絨毛細胞の糖輸送機構の解析(2 ヒト絨毛細胞の機能とその異常)
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The placenta serves as a functional organ for the exchange of various materials between mother and fetus. Glucose is one of major sources for fetar development and energy metabolism. The glucose transport is facilitated, independent of sodium ion, and mediated mainly through GLUT1 or GLUT3 in placenta. We analyzed the changes in human placental GLUT1 or GLUT3 level during pregnancy by cytochalasin B binding method, Northern blot analysis and immunoblot analysis. While the level of GLUT3 mRNA was the highest in the first trimester placenta, our findings showed that GLUT1 level increased during pregnancy in human placentas. To elucidate the potential different roles of GLUT1 and GLUT3 in human placenta, we examined the localization of these GLUTs by immunohisochemistry. The specific staining for GLUT1 by light and electron microscopy was localized mainly on the apical membrane and the basal membrane of the syncytiotrophoblast. The razor confocal microscopic findings with an anti-GLUT3 antibody showed that GLUT3 was localized mainly in the cytotrophoblast membrane. Next, we studied the changens in mouse placental GLUT1 levels in relation to changes in glucose concentration in vitro and in vivo. Treatment of mouse placental cells with 22 mM glucose resulted in the significant decrease in 2-deoxyglucose uptake, the GLUT1 mRNA levels and the GLUT1 protein levels. The similar findings were also observed in the placenta from STZ-treated diabetic mouse compared with that from control mouse. These findings suggest that glucose concentration may regulate the expression of mouse placental GLUT1. However, the levels of GLUT protein in human placenta from the patients with diabetes mellitus are not significantly higher than those from normanl pregnant women. The changes in placental GLUT1 and GLUT3 expression during trophoblast differentiation remain to be elucidated. In human trophoblasts, cAMP has been shown to stimulate not only morphological but also functional differentiation. Therefore, we performed 2-deoxyglucose uptake experiment, Northern blot analysis and immunoblot analysis in order to investigate whether the amounts of GLUT1 and GLUT3 in BeWo(choriocarcinoma)cells may change with 8-Br-cAMP induced differentiation. The stimulating effect of cAMP on glucose uptake was concentration dependent. Northern and immunoblot analyses revealed that the levels of mRNA and protein of GLUT1, but not GLUT3, were significantly increased by 8-Br-cAMP treatment. These findings suggested that 8-Br-cAMP stimulated GLUT1 expression with differentiation in BeWo cells. To analyze the regulatory mechanism of GLUT1 gene, we transfected GLUT1 promoter-CAT constructs into Rcho-1 cells. The region-76/-53 bp was important for basal GLUT1 transcription activity after deletion analysis of GLUT1 promoter. Electrophoresis mobility shift analysis revealed that Sp1 and Sp3 bound to the region. From the mutation analysis of Sp1 binding site with Sp1 or Sp3 over expression experiment, Sp1, not Sp3, may play an important role in regulating placental GLUT1 gene. In histrological data of human placenta, syncytiotrophoblast is superior to cytotrophoblast. From this phenomenon and our data, GLUT1, not GLU3, may play a mojor role in placental glucose transport in this third trimester. From our data from in vivo and in vitro mouse expriments, glucose concentration may regulate the expression of placental GLUT1. Our data showed that GLUT1 may be one of functional differentiation markers in human trophoblast and that cAMP and Sp1 may regulate the leves of placental GLUT1. To investigate the regulatory mechanism of GLUT may link to the anaklysis of the molecular mechanism for trophoblast differentiation.
- 社団法人日本産科婦人科学会の論文
- 2001-09-01
著者
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