Regulation of glucose transporters in sheep placenta

Abstract

Transplacental glucose transport is vital to fetal growth. Although the presence of glucose transporter-1 (GLUT1) and GLUT3 has been demonstrated in mammalian placenta, the factors regulating these genes remain unclear. Therefore, the overall aim of these studies was to clone ovine GLUT1 (oGLUT1) and oGLUT3 cDNAs, and to use these to investigate gene expression during ovine placental development and function. Ovine GLUT1 (~2.2 kb) and oGLUT3 (483 bp) cDNAs were isolated and cloned. Sequence analysis demonstrated that oGLUT1 showed high homology (97 - 99%) with other mammalian species, whereas oGLUT3 did not (84 - 88%). Northern analysis demonstrated that oGLUT1 mRNA abundance increased from d 45 to d 120 of gestation, then decreased towards term (d 145 ± 2), whereas oGLUT3 mRNA abundance increased throughout gestation. Western analysis showed oGLUT1 protein levels increased during late gestation, indicating post-transcriptional regulation of oGLUT1. Localisation experiments revealed spatio-temporal differences in ovine placental GLUT expression. In early gestation (d 45), oGLUT1 protein was restricted to fetal trophoblast cells. By mid gestation oGLUT1 immuno-signal was predominantly localised to maternal villous and endometrial tissue. By late gestation oGLUT1 mRNA was most strongly localised to maternal syncytiotrophoblast and villous tissue, whereas oGLUT3 was predominantly localised to fetal trophoblast cells. Placental oGLUT expression was regulated differently by acute (3 - 8 h) versus long-term (>6 d) alterations in late gestation maternal glucose supply. No evidence was found for regulation of placental oGLUT gene expression by long-term maternal undernutrition, but oGLUT1 and oGLUT3 mRNA and oGLUT1 protein were elevated by short-term (24 - 48 h) maternal hypoglycemia. Acute maternal hyperglycemia transiently increased oGLUT1 and oGLUT3 mRNA abundance, whereas oGLUT1 protein (but not mRNA) levels increased after long-term maternal hyperglycemia. Infusion studies provided no conclusive evidence for regulation of placental oGLUTs by long-term administration of growth hormone (GH) or insulin-like growth factor-1 (IGF-1) to the late gestation fetus. Following acute (4 h) fetal IGF-1 infusion, placental oGLUT3 mRNA abundance was greater in growth restricted (placental embolisation) than in normal fetuses, although the reason for this difference remained equivocal. This thesis describes isolation, cloning and sequence analysis of oGLUT1 and oGLUT3 cDNAs. These studies confirmed the presence of GLUTI and GLUT3 mRNA in ovine placenta, and demonstrated ontogenetic and nutritional regulation of placental oGLUT1 and oGLUT3. In addition, these results indicated that regulation of placental oGLUTs may occur at both transcriptional and post-transcriptional levels.