Identification of the Notch signal in intestinal epithelial cells and the failure of the intestinal differentiation in chronic colitis.

在本研究中，我们证实了Notch信号在肠上皮细胞分化中起着至关重要的作用，并揭示了Notch信号直接作用于肠上皮细胞的几个基因，有报道称HES 1基因的缺失通过Notch信号途径引起肠上皮细胞的分化，表明Notch信号调节肠上皮细胞的去分化。然而，Notch信号传导的功能一直是未知的。因此，我们旨在阐明Notch信号在肠上皮细胞中的作用。首先，我们利用多西环素诱导Notch胞内结构域（NICD）的表达，建立了Notch信号刺激系统。此外，我们还建立了Notch信号抑制系统，使用γ-分泌酶抑制剂抑制NICD的分离。NICD的表达导致肠上皮细胞的去分化，如Mucin 2基因和Hath 1基因的表达减少，而Notch信号的抑制导致肠上皮细胞的分化，Mucin 2基因和Hath 1基因的表达增加。因此，我们利用RNA微阵列技术，鉴定了Notch信号直接作用于的几个基因，以阐明Notch在肠道中的功能;由于Notch信号调节肠道细胞系的分化，我们检测了NICD在人肠道和人结肠炎中的表达。NICD在下隐窝表达，而Hes 1和Ki-67则表达。在IBD患者肠组织中，NICD在隐窝中的表达高于正常肠组织，提示Notch信号的增强抑制了IBD患者肠上皮细胞的分化。因此，我们评估了γ-分泌酶抑制剂对肠上皮细胞的影响。Notch信号的抑制可引起小鼠杯状细胞的增加，提示Notch信号可能成为IBD治疗的新靶点。

In this study, we demonstrated that Notch signaling plays a crucial role in the differentiation of intestinal epithelial cells, and then we revealed several genes directly targeted by Notch signal in intestinal epithelial cells.It has been reported that the deficient of HES1 gene via Notch signaling caused the differentiation of intestinal epithelial cells, indicating that Notch signaling regulates the dedifferentiation of intestinal epithelial cells. However the function of Notch signaling has been unknown. Therefore, we aimed to elucidate the effect of Notch signaling in intestinal epithelial cells. First, we established the system of Notch signal stimulation, using the expression of Notch intracellular domain (NICD) induced by doxycycline. Moreover, we established the system of Notch signal inhibition, using γ-secretase inhibitor that suppresses the separation of NICD. The expression of NICD caused the dedifferentiation of intestinal epithelial cells such as the decrease of Mucin2 gene and Hath1 gene, on the contrary, the inhibition of Notch signal caused the differentiation with the increase of Mucin2 gene and Hath1 gene. Therefore, we identified several genes directly targeted by Notch signal stimulation, using RNA micro array analysis for the clarification of Notch function in intestine.Because Notch signal regulates the differentiation of intestinal cell line, we examined the expression of NICD in human intestine and human colitis. NICD was expressed at the lower crypt, whereas Hes1 and Ki-67 were expressed. Moreover, in intestine of IBD patient, NICD was expressed at higher crypt than in normal intestine, suggesting that the increase of Notch signal suppress the differentiation of the intestinal epithelial cells in IBD. So we assessed the effect of γ-secretase inhibitor on intestinal epithelial cells. The inhibition of Notch signal caused the increase of goblet cells in mouse, indicating that Notch signal may be new target for the therapy of IBD.