RESUMEN
Spermidine/spermine N(1)-acetyltransferase (SSAT) regulates polyamine catabolism. Thioredoxin-1 (Trx-1) is a redox protein that is overexpressed in human cancer leading to increased cell proliferation, decreased apoptosis, and decreased patient survival. We report that SSAT mRNA expression is decreased in Trx-1 transfected MCF-7 human breast cancer cells. There is also a decrease in SSAT enzyme activity and lower putrescine levels but no change in spermine or spermidine levels. The expression of SSAT is regulated by the NF-E2-related factor 2 (Nrf-2) and polyamine modulated factor-1 (PMF-1) transcription factor complex. Trx-1 transfected MCF-7 cells showed decreased Nrf-2/PMF-1 DNA binding without a change in Nrf-2 or PMF-1 protein expression. The results suggest that Trx-1 may play a role in the redox regulation of SSAT expression and polyamine homeostasis that could contribute to the biological effects of Trx-1.
Asunto(s)
Acetiltransferasas/metabolismo , Neoplasias de la Mama/metabolismo , Proteínas de la Membrana/metabolismo , Tiorredoxinas/metabolismo , Apoptosis , Northern Blotting , Western Blotting , División Celular , Núcleo Celular/metabolismo , ADN/metabolismo , Proteínas de Unión al ADN/metabolismo , Humanos , Factor 2 Relacionado con NF-E2 , Análisis de Secuencia por Matrices de Oligonucleótidos , Oxidación-Reducción , Poliaminas/química , Unión Proteica , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Transfección , Células Tumorales CultivadasRESUMEN
The efficacy of difluoromethylornithine (DFMO) as a chemopreventive agent has been tested in vitro using a human epidermal cell (HEC) assay with growth inhibition and involucrin induction as endpoints. Suppression of polyamine content is currently being utilized as a biomarker in clinical trials for the chemopreventive efficacy of DFMO against colon cancer formation. We have now examined the effects of DFMO on suppression of polyamine content in the HEC assay. The findings indicate 1) the % change in spermidine to spermine ratio and the depletion of putrescine show excellent correlation with chemopreventive efficacy in vitro; 2) the effective concentrations in vitro overlap the plasma concentrations in the clinical trial. These observations serve as further validation of the usefulness of the HEC assay as a screen for chemopreventive efficacy.
Asunto(s)
Antineoplásicos/farmacología , Biomarcadores de Tumor/metabolismo , Eflornitina/farmacología , Carcinógenos/farmacología , División Celular , Células Cultivadas , Quimioprevención , Relación Dosis-Respuesta a Droga , Humanos , Queratinocitos/citología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Putrescina/metabolismo , Espermidina/metabolismo , Espermina/metabolismo , Tiofenos/farmacologíaRESUMEN
Both the sulfide and sulfone metabolites of sulindac, a nonsteroidal anti-inflammatory drug, display anticarcinogenic effects in experimental models. Sulindac sulfide inhibits cyclooxygenase (COX) enzyme activities and has been reported to suppress ras-dependent signaling. However, the mechanisms by which sulindac sulfone suppresses cancer growth are not as defined. We studied the effects of these sulindac metabolites in human colon cancer-derived Caco-2 cells that have been transfected with an activated K-ras oncogene. Stable transfected clones expressed high levels of COX-2 mRNA and protein, compared with parental cells. K-ras-transfected cells formed tumors more quickly when injected into severe combined immunodeficiency disease mice than parental cells, and this tumorigenesis was suppressed by treatment with sulindac. Sulindac sulfone inhibited COX-2 protein expression, which resulted in a decrease in prostaglandin synthase E2 production. Sulindac sulfide had little effect on COX-2 in this model, but did suppress prostaglandin synthase E2 production, presumably by inhibiting COX enzyme activity. These data indicate that the sulfide and sulfone derivatives of sulindac exert COX-dependent effects by distinct mechanisms.
Asunto(s)
Anticarcinógenos/farmacología , Neoplasias del Colon/enzimología , Inhibidores de la Ciclooxigenasa/farmacología , Genes ras/efectos de los fármacos , Isoenzimas/antagonistas & inhibidores , Sulindac/farmacología , Animales , Células CACO-2 , Células Clonales , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Dinoprostona/biosíntesis , Genes ras/fisiología , Humanos , Isoenzimas/biosíntesis , Proteínas de la Membrana , Ratones , Ratones SCID , Prostaglandina-Endoperóxido Sintasas/biosíntesis , Sulindac/análogos & derivados , Transfección , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Overexpression of the BltD gene in Bacillus subtilis causes acetylation of the polyamines spermidine and spermine. BltD is co-regulated with another gene, Blt, which encodes a multidrug export protein whose overexpression facilitates spermidine export [Woolridge, Vazquez-Laslop, Markham, Chevalier, Gerner and Neyfakh (1997) J. Biol. Chem. 272, 8864-8866]. Here we show that BltD acetylates both spermidine and spermine at primary propyl amine moieties, with spermine being the preferred substrate. In the presence of saturating concentrations of acetyl CoA, BltD rapidly acetylates spermine at both the N1 and N12 positions. The Km (app) values for spermine, spermidine and N1-acetylspermine are =67, 200 and 1200 microM, respectively. Diamines ranging from 1, 3-diaminopropane to 1,12-diaminododecane, monoacetylputrescine and N8-acetylspermidine were not substrates for BltD. Putrescine (1, 4-diaminobutane) and N8-acetylspermidine were competitive inhibitors of spermidine acetylation by BltD, with Ki values of 0.25 and 5.76 mM, respectively. CoA competitively inhibited both spermidine and acetyl-CoA interactions with BltD. These data and other results indicate that the mechanism of spermidine and spermine acetylation by BltD is a random-order mechanism of bi-molecular kinetics.
Asunto(s)
Acetiltransferasas/metabolismo , Bacillus subtilis/enzimología , Acetilcoenzima A/metabolismo , Acetilación , Acetiltransferasas/antagonistas & inhibidores , Unión Competitiva , Coenzima A/metabolismo , Diaminas/metabolismo , Cinética , Espermidina/análogos & derivados , Espermidina/metabolismo , Espermina/análogos & derivados , Espermina/metabolismo , Especificidad por Sustrato , TermodinámicaRESUMEN
Chinese hamster ovary (CHO) cells were analyzed for their ability to reassemble microfilament bundles, to remain attached to a tissue culture surface, or to initiate and complete attachment onto a substrate after heat shock (45 degrees C/10 min). The cells remained attached to the tissue culture surface during and after the heat shock while the actin microfilament bundles were reversibly disrupted. Heat shock inhibited the ability of the cells to initiate and complete attachment onto a new tissue culture surface or onto a plastic surface coated with vitronectin. An inspection of the proteins present in substrate-attached material (SAM) revealed 11 major proteins containing glucosamine whose apparent Mr values were 250,000, 200,000, 150,000, 140,000, 90,000, 86,000, 82,000, 68,000, 54,000, 47,000, and 46,000. Three of the proteins (p200, p150, and p46) bound to wheat germ agglutinin while p150 and p140 bound to concanavalin A. The composition of the 11 proteins of the SAM fraction synthesized previous to the heat shock was not altered during heat shock. However, the appearance of the newly synthesized proteins in the SAM fraction was delayed by heat shock (0.5 h for p150 and 6 h for p82). The ability of heat-shocked cells to reattach onto a vitronectin-coated surface correlated with the appearance of newly synthesized p150 and p82 in the SAM fraction. Our results suggest that in addition to the microfilament bundles, heat shock may reversibly disrupt the cellular adhesion site. Further, p150 and p82, proteins whose appearance in the SAM fraction is delayed by heat shock, may be involved in the cellular attachment onto substrates, including vitronectin.
Asunto(s)
Citoesqueleto de Actina/metabolismo , Moléculas de Adhesión Celular/metabolismo , Adhesión Celular/fisiología , Citoesqueleto/metabolismo , Calor , Citoesqueleto de Actina/ultraestructura , Actinas/metabolismo , Animales , Técnicas In Vitro , Factores de TiempoRESUMEN
Surgical techniques for correcting midfacial deformities have used bone grafting with direct wire fixation, occlusal overcorrection, and maxillomandibular fixation for 6 to 8 weeks. With the advent of rigid internal fixation, the need for occlusal overcorrection and maxillomandibular fixation has been reduced or eliminated. A technique of rigid internal fixation using titanium mesh is presented. Two representative cases and their 18-month followups are described.
Asunto(s)
Maxilar/cirugía , Osteotomía/métodos , Mallas Quirúrgicas , Humanos , Inmovilización , TitanioRESUMEN
Hemifacial microsomia is a combination of malformations of the first and second branchial arch derivatives. Reconstruction of three adult cases of hemifacial microsomia with varying degrees of deformity was presented.
Asunto(s)
Asimetría Facial/cirugía , Adolescente , Adulto , Trasplante Óseo , Asimetría Facial/fisiopatología , Femenino , Humanos , Masculino , Maloclusión/cirugía , Mandíbula/cirugía , Maxilar/cirugía , Desarrollo Maxilofacial , Osteotomía/métodos , Trasplante AutólogoRESUMEN
Two cases of subcutaneous emphysema that occurred after Le Fort I osteotomies are reported, and diagnosis and treatment are discussed.