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The effects of imeglimin, a novel anti-diabetes agent, on β-cell function remain unclear. Here, we unveiled the impact of imeglimin on β-cell survival. Treatment with imeglimin augmented mitochondrial function, enhanced insulin secretion, promoted β-cell proliferation, and improved β-cell survival in mouse islets. Imeglimin upregulated the expression of endoplasmic reticulum (ER)-related molecules including Chop (Ddit3), Gadd34 (Ppp1r15a), Atf3, and Sdf2l1, and decreased eIF2α phosphorylation, after treatment with thapsigargin, and restored global protein synthesis in β-cells under ER stress. Imeglimin failed to protect ER stress-induced β-cell apoptosis in CHOP-deficient islets or in the presence of GADD34 inhibitor. Treatment with imeglimin showed a significant decrease in the number of apoptotic β-cells and increased β-cell mass in Akita mice. Imeglimin also protected against β-cell apoptosis in both human islets and human pluripotent stem cell (hPSC)-derived β-like cells. Taken together, imeglimin modulates ER homeostasis pathway, which results in the prevention of β-cell apoptosis both in vitro and in vivo.

Metformin is becoming a popular treatment before and during pregnancy but current literature on in utero exposure to metformin lacks long-term clinical trials and mechanistic studies. Current literature on the effects of metformin on mature pancreatic β cells highlighted its dual, opposing, protective or inhibitory, effects depending on metabolic environments. However, the impact of metformin on developing human pancreatic β cells remains unknown. Here, we investigated the potential effects of metformin exposure on human pancreatic β cell development and function in vitro. In the absence of metabolic challenges such as high levels of glucose and fatty acids, metformin exposure impaired the development and function of pancreatic β cells, with downregulation of pancreatic genes and dysfunctional mitochondrial respiration. It also affected the insulin secretion function of pancreatic β cells. These findings call for further in-depth evaluation of the exposure of human embryonic and fetal tissue during pregnancy to metformin, and its implications on long-term offspring health.

Neonatal diabetes affects 1 in 90,000-200,000 live births. Permanent neonatal diabetes mellitus (PNDM) presents itself in the first few months of life, and patients require treatment henceforth. PNDM can be caused by mutations in the insulin gene, INS, most of which are heterozygous. This study involves two INS gene mutations: C109Y and G32V. The first mutation was reported in a Singaporean family, and the latter has not been reported yet. Patients with either mutation are unable to secrete insulin despite having one wild type (WT) allele. We hypothesize that both C109Y and G32V mutations result in misfolding of the proinsulin protein, leading to β-cell dysfunction. The misfolded proinsulin is then able to exert its dominant negative effect on WT proinsulin, either directly through heteromerization, or indirectly through altering endoplasmic reticulum (ER) integrity, preventing insulin secretion and eventually insulin synthesis and/or even β-cell death. We computationally modelled both C109Y and G32V proinsulin structures, and found that both mutations result in conformational changes that likely affect further insulin processing. We also overexpressed C109Y and G32V preproinsulin in a mouse insulinoma cell line, MIN6. Analysis of electron microscopy images revealed differences in the size and number of insulin granules. In addition, we obtained skin fibroblasts from patients with the C109Y mutation and their family members, reprogrammed them into induced pluripotent stem cells (iPSCs), and characterized them. We differentiated the iPSCs into β-like cells and saw no difference in size between WT and mutant cells. Current efforts are underway to determine the impact of the mutant insulin gene in the iPSC-derived β-like cells. Our findings will reveal mechanistic insights into this neonatal diabetes condition caused by the mutant insulin gene and potentially identify therapeutic approaches to reinstate the function of the existing wild type insulin gene. Disclosure N. Amirruddin: None. Y. Tan: None. C.S. Verma: Other Relationship; Self; sinopsee therapeutics. D. Gardner: None. Y. Bee: None. S. Hoon: None. A. Teo: None. Funding Institute of Molecular and Cell Biology; Khoo Teck Puat Hospital; National University Health System; Agency for Science, Technology and Research; National Medical Research Council; Lee Foundation; Skin Research Institute of Singapore

The differentiation of human pluripotent stem cells (hPSCs) into desired cell types such as the pancreatic cells involve cellular proliferation and apoptosis during cell fate transitions. Although different rates of cell growth and death are typically observed during differentiation, its implications for pancreatic tissue formation in humans remain unclear. Here, we profiled the expression of BCL-2 family of proteins during pancreatic specification from hPSCs and observed an upregulation of BCL- XL, downregulation of BAK and corresponding downregulation of cleaved CASP3 representative of apoptosis. The inhibition or downregulation of BCL-xL reciprocally increased apoptosis and resulted in a decreased gene expression of pancreatic markers despite a compensatory increase in companion anti-apoptotic protein BCL2. RNA-Seq analyses revealed a downregulation of multiple metabolic genes upon inhibition of BCL-xL function. Bioenergetics assays then revealed a broad downregulation of both glycolysis and oxidative phosphorylation when BCL-xL function was inhibited. In conclusion, we have identified anti-apoptotic protein BCL-xL to be necessary for suppressing the expression of pro-apoptotic protein BAK to facilitate proper pancreas formation in human cells. This is the first report that links a member of the BCL-2 family with pancreatic specification. Therefore, modulation of these two BCL-2 family of proteins can potentially increase the survival and robustness of pancreatic progenitors that ultimately defines human pancreatic beta cell mass and function. Disclosure L. Loo: None. A. Teo: None. S. Ghosh: None. A. Soetedjo: None. L. Nguyen: None. V.G. Krishnan: None. S. Hoon: None. Funding Agency for Science, Technology and Research; Institute of Molecular and Cell Biology

Type 1 diabetes is characterized by infiltration of pancreatic islets with immune cells, leading to insulin deficiency. Although infiltrating immune cells are traditionally considered to negatively impact β-cells by promoting their death, their contribution to proliferation is not fully understood. Here we report that islets exhibiting insulitis also manifested proliferation of β-cells that positively correlated with the extent of lymphocyte infiltration. Adoptive transfer of diabetogenic CD4+ and CD8+ T cells, but not B cells, selectively promoted β-cell proliferation in vivo independent from the effects of blood glucose or circulating insulin or by modulating apoptosis. Complementary to our in vivo approach, coculture of diabetogenic CD4+ and CD8+ T cells with NOD.RAG1−/− islets in an in vitro transwell system led to a dose-dependent secretion of candidate cytokines/chemokines (interleukin-2 [IL-2], IL-6, IL-10, MIP-1α, and RANTES) that together enhanced β-cell proliferation. These data suggest that soluble factors secreted from T cells are potential therapeutic candidates to enhance β-cell proliferation in efforts to prevent and/or delay the onset of type 1 diabetes.