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Pancreatic islet cellular oscillators have been recently demonstrated to play a critical role in regulating glucose homeostasis in physiological conditions. However, little is known about changes of molecular clockwork upon islet dysfunction, and its role in beta (b)-cell regeneration. We aimed to assess diurnal regulation of b-cell proliferation and circadian transcriptome of alpha (a)- and b-cells following massive b-cell ablation in rtTA/TET-DTA mouse model. Diurnal proliferation of remnant b-cells was assessed by injecting bromodeoxyuridine (BrdU) before sacrificing in several time-points across 24 h. The role of functional islet oscillators in b-cell regeneration was studies in clock deficient mice. Our data suggest that both absolute levels and diurnal profiles of blood glucose, insulin and glucagon are altered following b-cell ablation. Temporal RNAseq analyses suggest massive changes in b-cell temporal transcriptome landscape, with modest alterations in a-cell transcriptional regulation. Complimentary differential RNAseq analysis of b-cells between intact islets and after ablation showed altered expression of genes involved in hormone secretory pathway, and activation of transcripts responsible for cell proliferation. In line with the circadian rhythmicity of cell-cycle related transcripts, the incorporation of BrdU by residual b-cells exhibited circadian rhythmicity. Moreover, we show that also in young pups, b-cell proliferation follows diurnal pattern. Importantly, the arrhythmic Bmal1 KO mice failed to regenerate b-cells after ablation. To conclude, loss of b-cell mass in rtTA/TET-DTA mice affects gene transcription in residual b-cells, but not in a-cells, while the temporal transcriptome profile is perturbed in both cell types, paralleled with impaired daily secretion of insulin and glucagon. Noteworthy, the key component of molecular clock Bmal1 is required for proper regeneration and functional recovery after massive b-cell ablation. Disclosure V. Petrenko: None. M. Stolovich-Rain: None. B. Vandereycken: None. S. Chera: None. Y. Dor: None. C. Dibner: None. Funding Swiss National Science Foundation (31003A_166700/1, 310030_184708/1)

Type 1 diabetes (T1D) is an autoimmune disease where pancreatic β-cells are destroyed by islet-infiltrating T cells. Although a role for β-cell defects has been suspected, β-cell abnormalities are difficult to demonstrate. We show a β-cell DNA damage response (DDR), presented by activation of the 53BP1 protein and accumulation of p53, in biopsy and autopsy material from patients with recently diagnosed T1D as well as a rat model of human T1D. The β-cell DDR is more frequent in islets infiltrated by CD45+ immune cells, suggesting a link to islet inflammation. The β-cell toxin streptozotocin (STZ) elicits DDR in islets, both in vivo and ex vivo, and causes elevation of the proinflammatory molecules IL-1β and Cxcl10. β-Cell–specific inactivation of the master DNA repair gene ataxia telangiectasia mutated (ATM) in STZ-treated mice decreases the expression of proinflammatory cytokines in islets and attenuates the development of hyperglycemia. Together, these data suggest that β-cell DDR is an early event in T1D, possibly contributing to autoimmunity.

The molecular program underlying infrequent replication of pancreatic β-cells remains largely inaccessible. Using transgenic mice expressing green fluorescent protein in cycling cells, we sorted live, replicating β-cells and determined their transcriptome. Replicating β-cells upregulate hundreds of proliferation-related genes, along with many novel putative cell cycle components. Strikingly, genes involved in β-cell functions, namely, glucose sensing and insulin secretion, were repressed. Further studies using single-molecule RNA in situ hybridization revealed that in fact, replicating β-cells double the amount of RNA for most genes, but this upregulation excludes genes involved in β-cell function. These data suggest that the quiescence-proliferation transition involves global amplification of gene expression, except for a subset of tissue-specific genes, which are “left behind” and whose relative mRNA amount decreases. Our work provides a unique resource for the study of replicating β-cells in vivo.

Understanding and manipulating pancreatic beta-cell proliferation is a major challenge for pancreas biology and diabetes therapy. Recent studies have raised the possibility that human beta-cells can undergo dedifferentiation and give rise to highly proliferative mesenchymal cells, which retain the potential to redifferentiate into beta-cells. To directly test whether cultured beta-cells dedifferentiate, we applied genetic lineage tracing in mice. Differentiated beta-cells were heritably labeled using the Cre-lox system, and their fate in culture was followed. We provide evidence that mouse beta-cells can undergo dedifferentiation in vitro into an insulin-, pdx1-, and glut2-negative state. However, dedifferentiated beta-cells only rarely proliferate under standard culture conditions and are eventually eliminated from cultures. Thus, the predominant mesenchymal cells seen in cultures of mouse islets are not of a beta-cell origin.

Salpeter SJ, Khalaileh A, Weinberg-Corem N, Ziv O, Glaser B, Dor Y. Systemic regulation of the age-related decline of …