Notch signalling suppresses apoptosis in adult human and mouse pancreatic islet cells
Notch signalling suppresses apoptosis in adult human and mouse pancreatic islet cells
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DOI：
10.1007/s00125-007-0835-5
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发表时间：
2007-12-01
期刊：
影响因子：
8.2
通讯作者：
Johnson, J. D.
中科院分区：
文献类型：
--
作者：
Dror, V.;Nguyen, V.;Johnson, J. D.
目的/假设糖尿病的发病机制和胰岛移植的成功取决于胰腺β细胞命运的控制。Notch信号通路是必不可少的正常产前胰腺发育，但这种基因网络在成人islets的存在和功能得到了少得多的attention.Methods Notch信号成分的存在进行了评估，在体外使用RT-PCR，蛋白质印迹和免疫荧光。改变Notch信号对胰岛素分泌和程序性细胞死亡的功能后果examined.Results成年小鼠胰岛，人类胰岛和小鼠胰岛素瘤MIN 6细胞具有Notch途径的关键组成部分。RT-PCR、蛋白质印迹和免疫荧光表明Notch靶基因neurogenin 3（Ngn 3，也称为Neurog 3）也存在于成年胰岛细胞中。用N-[N-（3，5-二氟苯乙酰基-L-丙氨酰）] S-苯基甘氨酸叔丁酯（DAPT）抑制Notch信号传导增加成年胰岛中Ngn 3 mRNA表达和蛋白水平。激活的notch同源物1（NOTCH 1）蛋白水平降低血清撤出后，以及与磷脂酰肌醇3-激酶抑制剂，或羟基2-萘甲基膦酸，胰岛素受体抑制剂治疗后。虽然在DAPT中培养的胰岛没有表现出胰岛素分泌缺陷，表明分化没有改变，但抑制γ-分泌酶依赖性Notch活化导致MIN 6细胞和人胰岛中caspase-3依赖性细胞凋亡的剂量依赖性增加。相反，γ-分泌酶的过度活性导致切割的NOTCH 1的积累和保护免于apoptosis.Conclusions/interpretation这些结果一起表明，Notch/Ngn 3信号网络是完整的，在成年胰岛中起作用。该途径代表了用于调节糖尿病中的β细胞命运、胰岛移植和在体外获得β细胞替代物的努力的有吸引力的靶点。
Aims/hypothesis The pathogenesis of diabetes and the success of islet transplantation depend on the control of pancreatic beta cell fate. The Notch signalling pathway is essential for normal prenatal pancreatic development, but the presence and function of this gene network in adult islets has received much less attention.Methods The presence of Notch signalling components was assessed in vitro using RT-PCR, western blotting and immunofluorescence. The functional consequences of altering Notch signalling on insulin secretion and programmed cell death were examined.Results Adult mouse islets, human islets and mouse insulinoma MIN6 cells possess key components of the Notch pathway. RT-PCR, western blotting and immunofluorescence indicated that the Notch target gene, neurogenin3 (Ngn3, also known as Neurog3), is also present in adult islet cells. Inhibiting Notch signalling with N-[N-(3,5-difluorophenacetyl-L-alanyl)]S-phenylglycine t-butyl ester (DAPT) increased Ngn3 mRNA expression and protein levels in adult islets. The activated notch homologue 1 (NOTCH1) protein level was decreased upon serum withdrawal, as well as after treatment with a phosphatidylinositol 3-kinase inhibitor, or hydroxy2- naphthalenylmethylphosphonic acid, an insulin receptor inhibitor. While islets cultured in DAPT did not exhibit defects in insulin secretion, indicating that differentiation is unaltered, inhibiting gamma-secretase-dependent Notch activation led to a dose-dependent increase in caspase-3- dependent apoptosis in both MIN6 cells and human islets. Conversely, gamma-secretase overactivity resulted in an accumulation of cleaved NOTCH1 and protection from apoptosis.Conclusions/interpretation Together these results show that the Notch/Ngn3 signalling network is intact and functional in adult islets. This pathway represents an attractive target for modulating beta cell fate in diabetes, islet transplantation and efforts to derive beta cell surrogates in vitro.