RESUMEN
We report that a decrease in facilitative glucose transporter (GLUT1) expression and reduced glucose transport trigger apoptosis in the murine blastocyst. Inhibition of GLUT1 expression either by high glucose conditions or with antisense oligodeoxynucleotides significantly lowers protein expression and function of GLUT1 and as a result induces a high rate of apoptosis at the blastocyst stage. Similar to wild-type mice, embryos from streptozotocin-induced diabetic Bax -/- mice experienced a significant decrease in glucose transport compared with embryos from non-diabetic Bax -/- mice. However, despite this decrease, these blastocysts demonstrate significantly fewer apoptotic nuclei as compared with blastocysts from hyperglycemic wild-type mice. This decrease in preimplantation apoptosis correlates with a decrease in resorptions and malformations among the infants of the hyperglycemic Bax -/- mice versus the Bax +/+ and +/- mice. These findings suggest that hyperglycemia by decreasing glucose transport acts as a cell death signal to trigger a BAX-dependent apoptotic cascade in the murine blastocyst. This work also supports the hypothesis that increased apoptosis at a blastocyst stage because of maternal hyperglycemia may result in loss of key progenitor cells and manifest as a resorption or malformation, two adverse pregnancy outcomes more common in diabetic women.
Asunto(s)
Apoptosis/fisiología , Proteínas de Transporte de Monosacáridos/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2 , Proteínas Proto-Oncogénicas/fisiología , Animales , Apoptosis/efectos de los fármacos , Diabetes Mellitus Experimental/prevención & control , Femenino , Enfermedades Fetales/prevención & control , Regulación de la Expresión Génica/efectos de los fármacos , Transportador de Glucosa de Tipo 1 , Ratones , Sondas Moleculares , Proteínas de Transporte de Monosacáridos/genética , Oligonucleótidos Antisentido/farmacología , Proteínas Proto-Oncogénicas/genética , Proteína X Asociada a bcl-2RESUMEN
Mammalian preimplantation blastocysts exhibit insulin-stimulated glucose uptake despite the absence of the only known insulin-regulated transporter, GLUT4. We describe a previously unidentified member of the mammalian facilitative GLUT superfamily that exhibits approximately 20-25% identity with other murine facilitative GLUTs. Insulin induces a change in the intracellular localization of this protein, which translates into increased glucose uptake into the blastocyst, a process that is inhibited by antisense oligoprobes. Presence of this transporter may be necessary for successful blastocyst development, fuel metabolism, and subsequent implantation. Moreover, the existence of an alternative transporter may explain examples in other tissues of insulin-regulated glucose transport in the absence of GLUT4.
Asunto(s)
Blastocisto/metabolismo , Glucosa/metabolismo , Hipoglucemiantes/farmacología , Insulina/farmacología , Proteínas de Transporte de Monosacáridos/metabolismo , Secuencia de Aminoácidos , Animales , Transporte Biológico , Regulación de la Expresión Génica , Proteínas Facilitadoras del Transporte de la Glucosa , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Proteínas de Transporte de Monosacáridos/genética , Análisis de Secuencia de ProteínaRESUMEN
Several cyclopentene GABA analogues were synthesized as conformationally rigid analogues of the epilepsy drug vigabatrin and tested as inhibitors and substrates of gamma-aminobutyric acid aminotransferase (GABA-AT). None of these compounds produced time-dependent inhibition. (1R, 4S)-(+)-4-Amino-2-cyclopentene-1-carboxylic acid ((+)-3), (4R)-(-)-4-amino-1-cyclopentene-1-carboxylic acid ((-)-4), and d, l-3-amino-1-cyclopentene-1-carboxylic acid (6) are good substrates. The K(m) and k(cat) values for the latter two compounds are very similar to those of GABA, suggesting that they bind in an orientation similar to that of GABA. The K(m) value for (+)-3 is 24 times lower than that for GABA, although its k(cat) value is only one-fourth that for GABA; nonetheless, it is a better substrate for GABA-AT than is GABA. All of these compounds, as well as the enantiomers of 3 and 4 and d, l-trans-4-amino-2-cyclopentene-1-carboxylic acid (5), are competitive inhibitors of GABA-AT. These results demonstrate the effects of the carboxylate group orientation and the stereochemistry of the amino and carboxylate groups on the substrate activity and inhibitor activity, and this should be important to the future design of inhibitors of GABA-AT.