Molecular Mechanisms for Pancreatic Islet Disfunction in Development of Diabetes Mellitus-a study using WFS1 knockout mice

WFS1缺陷型胰岛素瘤细胞系的建立为了研究wfs1基因缺失对同源β细胞群的影响，我们将wfs1^-lt；-/-&gt；小鼠与在胰岛素启动子作用下表达猴病毒40大T抗原的β小鼠杂交，建立了WFS1-细胞系。我们发现高糖引起的胰岛素分泌受损，这可以通过腺病毒介导的野生型WFS1的表达而得到恢复，但在Wolfram综合征患者中发现的突变的WFS1蛋白没有恢复。WFS1蛋白在细胞内钙稳态中的作用。我们分析了Wfs1基因敲除和过表达的HEK293细胞，发现Wfs1蛋白通过正向调节内质网钙摄取来调节[Ca^&lt；2+&gt；]_&lt；er&gt；，这与内质网应激下β细胞内[Ca^&lt；2+&gt；]_&lt；细胞反应的变化有关。WFS1缺陷的胰岛表达谱显示4E-BPI表达增加。我们发现，在内质网应激下，糖尿病小鼠胰岛中4E-BP1的表达也增强。4E-BP1的诱导由转录因子ATF4介导。4E-BP1缺失的MIN6β-细胞更容易受到内质网应激诱导的细胞凋亡，C/EBP同源蛋白的诱导作用更强。此外，在内质网应激的小鼠糖尿病模型中，Eif4ebp1缺失加剧了高血糖，并加速了(3-细胞衰竭)。因此，4E-BP1的诱导对于内质网应激下的3-细胞是一个促进生存的信号，也是糖尿病的潜在治疗靶点。WFS1缺乏的胰岛分泌的高血糖素增加。丙酮酸此前被证明可以刺激胰升糖素的分泌。在这项研究中，我们发现在WFS1缺陷的胰岛中，α细胞数量增加，丙酮酸刺激的胰升糖素分泌增加。在Wolfram综合征的小鼠模型中，多种异常被认为是糖尿病发生的原因。

Establishment of WFS1-deficient insulinoma cell lines. To examine the influence of WFS1-deficiency specifically in a homogenous β-cell population, β-cell lines were established by crossing wfs1^<-/-> mice with IT6 mice expressing simian virus 40 large T antigen under the insulin promoter. We found impaired insulin secretion in response to high glucose, which was recovered by adenovirus-mediated expression of wild-type WFS 1 but not of mutant WFS1 proteins found in Wolfram-syndrome patients.Role of WFS1 protein in intracellular calcium homeostasis. We analyzed WFS1-knockdown and-overexpressing HEK293 cells, and found that WFS1 protein modulates [Ca^<2+>]_<er> by positively regulating the ER Ca^<2+> uptake, which is associated with changes in the [Ca^<2+>]_<cyt> response evoked by Ca^<2+> store depletion.Identification of the translational suppressor 4E-BP1 as a pro-survival factor in β-cells under ER stress. Expression profiling in WFS1-deficient islets revealed increased 4E-BPI expression. We found that expression of 4E-BP1 was also enhanced in islets under ER stress in mouse models of diabetes. 4E-BP1 induction was found to be mediated by the transcription factor ATF4. 4E-BP1-deficient MIN6 β-cells were more vulnerable to apoptosis triggered by ER stress, with greater induction of C/EBP homologue protein. Furthermore, Eif4ebp1 deletion exacerbated hyperglycemia, with accelerated (3-cell failure, in mouse diabetes models with ER stress. Thus, 4E-BP1 induction is a pro-survival signal for (3-cells under ER stress and a potential therapeutic target for diabetes.Increased glucagon secretion from WFS1-deficient islets. Pyruvate was previously shown to stimulate glucagon secretion. In this study, we found increased alpha cell numbers and enhanced pyruvate-stimulated glucagon secretion in WFS1-deficient islets.Multiple abnormalities are suggested to contribute development of diabetes in the mouse model of Wolfram syndrome.