Effect of daf2 on insulin secreating cells and target cells

(DAF2在胰岛素分泌细胞中的作用)为了检测DAF2基因在胰岛素分泌细胞中的作用，我们使用了表达DAF2基因的胰腺细胞。已有报道表明，DAF2基因突变可抑制细胞内的细胞凋亡。虽然脑膜素是一种核蛋白，并与DAF2相互作用，但它在包括胰岛素瘤在内的男性1相关内分泌肿瘤的发病机制中的作用尚不清楚。在这项研究中，我们研究了DAF2和薄荷素对人胰岛素产生的影响。将该表达载体与大鼠胰岛素基因5‘侧翼区(PINS-Luc)235bp的荧光素酶报告基因(PINS-Luc)共转染INS-1细胞。胰岛素基因启动子活性在共转染组细胞中显著低于模型转染组。将该表达载体导入大鼠胰岛素瘤细胞株INS-1，用稳定表达的细胞克隆检测胰岛素的分泌。在高表达Menin的细胞中，胰岛素的分泌和DNA的胸腺嘧啶核苷掺入都受到抑制。另一方面，与模型细胞相比，转染膜蛋白的细胞凋亡率增加。(DAF2在血管平滑肌细胞清道夫受体基因表达中的作用)Gas6是由生长停滞基因6编码的分泌蛋白，最初被鉴定为Ax1受体酪氨酸激酶的配体，并上调A类清道夫受体(SRA)的表达，参与人血管平滑肌细胞(HvCs)动脉粥样硬化和泡沫细胞的形成。以上结果表明，DAF2可诱导Gas6介导的血管内皮细胞凋亡。

(Role of daf2 on insulin-secreting cells)To examine the role of daf2 gene on insulin-secreting cells, we used the pancreatic-cells expressing daf2 gene. Previous report indicated that the mutation of daf2 induce the inhibitory effect of apoptosis in the cells. Although menin is a nuclear protein and interacts with daf2, its role in the pathogenesis of MEN 1-related endocrine tumors including insulinoma remains unknown. In this study, we examined the effects of daf2 and menin on the human insulin production. INS-1 cells were then co-transfected with the menin expression vector and a luciferase reporter gene linked to 235 bp from the rat insulin gene 5'-flanking region (pINS-LUC). Promoter activity of the insulin gene was significantly decreased in the co-transfected cells as compared to mock-transfected controls. Rat insulinoma cell line, INS-1 were transfected with the menin expression vector and stably transfected cell clones were used to examine the insulin secretion. In the cells with high level expression of menin, both insulin secretion and thymidine incorporation into DNA were inhibited as compared to mock-transfected cells. On the other hand, the rate of apoptosis of menin-transfected cells was increased as compared to mock-transfeced cells.(The role of daf2 on scavenger receptor gene expression in vascular smooth muscle cell)The secreted protein, Gas6 is encoded by the growth arrest-specific gene 6, has been originally identified as the ligand of the Axl receptor tyrosine kinase, and upregulated the expression of the class A scavenger receptor (SRA) in the pathogenesis of atherosclerosis and foam cell formation in human vascular smooth muscle cells (HVSMCs). These results indicated that daf2 induced the apoptosis in the smooth muscle cell mediated by Gas6.