RESUMEN
Fourth-year medical students at the University of Colorado School of Medicine distributed cards to patients in the emergency department asking, "What Worries You Most?" The patients' responses provided insight about their most pressing concerns, often unrelated to their "chief complaints."
Asunto(s)
Servicio de Urgencia en Hospital , Pacientes/psicología , Estrés Psicológico , Femenino , Humanos , Masculino , Atención Primaria de Salud , Encuestas y CuestionariosRESUMEN
A disintegrin and metalloproteinase 8 (ADAM8), a catalytically active member of the ADAMs family of enzymes, is expressed primarily on immune cells and thus probably involved in inflammatory responses. ADAM8 is also produced by chondrocytes, and recombinant ADAM8 can induce cartilage catabolism. We therefore decided to test the role of ADAM8 in autoimmune inflammatory arthritis using transgenic mice expressing catalytically inactive ADAM8. Transgenic DBA/1J mice expressing an inactivating point mutation in the ADAM8 gene to change Glu330 to Gln330 (ADAM8(EQ)) were generated to evaluate the proteolytic function of ADAM8 in an lipopolysaccharide-synchronized collagen-induced arthritis (LPS-CIA) model of autoimmune arthritis. The systemic inflammatory reaction to LPS was also evaluated in these mice. Expression profiling of paw joints from wild-type mice revealed that ADAM8 mRNA levels increased at the onset of clinical arthritis and correlated well with cellular macrophage markers. When subjected to LPS-CIA, ADAM8(EQ) mice demonstrated decreased incidence and severity of clinical arthritis compared to wild-type mice. Histological examination of paw joints from ADAM8(EQ) mice confirmed marked attenuation of synovial inflammation, cartilage degradation and bone resorption when compared to wild-type mice. However, transgenic mice and wild-type mice responded similarly to LPS-induced systemic inflammation with regard to mortality, organ weights, neutrophil sequestration and serum cytokine/chemokine production. We conclude that ADAM8 proteolytic activity plays a key role in the development of experimental arthritis and may thus be an attractive target for the treatment of arthritic disorders while minimizing risk of immunocompromise.
Asunto(s)
Proteínas ADAM/fisiología , Antígenos CD/fisiología , Artritis Experimental/metabolismo , Artritis Reumatoide/metabolismo , Proteínas de la Membrana/fisiología , Proteínas ADAM/genética , Proteínas ADAM/metabolismo , Animales , Antígenos CD/genética , Antígenos CD/metabolismo , Artritis Experimental/inmunología , Artritis Experimental/patología , Artritis Reumatoide/inmunología , Artritis Reumatoide/patología , Autoanticuerpos/biosíntesis , Catálisis , Células Cultivadas , Colágeno Tipo II/inmunología , Citocinas/sangre , Progresión de la Enfermedad , Perfilación de la Expresión Génica/métodos , Ácido Glutámico/genética , Lipopolisacáridos/inmunología , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos DBA , Ratones Transgénicos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Tamaño de los Órganos , Mutación Puntual , Índice de Severidad de la EnfermedadRESUMEN
Chiral N,N-disubstituted trifluoro-3-amino-2-propanols represent a recently discovered class of compounds that inhibit the neutral lipid transfer activity of cholesteryl ester transfer protein (CETP). These compounds all contain a single chiral center that is essential for inhibitory activity. (R,S)SC-744, which is composed of a mixture of the two enantiomers, inhibits CETP-mediated transfer of [(3)H]cholesteryl ester ([(3)H]CE) from HDL donor particles to LDL acceptor particles with an IC(50) = 200 nM when assayed using a reconstituted system in buffer and with an IC(50) = 6 microM when assayed in plasma. Upon isolation of the enantiomers, it was found that the (R,+) enantiomer, SC-795, was about 10-fold more potent than the mixture, and that the (S,-) enantiomer, SC-794, did not have significant inhibitory activity (IC(50) > 0.8 microM). All of the activity of the (S,-)SC-794 enantiomer could be accounted for by contamination of this sample with a residual 2% of the highly potent (R,+) enantiomer, SC-795. The IC(50) of (R,+)SC-795, 20 nM, approached the concentration of CETP (8 nM) in the buffer assay. These chiral N,N-disubstituted trifluoro-3-amino-2-propanols were found to associate with both LDL and HDL, but did not disrupt overall lipoprotein structure. They did not affect the on or off rates of CETP binding to HDL disk particles. Inhibition was highly specific since the activities of phospholipid transfer protein and lecithin cholesterol acyl transferase were not affected. Competition experiments showed that the more potent enantiomer (R)SC-795 prevented cholesteryl ester binding to CETP, and direct binding experiments demonstrated that this inhibitor bound to CETP with high affinity and specificity. It is estimated, based on the relative concentrations of inhibitor and lipid in the transfer assay, that (R)SC-795 binds approximately 5000-fold more efficiently to CETP than the natural ligand, cholesteryl ester. We conclude that these chiral N,N-disubstituted trifluoro-3-amino-2-propanol compounds do not affect lipoprotein structure or CETP-lipoprotein recognition, but inhibit lipid transfer by binding to CETP reversibly and stereospecifically at a site that competes with neutral lipid binding.
Asunto(s)
Proteínas Portadoras/antagonistas & inhibidores , Ésteres del Colesterol/antagonistas & inhibidores , Glicoproteínas , Proteínas de Transferencia de Fosfolípidos , Propanolaminas/farmacología , Triglicéridos/antagonistas & inhibidores , Animales , Unión Competitiva/efectos de los fármacos , Células CHO , Proteínas Portadoras/metabolismo , Proteínas de Transferencia de Ésteres de Colesterol , Ésteres del Colesterol/metabolismo , Cricetinae , Disulfuros/química , Disulfuros/farmacología , Sinergismo Farmacológico , Electroforesis en Gel de Agar , Humanos , Lipoproteínas HDL/antagonistas & inhibidores , Lipoproteínas HDL/metabolismo , Lipoproteínas LDL/antagonistas & inhibidores , Lipoproteínas LDL/metabolismo , Proteínas de la Membrana/antagonistas & inhibidores , Fosfatidilcolina-Esterol O-Aciltransferasa/metabolismo , Fosfolípidos/antagonistas & inhibidores , Propanolaminas/química , Estereoisomerismo , Relación Estructura-Actividad , Factores de TiempoAsunto(s)
Proteínas Portadoras/metabolismo , Ensayo de Inmunoadsorción Enzimática , Glicoproteínas , Animales , Anticuerpos Monoclonales/inmunología , Proteínas Portadoras/sangre , Proteínas Portadoras/inmunología , Colesterol/metabolismo , Proteínas de Transferencia de Ésteres de Colesterol , Ésteres del Colesterol/metabolismo , Humanos , Lipoproteínas HDL/metabolismo , Lipoproteínas LDL/metabolismo , RadioinmunoensayoRESUMEN
It has been suggested that at the time of innervation, developing neurites release one or more soluble factors that locally induce acetylcholine receptor (AChR) aggregate formation at the synaptic site. To test this hypothesis, we developed a model system that mimics the local, neural induction of AChR aggregation at developing synapses. Partially purified protein derived from fetal pig brain was applied locally to the surface of cultured myotubes via a micropipet. We found that local application of this factor for as little as 30 min induced the formation of AChR aggregates that were restricted to a region within 30 microns around the release site. In addition, the locally applied factor induced a local AChR aggregation response, but did not cause a detectable change in the myotube resting membrane potential at the release site. Our data support a soluble factor hypothesis and suggest that neither cell-cell contact nor local electric fields are necessary for the initial induction of AChR aggregation.