Center for Fetal Cellular and Molecular Therapy, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, United States of America.
PLoS One. 2013 Sep 3;8(9):e74632. doi: 10.1371/journal.pone.0074632. eCollection 2013.
Previous work in our laboratory demonstrated that over-expression of human insulin-like growth factor -1 (hIGF-1) in the placenta corrects fetal weight deficits in mouse, rat, and rabbit models of intrauterine growth restriction without changes in placental weight. The underlying mechanisms of this effect have not been elucidated. To investigate the effect of intra-placental IGF-1 over-expression on placental function we examined glucose transporter expression and localization in both a mouse model of IUGR and a model of human trophoblast, the BeWo Choriocarcinoma cell line.
At gestational day 18, animals were divided into four groups; sham-operated controls, uterine artery branch ligation (UABL), UABL+Ad-hIGF-1 (10(8) PFU), UABL+Ad-LacZ (10(8) PFU). At gestational day 20, pups and placentas were harvested by C-section. For human studies, BeWo choriocarcinoma cells were grown in F12 complete medium +10%FBS. Cells were incubated in serum-free control media ± Ad-IGF-1 or Ad-LacZ for 48 hours. MOIs of 10∶1 and 100∶1 were utilized. The RNA, protein expression and localization of glucose transporters GLUT1, 3, 8, and 9 were analyzed by RT-PCR, Western blot and immunohistochemistry.
In both the mouse placenta and BeWo, GLUT1 regulation was linked to altered protein localization. GLUT3, localized to the mouse fetal endothelial cells, was reduced in placental insufficiency but maintained with Ad-I GF-1 treatment. Interestingly, GLUT8 expression was reduced in the UABL placenta but up-regulated following Ad-IGF-1 in both mouse and human systems. GLUT9 expression in the mouse was increased by Ad-IGF-1 but this was not reflected in the BeWo, where Ad-IGF-1 caused moderate membrane relocalization.
Enhanced GLUT isoform transporter expression and relocalization to the membrane may be an important mechanism in Ad-hIGF-1mediated correction of placental insufficiency.
本实验室先前的工作表明，在宫内生长受限的小鼠、大鼠和兔模型中，过表达人胰岛素样生长因子-1（hIGF-1）可纠正胎儿体重不足，而胎盘重量不变。其潜在机制尚未阐明。为了研究胎盘内 IGF-1 过表达对胎盘功能的影响，我们在宫内生长受限的小鼠模型和人类滋养层细胞模型——BeWo 绒毛膜癌细胞系中检查了葡萄糖转运体的表达和定位。
在妊娠第 18 天，将动物分为四组：假手术对照组、子宫动脉分支结扎组（UABL）、UABL+Ad-hIGF-1（10^8 PFU）、UABL+Ad-LacZ（10^8 PFU）。在妊娠第 20 天，通过剖腹产收获幼仔和胎盘。对于人类研究，BeWo 绒毛膜癌细胞在 F12 完全培养基+10%FBS 中生长。将细胞在无血清对照培养基中孵育，±Ad-IGF-1 或 Ad-LacZ 48 小时。使用 MOI 为 10∶1 和 100∶1。通过 RT-PCR、Western blot 和免疫组织化学分析葡萄糖转运体 GLUT1、3、8 和 9 的 RNA、蛋白表达和定位。
在小鼠胎盘和 BeWo 中，GLUT1 的调节与改变的蛋白定位有关。GLUT3 定位于胎儿内皮细胞，在胎盘功能不全时减少，但用 Ad-IGF-1 治疗后得到维持。有趣的是，GLUT8 在 UABL 胎盘中表达减少，但在小鼠和人类系统中，在用 Ad-IGF-1 处理后上调。在小鼠中，Ad-IGF-1 增加了 GLUT9 的表达，但在 BeWo 中没有反映出来，在 BeWo 中，Ad-IGF-1 导致中等程度的膜再定位。
增强的 GLUT 同工型转运体表达和向膜的再定位可能是 Ad-hIGF-1 介导的纠正胎盘功能不全的重要机制。
