GC/MS in neutrophil homogenates under various conditions. The addition of unlabelled lyso phospholipids to neutrophil homogenates stimulated the time-dependent formation of 1-[3H]alkyl-2-lyso-GPC from 1-[3H]alkyl-2-acyl-GPC. Without exogenous lyso phospholipids, little 1-[3H]alkyl-2-lyso-GPC was formed in this reaction. The activity which catalyzed the formation of 1-[3H]alkyl-2-lyso-GPC had characteristics identical to CoA-IT as indicated by the fact that both reactions were: independent of Ca2+, Mg2+, CoA and CoA fatty acids, located in microsomal fractions, and stable in 10 mM dithiothreitol. In sharp contrast to the aforementioned reaction, addition of lyso phospholipids did not affect the quantity of arachidonic acid released from membrane phospholipids. Furthermore, there was a Ca(2+)-independent release of arachidonic acid from membrane phospholipid that was increased 4 to 5-fold after the addition of 5 mM Ca2+. Finally, Ca(2+)-dependent arachidonic acid release was inhibited by putative phospholipase A2 inhibitors, aristolochic acid and scalaradial, at concentrations where neither the production of 1-[3H]alkyl-2-lyso-GPC nor Ca(2+)-independent arachidonic acid release was altered. Together these data imply that there may be different mechanisms involved in the formation of 1-alkyl-2-lyso-GPC and arachidonic acid from membrane phospholipids.
Assuntos
Ácido Araquidônico/metabolismo , Cálcio/metabolismo , Neutrófilos/metabolismo , Fator de Ativação de Plaquetas/análogos & derivados , Aciltransferases/metabolismo , Membrana Celular/metabolismo , Humanos , Fosfatidilcolinas/metabolismo , Fosfolipases A/antagonistas & inibidores , Fosfolipases A/metabolismo , Fosfolipases A2 , Fosfolipídeos/metabolismo , Fator de Ativação de Plaquetas/biossínteseRESUMO
The interactions of certain gold(I) and gold(III) complexes with isolated plasmid pBR322 DNA were defined and compared to those of cis-diamminedichloroplatinum(II), CDDP, using an agarose gel electrophoresis assay. Trichloro(pyridine)gold(III) appeared to bind to DNA as evidenced by its ability to produce dose-dependent changes in the electrophoretic mobilities of closed circular, supercoiled, closed circular, relaxed, and open circular plasmid DNAs. These effects suggest that the gold containing complex induces conformational changes in the plasmid as a result of the compound binding to the DNA and the subsequent unwinding of the double helix and shorting of the DNA. Auranofin [(2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranosato-S)-triethyl phosphine gold(I)] did not appear to interact with DNA under any conditions. However, its analog chloro(triethylphosphine) gold(I) interacted with DNA at pH 9.5 in borate buffer and produced electrophoretic mobility changes in pBR322 DNA which were different from those produced by the gold(III) complexes that were evaluated. Binding of chloro(triethylphosphine) gold(I) was inhibited by the co-addition of the thiosugar portion of auranofin suggesting preferential binding of the gold moiety to thiosugar, which results in the production of auranofin (or a sugar containing) gold complex and inhibition of gold binding to DNA. The interactions of a number of gold compounds with DNA were also evidenced by their abilities to inhibit the binding of ethidium bromide to DNA. The results from these studies indicate that: gold containing complexes can bind to, and produce conformational changes in, DNA; gold(I) and gold(III) complexes may interact with DNA via different chemical mechanisms to produce different conformational changes in DNA; and certain coordinating ligands in gold complexes (e.g. Cl, Br and SCN) can be exchanged for binding sites on DNA by gold.
Assuntos
DNA/metabolismo , Ouro/farmacologia , Animais , Bovinos , Cisplatino/farmacologia , Eletroforese em Gel de Ágar , Etídio , Fluorescência , Ouro/metabolismo , Ligantes , Conformação de Ácido Nucleico , Plasmídeos , Relação Estrutura-AtividadeRESUMO
AIM: In this study, we investigated the prognostic significance of the number of examined lymph nodes in node-negative gastric adenocarcinoma (GC). PATIENTS AND METHODS: A total of 1194 node-positive and 1030 node-negative GC patients undergoing potentially curative gastrectomy was enrolled in this study. Patients were stratified into 3 groups according to the number of examined lymph nodes: group 1, ≤ 15; group 2, 16-25; group 3, >25. RESULTS: Patients with node-negative GC had significantly favorable survival compared with those with node-positive. Among patients with node-negative T2-T4 disease, the percentage of locoregional relapse was higher in those with <25 examined lymph nodes than in those with ≥ 25 examined lymph nodes. The number of examined lymph nodes affected the overall survival rates for patients with node-negative T2-T4 GC but not for patients with T1 lesions. Tumor size, tumor location, the number of examined lymph nodes, T status, and the presence of perineural invasion were significant prognostic factors as determined by multivariate analysis in node-negative GC. CONCLUSIONS: No survival benefit of examining ≥ 15 lymph nodes was noted for patients with node-negative T1 GC. Extensive lymphadenectomy in patients with node-negative T2-T4 lesions in whom the number of examined lymph nodes was >25 had favorable survival.
Assuntos
Adenocarcinoma/patologia , Excisão de Linfonodo , Linfonodos/patologia , Neoplasias Gástricas/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Linfonodos/cirurgia , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Prognóstico , Resultado do TratamentoRESUMO
BACKGROUND: The burden of gastroesophageal reflux disease (GERD) is increasing in the Asia area and the majority of GERD patients have non-erosive reflux disease (NERD). AIM: To evaluate the efficacy and safety of sodium alginate suspension compared to omeprazole in adult subjects with NERD. METHODS: In this 4-week, double-blind, parallel study, 195 NERD subjects were randomised to one of two treatment groups: sodium alginate suspension 20 mL three times a day and omeprazole 20 mg once daily. The primary efficacy endpoint was the percentage of patients achieving adequate heartburn or regurgitation relief at day 28 assessed by patient diary. The secondary efficacy endpoints included percentage of patients achieving adequate heartburn or regurgitation relief, change from baseline of the Reflux Disease Questionnaire total score at day 14 and 28 from baseline, and patients' overall satisfaction. RESULTS: In this study, 183 subjects were included in the intent-to-treat population, and 172 subjects were included in the per-protocol population. Non-inferiority of sodium alginate to omeprazole was demonstrated in the intent-to-treat population [difference, 2.7% (53.3% vs. 50.5%, P = 0.175), 95% lower confidence interval -11.9%, above the preset margin of -19%]. All of the secondary efficacy endpoints were comparable between two groups. The incidence of adverse event was relatively low and there was no difference between the two groups (5.4% vs. 5.5% for sodium alginate vs. omeprazole). No severe adverse event was noted in this study. CONCLUSION: The study showed that sodium alginate was as effective as omeprazole for symptomatic relief in patients with non-erosive reflux disease (Clinicaltrials.gov NCT01338077).
Assuntos
Alginatos/uso terapêutico , Refluxo Gastroesofágico/tratamento farmacológico , Omeprazol/uso terapêutico , Inibidores da Bomba de Prótons/uso terapêutico , Administração Oral , Adulto , Alginatos/administração & dosagem , Método Duplo-Cego , Feminino , Refluxo Gastroesofágico/patologia , Fármacos Gastrointestinais/administração & dosagem , Fármacos Gastrointestinais/uso terapêutico , Ácido Glucurônico/administração & dosagem , Ácido Glucurônico/uso terapêutico , Azia/tratamento farmacológico , Azia/etiologia , Ácidos Hexurônicos/administração & dosagem , Ácidos Hexurônicos/uso terapêutico , Humanos , Masculino , Pessoa de Meia-Idade , Omeprazol/administração & dosagem , Satisfação do Paciente , Inibidores da Bomba de Prótons/administração & dosagem , Inquéritos e Questionários , Suspensões , Resultado do TratamentoAssuntos
Aciltransferases/antagonistas & inibidores , Ácido Araquidônico/antagonistas & inibidores , Ácido Araquidônico/metabolismo , Inibidores Enzimáticos/farmacologia , Fator de Ativação de Plaquetas/biossíntese , Animais , Benzenossulfonatos/farmacologia , Medula Óssea/efeitos dos fármacos , Células da Medula Óssea , Humanos , Imidazóis/farmacologia , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos CBA , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Compostos Organofosforados/farmacologia , Ureia/análogos & derivados , Ureia/farmacologiaRESUMO
CoA-independent transacylase (CoA-IT) catalyzes the transfer of arachidonic acid from acyl- to alkyl-linked phospholipids. The removal of arachidonic acid from the sn-2 position of the donor phospholipid is a PLA2-like reaction. However, examination of CoA-IT in U937 cells demonstrated that CoA-IT has many characteristics that are distinct from those of PLA2 enzymes, including activity in the absence of Ca2+, activity that was heat and acid unstable and stable in 10 mM 2-mercaptoethanol and that was inhibited by detergents. Compounds that inhibit PLA2 activity did not inhibit CoA-IT activity, including quinacrine, aristolochic acid and arachidonic acid. All of these characteristics of CoA-IT are in contrast to those of most PLA2 enzymes. These data suggest that CoA-IT is biochemically different from, and has a mechanism of action unique from PLA2 enzymes.
Assuntos
Aciltransferases/metabolismo , Fosfolipases A/metabolismo , Aciltransferases/antagonistas & inibidores , Células Cultivadas , Detergentes/farmacologia , Mercaptoetanol/farmacologia , Microssomos/enzimologia , Fosfolipases A/antagonistas & inibidores , Fosfolipases A2RESUMO
The present studies were conducted to understand better the regulation of phospholipase A2 (PLA2)-dependent mobilization of lipid mediators by arachidonic acid (C20:4). After stimulation of human neutrophils, g.l.c./m.s. analysis of non-esterified fatty acids indicated that the quantity of C20:4 increased as a function of time after stimulation, from undetectable quantities to > 800 pmol/10(7) cells. In contrast with C20:4, the quantities of other free fatty acids such as oleic and linoleic were high in resting cells and did not change after stimulation. Some 15% of the C20:4 released from cellular lipids remained cell-associated. To examine the effect of C20:4 on its own release, neutrophils were exposed to [2H8]C20:4, to differentiate it by g.l.c./m.s. from naturally occurring C20:4. In A23187-stimulated neutrophils, low concentrations (5-10 microM) of [2H8]C20:4 added just before A23187 increased the quantity of C20:4 produced by the cell, whereas higher concentrations (30-50 microM) decreased the quantity of C20:4 released from phospholipids. As other measures of PLA2 activity, the effects of C20:4 on production of platelet-activity factor (PAF) and leukotriene B4 (LTB4) were assessed. C20:4 treatment just before stimulation of neutrophils blocked PAF and LTB4 production in a concentration-dependent manner (IC50 10-20 microM). The effect of C20:4 was not blocked by the cyclo-oxygenase inhibitor naproxine (10 microM), nor could it be mimicked by 1 microM LTB4, 5-hydroxyeicosa-6,8,11,14-tetraenoic acid (5HETE), 5-hydroperoxyeicosa-6,8,11,14-tetraenoic acid (5HPETE) or 15-hydroxyeicosa-5,8,11,13-tetraenoic acid (15HETE). The 5-lipoxygenase (5LO) inhibitor zileuton induced a concentration-dependent decrease in PAF, with a maximal effect of a 50% decrease at 10-50 microM. The decrease in PAF by the 5LO inhibitor could not be circumvented by addition of 1 microM 5HETE, 5HPETE and LTB4, and may be attributed to the capacity of zileuton to increase the quantity of C20:4 in A23187-treated neutrophils. The inhibitory effect of C20:4 (20-40 microM) on PAF production could be antagonized by the protein kinase C inhibitor staurosporine (30 nM), but not by inhibitors of protein kinase A, tyrosine kinase or calmodulin kinase II. Taken together, these data demonstrate that C20:4 is selectively released from membrane phospholipids of A23187-stimulated neutrophils, and this C20:4 may play an important role in regulating the mobilization of C20:4 by altering PLA2 activity.