miRNA-148b-3p调节胎盘滋养细胞GLUT1表达与ICP子代糖代谢紊乱关系的研究
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摘要
目的研究miRNA-148b-3p在妊娠期肝内胆汁淤积症(intrahepatic cholestasis of pregnancy,ICP)孕妇胎盘组织中的表达及对滋养细胞葡萄糖转运蛋白-1(glucose transporter 1,GLUT1)的调控,探索ICP孕妇子代糖代谢异常的可能机制。方法收集2017年3月至2018年1月于四川大学华西第二医院产科行剖宫分娩的ICP孕妇30例,正常妊娠孕妇30例,分别收集胎盘组织、母血及脐血,提取胎盘组织miRNA,母血及脐血送检生化指标。采用实时荧光定量PCR验证miR-148b-3p在两组胎盘中的表达差异。通过lipofectaine3000脂质体包裹miR-148b-3p inhibitor/mimics转染人滋养细胞HTR-8,实时荧光定量PCR验证转染后细胞中miR-148b-3p的表达水平;Western blot检测转染后细胞中GLUT1的表达水平。结果 ICP孕妇母血空腹血糖(fasting blood glucose,FPG)及其胎儿脐血胰岛素水平较对照组增高(P<0.05)。ICP组胎盘组织中miR-148b-3p表达量较对照组下降(P<0.05)。转染miR-148b-3p inhibitor后,与转染inhibitor control比较,HTR-8细胞中miR-148b-3p的表达水平下降(P<0.05),GLUT1蛋白的表达量升高(P<0.05);转染miR-148b-3p mimics后,与转染mimics control比较,HTR-8细胞中miR-148b-3p的表达水平上升(P<0.05),GLUT1蛋白的表达量降低(P<0.05)。结论 miR-148b-3p可能通过对胎盘滋养细胞GLUT1表达的负向调控参与了ICP子代糖代谢异常的发生。
Objective To investigate the expression of miRNA-148b-3p and its target gene in the placenta between normal pregnant women and pregnant women with intrahepatic cholestasis of pregnancy(ICP) and to explore the possible mechanism of glucose metabolism of offspring with maternal cholestasis. Methods There were 30 cases of normal pregnant women and 30 cases of pregnant women with ICP recruited in the study, all of whom underwent cesarean delivery from Mar. 2017 to Jan. 2018. Placenta tissues, maternal blood and cord blood were collected in each case. Maternal blood and cord blood were sent for biochemical detection. miRNA of placenta tissues was extracted and qRT-PCR was used to measure the expression of miR-148b-3p in the placenta. Normal HTR-8 cells were transfected with miR-148b-3p inhibitor/mimics wrapped with lipofectaine3000. qRT-PCR was used to measure the expression of miR-148b-3p, and Western blot was used to measure the expression of glucose transporter 1(GLUT1) after transfection. Results Maternal fasting blood glucose(FPG) and its fetal cord blood insulin levels in the ICP group were significantly higher than those of control. The expression of miR-148b-3p in the placenta of ICP group was lower than that of control group(P<0.05). Compared with inhibitor control group, the expression of miR-148b-3p was decreased in HTR-8 cells transfected with miR-148b-3p inhibitor(P<0.05), while the expression of GLUT1 was increased(P<0.05). Compared with mimics control group, the expression of miR-148b-3p was increased in HTR-8 cells transfected with miR-148b-3p mimics(P<0.05), while the expression of GLUT1 was decreased(P<0.05). Conclusion miR-148b-3p might participate in glucose metabolism of offspring with maternal cholestasis through the negative regulation of GLUT1 expression in placental trophoblast cells.
Objective To investigate the expression of miRNA-148b-3p and its target gene in the placenta between normal pregnant women and pregnant women with intrahepatic cholestasis of pregnancy(ICP) and to explore the possible mechanism of glucose metabolism of offspring with maternal cholestasis. Methods There were 30 cases of normal pregnant women and 30 cases of pregnant women with ICP recruited in the study, all of whom underwent cesarean delivery from Mar. 2017 to Jan. 2018. Placenta tissues, maternal blood and cord blood were collected in each case. Maternal blood and cord blood were sent for biochemical detection. miRNA of placenta tissues was extracted and qRT-PCR was used to measure the expression of miR-148b-3p in the placenta. Normal HTR-8 cells were transfected with miR-148b-3p inhibitor/mimics wrapped with lipofectaine3000. qRT-PCR was used to measure the expression of miR-148b-3p, and Western blot was used to measure the expression of glucose transporter 1(GLUT1) after transfection. Results Maternal fasting blood glucose(FPG) and its fetal cord blood insulin levels in the ICP group were significantly higher than those of control. The expression of miR-148b-3p in the placenta of ICP group was lower than that of control group(P<0.05). Compared with inhibitor control group, the expression of miR-148b-3p was decreased in HTR-8 cells transfected with miR-148b-3p inhibitor(P<0.05), while the expression of GLUT1 was increased(P<0.05). Compared with mimics control group, the expression of miR-148b-3p was increased in HTR-8 cells transfected with miR-148b-3p mimics(P<0.05), while the expression of GLUT1 was decreased(P<0.05). Conclusion miR-148b-3p might participate in glucose metabolism of offspring with maternal cholestasis through the negative regulation of GLUT1 expression in placental trophoblast cells.
引文
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[13] 贺晶,杨慧霞,段涛,等.妊娠期肝内胆汁淤积症诊疗指南(2015).临床肝胆病杂志,2015,31(10):1575-1578.
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[20] MOUILLET JF,OUYANG Y,COYNE CB,et al.MicroRNAs in placental health and disease.Am J Obstet Gynecol,2015,213(4 Suppl):S163-S172.
[21] POIRIER C,DESGAGNE V,GUERIN R,et al.MicroRNAs in pregnancy and gestational diabetes mellitus:emerging role in maternal metabolic regulation.Curr Diab Rep,2017,17(5):35.
[22] MAVRELI D,PAPANTONIOU N,KOLIALEXI A.miRNAs in pregnancy-related complications:an update.Expert Rev Mol Diagns,2018,18(7):587-589.
[23] CHEN Y,SONG YX,WANG ZN.The microRNA-148/152 family:multi-faceted players.Mol Cancer,2013,12:43.
[24] MIAO C,ZHANG J,ZHAO K,et al.The significance of microRNA-148/152 family as a prognostic factor in multiple human malignancies:a meta-analysis.Oncotarget,2017,8(26):43344-43355.
[25] ZHANG H,YE Q,DU Z,et al.MiR-148b-3p inhibits renal carcinoma cell growth and pro-angiogenic phenotype of endothelial cell potentially by modulating FGF2.Biomed Pharmacother,2018,107:359-367[2018-11-20].https://doi.org/10.1016/j.biopha.2018.07.054.
[26] WANG Y,LI J,KUANG D,et al.miR-148b-3p functions as a tumor suppressor in GISTs by directly targeting KIT.Cell Commun Signal,2018,16(1):16.
[27] LI X,JIANG M,CHEN D,et al.miR-148b-3p inhibits gastric cancer metastasis by inhibiting the Dock6/Rac1/Cdc42 axis.J Exp Clin Cancer Res,2018,37(1):71.
[28] DING X,LIU J,LIU T,et al.miR-148b inhibits glycolysis in gastric cancer through targeting SLC2A1.Cancer Med,2017,6(6):1301-1310.
[29] AGARWAL V,BELL GW,NAM JW,et al.Predicting effective microRNA target sites in mammalian mRNAs.Elife,2015,4:e05005[2018-11-20].https://doi.org/10.7554/eLife.05005.001.
[30] BARKER DJ.The origins of the developmental origins theory.J Intern Med,2007,261(5):412-417.
[31] SILVEIRA PP,PORTELLA AK,GOLDANI MZ,et al.Developmental origins of health and disease (DOHaD).J Pediatr (Rio J),2007,83(6):494-504.
[32] MEAS T.Fetal origins of insulin resistance and the metabolic syndrome:a key role for adipose tissue?Diabetes Metab,2010,36(1):11-20.
[33] KANAKA-GANTENBEIN C.Fetal origins of adult diabetes.Ann N Y Acad Sci,2010,1205:99-105.
[34] HANSON MA,GLUCKMAN PD.Developmental origins of health and disease:new insights.Basic Clin Pharmacol Toxicol,2008,102(2):90-93.
[2] WIKSTR?M SHEMER E,MARSCHALL HU,LUDVIGSSON JF,et al.Intrahepatic cholestasis of pregnancy and associated adverse pregnancy and fetal outcomes:a 12-year population-based cohort study editorial comment.BJOG,2013,120(6):717-723.
[3] KENYON AP,PIERCY CN,GIRLING J,et al.Obstetric cholestasis,outcome with active management:a series of 70 cases.BJOG,2002,109(3):282-288.
[4] REZAI S,LAM J,HENDERSON CE.Intrahepatic cholestasis of pregnancy:maternal and fetal outcomes associated with elevated bile acid levels.Am J Obstet Gynecol,2015,213(1):114[2018-11-14].https://doi.org/10.1016/j.ajog.2015.03.040.
[5] MARTINEAU MG,RAKER C,DIXON PH,et al.The metabolic profile of intrahepatic cholestasis of pregnancy is associated with impaired glucose tolerance,dyslipidemia,and increased fetal growth.Diabetes Care,2015,38(2):243-248.
[6] MARTINEAU M,RAKER C,POWRIE R,et al.Intrahepatic cholestasis of pregnancy is associated with an increased risk of gestational diabetes.Eur J Obstet Gynecol Reprod Biol,2014,176:80-85[2018-11-14].https://doi.org/10.1016/j.ejogrb.2013.12.037.
[7] PAPACLEOVOULOU G,ABU-HAYYEH S,NIKOLOPOULOU E,et al.Maternal cholestasis during pregnancy programs metabolic disease in offspring.J Clin Invest,2013,123(7):3172-3181.
[8] BAUMANN MU,DEBORDE S,ILLSLEY NP.Placental glucose transfer and fetal growth.Endocrine,2002,19(1):13-22.
[9] ACOSTA O,RAMIREZ VI,LAGER S,et al.Increased glucose and placental GLUT-1 in large infants of obese nondiabetic mothers.Am J Obstet Gynecol,2015,212(2):227.e1-e7[2018-11-14].https://doi.org/10.1016/j.ajog.2014.08.009.
[10] WEI W,HU YY.Expression of hypoxia-regulated genes and glycometabolic genes in placenta from patients with intrahepatic cholestasis of pregnancy.Placenta,2014,35(9):732-736.
[11] ALVAREZ-GARCIA I,MISKA EA.MicroRNA functions in animal development and human disease.Development,2005,132(21):4653-4662.
[12] KROL J,LOEDIGE I,FILIPOWICZ W.The widespread regulation of microRNA biogenesis,function and decay.Nat Rev Genet,2010,11(9):597-610.
[13] 贺晶,杨慧霞,段涛,等.妊娠期肝内胆汁淤积症诊疗指南(2015).临床肝胆病杂志,2015,31(10):1575-1578.
[14] ILLSLEY NP.Glucose transporters in the human placenta.Placenta,2000,21(1):14-22.
[15] BARTA E,DRUGAN A.A clinical study which relates to a theoretical simulation of the glucose transport in the human placenta under various diabetic conditions.J Perinat Med,2016,44(4):405-410.
[16] BAUMANN MU,ZAMUDIO S,ILLSLEY NP.Hypoxic upregulation of glucose transporters in BeWo choriocarcinoma cells is mediated by hypoxia-inducible factor-1.Am J Physiol Cell Physiol,2007,293(1):C477-C485.
[17] JANSSON T,EKSTRAND Y,WENNERGREN M,et al.Placental glucose transport in gestational diabetes mellitus.Am J Obstet Gynecol,2001,184(2):111-116.
[18] SHI Z,ZHAO C,GUO X,et al.Differential expression of microRNAs in omental adipose tissue from gestational diabetes mellitus subjects reveals miR-222 as a regulator of ERalpha expression in estrogen-induced insulin resistance.Endocrinology,2014,155(5):1982-1990.
[19] RAO ZZ,ZHANG XW,DING YL,et al.miR-148a-mediated estrogen-induced cholestasis in intrahepatic cholestasis of pregnancy:role of PXR/MRP3.PLoS One,2017,12(6):e0178702[2018-11-20].https://doi.org/10.1371/journal.pone.0178702.
[20] MOUILLET JF,OUYANG Y,COYNE CB,et al.MicroRNAs in placental health and disease.Am J Obstet Gynecol,2015,213(4 Suppl):S163-S172.
[21] POIRIER C,DESGAGNE V,GUERIN R,et al.MicroRNAs in pregnancy and gestational diabetes mellitus:emerging role in maternal metabolic regulation.Curr Diab Rep,2017,17(5):35.
[22] MAVRELI D,PAPANTONIOU N,KOLIALEXI A.miRNAs in pregnancy-related complications:an update.Expert Rev Mol Diagns,2018,18(7):587-589.
[23] CHEN Y,SONG YX,WANG ZN.The microRNA-148/152 family:multi-faceted players.Mol Cancer,2013,12:43.
[24] MIAO C,ZHANG J,ZHAO K,et al.The significance of microRNA-148/152 family as a prognostic factor in multiple human malignancies:a meta-analysis.Oncotarget,2017,8(26):43344-43355.
[25] ZHANG H,YE Q,DU Z,et al.MiR-148b-3p inhibits renal carcinoma cell growth and pro-angiogenic phenotype of endothelial cell potentially by modulating FGF2.Biomed Pharmacother,2018,107:359-367[2018-11-20].https://doi.org/10.1016/j.biopha.2018.07.054.
[26] WANG Y,LI J,KUANG D,et al.miR-148b-3p functions as a tumor suppressor in GISTs by directly targeting KIT.Cell Commun Signal,2018,16(1):16.
[27] LI X,JIANG M,CHEN D,et al.miR-148b-3p inhibits gastric cancer metastasis by inhibiting the Dock6/Rac1/Cdc42 axis.J Exp Clin Cancer Res,2018,37(1):71.
[28] DING X,LIU J,LIU T,et al.miR-148b inhibits glycolysis in gastric cancer through targeting SLC2A1.Cancer Med,2017,6(6):1301-1310.
[29] AGARWAL V,BELL GW,NAM JW,et al.Predicting effective microRNA target sites in mammalian mRNAs.Elife,2015,4:e05005[2018-11-20].https://doi.org/10.7554/eLife.05005.001.
[30] BARKER DJ.The origins of the developmental origins theory.J Intern Med,2007,261(5):412-417.
[31] SILVEIRA PP,PORTELLA AK,GOLDANI MZ,et al.Developmental origins of health and disease (DOHaD).J Pediatr (Rio J),2007,83(6):494-504.
[32] MEAS T.Fetal origins of insulin resistance and the metabolic syndrome:a key role for adipose tissue?Diabetes Metab,2010,36(1):11-20.
[33] KANAKA-GANTENBEIN C.Fetal origins of adult diabetes.Ann N Y Acad Sci,2010,1205:99-105.
[34] HANSON MA,GLUCKMAN PD.Developmental origins of health and disease:new insights.Basic Clin Pharmacol Toxicol,2008,102(2):90-93.