P-糖蛋白过表达对INS-1和大鼠胰岛β细胞分化的影响
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摘要
目的探讨P-糖蛋白过表达对大鼠胰岛β细胞系INS-1细胞和大鼠胰岛β细胞分化的影响。方法分别构建INS-1细胞(Ad-abcb1b腺病毒感染)和SD大鼠(Ins2-abcb1b随机插入)P-糖蛋白过表达模型。以阴性对照病毒和野生型SD大鼠为对照组,P-糖蛋白过表达组和转基因SD大鼠为实验组,萃取细胞和胰岛总RNA,定量PCR分别检测INS-1细胞和胰岛组织Abcb1b、Pdx1、Ins1、Ins2、Mafa、Mafb、Ngn3、Nkx6.1、Nkx2.2、Neurod1、Pax6等基因的表达。结果与对照组相比,P-糖蛋白过表达后,INS-1细胞Ngn3、Mafb转录水平升高(P<0.05),Mafa转录水平降低(P<0.001),其余基因表达差异无统计学意义。在野生型和转基因大鼠胰岛组织中,各基因转录水平差异均无统计学意义(P>0.05)。结论 P-糖蛋白过表达可能使INS-1细胞去分化,而对胰岛β细胞的分化无明显影响。
Objective To investigate the effect of P-glycoprotein overexpression on the differentiation of INS-1 cells and pancreatic β cells. Methods INS-1 cells were infected with adenovirus overexpressing P-glycoprotein. Ins2-abcblb gene was randomly inserted into the rat islets, and P-glycoprotein overexpression model was successfully constructed. Empty adenovirus and wild-type SD rats were used as control groups. INS-1 cells and P-glycoprotein overexpression rats were used as experimental group. Total RNA was extracted both in INS-1 cells and rat islets. Transcription factors such as Abcb1 b,Pdxl, Ins1, Ins2, Mafa, Mafb, Ngn3, Nkx6.1, Nkx2.2, Neurodl and Pax6 were detected by real-time quantitative PCR.Results Compared with the control group, the transcription levels of Ngn3 and Mafb in INS-1 cells increased(P < 0.05),while Mafa decreased(P < 0.001). No significance was detected in other transcription factors. Unlike INS-1 cells, all of the transcription factors of islets were not affected(P > 0.05). Conclusion The overexpression of P-glycoprotein may dedifferentiate INS-1 cells, but has no effect on the differentiation of rat pancreatic beta cells.
Objective To investigate the effect of P-glycoprotein overexpression on the differentiation of INS-1 cells and pancreatic β cells. Methods INS-1 cells were infected with adenovirus overexpressing P-glycoprotein. Ins2-abcblb gene was randomly inserted into the rat islets, and P-glycoprotein overexpression model was successfully constructed. Empty adenovirus and wild-type SD rats were used as control groups. INS-1 cells and P-glycoprotein overexpression rats were used as experimental group. Total RNA was extracted both in INS-1 cells and rat islets. Transcription factors such as Abcb1 b,Pdxl, Ins1, Ins2, Mafa, Mafb, Ngn3, Nkx6.1, Nkx2.2, Neurodl and Pax6 were detected by real-time quantitative PCR.Results Compared with the control group, the transcription levels of Ngn3 and Mafb in INS-1 cells increased(P < 0.05),while Mafa decreased(P < 0.001). No significance was detected in other transcription factors. Unlike INS-1 cells, all of the transcription factors of islets were not affected(P > 0.05). Conclusion The overexpression of P-glycoprotein may dedifferentiate INS-1 cells, but has no effect on the differentiation of rat pancreatic beta cells.
引文
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[14]Li N,Yang Z,Li Q,et al.Ablation of somatostatin cells leads to impaired pancreatic islet function and neonatal death in rodents[J].Cell Death Dis,2018,9(6):682.doi:10.1038/s41419-018-0741-4.
[2]Su L,Cheng YC,Lee WM,et al.Abcb1a and Abcb1b genes function differentially in blood-testis barrier dynamics in the rat[J].Cell Death Dis,2017,8(9):e3038.doi:10.1038/cddis.2017.435.
[3]Tang YZ,Li DQ,Sun FJ,et al.P-glycoprotein regulating biphasic insulin secretion in rat pancreatic beta cells[J].Chin Med J(Engl),2009,122(21):2587-2592.
[4]Talchai C,Xuan S,Lin HV,et al.Pancreaticβcell dedifferentiation as a mechanism of diabeticβcell failure[J].Cell,2012,150(6):1223-1234.doi:10.1016/j.cell.2012.07.029.
[5]Lee JS,Jung ID,Lee CM,et al.Venlafaxine inhibits the development and differentiation of dendritic cells through the regulation of P-glycoprotein[J].Int Immunopharmacol,2011,11(9):1348-1357.doi:10.1016/j.intimp.2011.04.019.
[6]Nishimura W,Takahashi S,Yasuda K.MafA is critical for maintenance of the mature beta cell phenotype in mice[J].Diabetologia,2015,58(3):566-574.doi:10.1007/s00125-014-3464-9.
[7]Samant MD,Jakson CM,Felix CL,et al.Multi-drug resistance ABC transporter inhibition enhances murine ventral prostate stem/progenitor cell differentiation[J].Stem Cells Dev,2015,24(10):1236-1251.doi:10.1089/scd.2014.0293.
[8]Duan FF,Liu JH,March JC.Engineered commensal bacteria reprogram intestinal cells into glucose-responsive insulin-secreting cells for the treatment of diabetes[J].Diabetes,2015,64(5):1794-1803.doi:10.2337/db14-0635.
[9]Shimoda M,Kanda Y,Hamamoto S,et al.The human glucagonlike peptide-1 analogue liraglutide preserves pancreatic beta cells via regulation of cell kinetics and suppression of oxidative and endoplasmic reticulum stress in a mouse model of diabetes[J].Diabetologia,2011,54(5):1098-1108.doi:10.1007/s00125-011-2069-9.
[10]Suarez-Pinzon WL,Power RF,Yan Y,et al.Combination therapy with glucagon-like peptide-1 and gastrin restores normoglycemia in diabetic NOD mice[J].Diabetes,2008,57(12):3281-3288.doi:10.2337/db08-0688.
[11]Sasaki S,Miyatsuka T,Matsuoka TA,et al.Activation of GLP-1and gastrin signalling induces in vivo reprogramming of pancreatic exocrine cells into beta cells in mice[J].Diabetologia,2015,58(11):2582-2591.doi:10.1007/s00125-015-3728-z.
[12]Chung CH,Hao E,Piran R,et al.Pancreatic beta-cell neogenesis by direct conversion from mature alpha-cells[J].Stem cells,2010,28(9):1630-1638.doi:10.1002/stem.482.
[13]Thorel F,Nepote V,Avril I,et al.Conversion of adult pancreatic alpha-cells to beta-cells after extreme beta-cell loss[J].Nature,2010,464(7292):1149-1154.doi:10.1038/nature08894.
[14]Li N,Yang Z,Li Q,et al.Ablation of somatostatin cells leads to impaired pancreatic islet function and neonatal death in rodents[J].Cell Death Dis,2018,9(6):682.doi:10.1038/s41419-018-0741-4.