期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2020
卷号：117
期号：16
页码：8912-8923
DOI：10.1073/pnas.2001936117
出版社：The National Academy of Sciences of the United States of America
摘要：Pancreatic islets regulate glucose homeostasis through coordinated actions of hormone-secreting cells. What underlies the function of the islet as a unit is the close approximation and communication among heterogeneous cell populations, but the structural mediators of islet cellular cross talk remain incompletely characterized. We generated mice specifically lacking β-cell primary cilia, a cellular organelle that has been implicated in regulating insulin secretion, and found that the β-cell cilia are required for glucose sensing, calcium influx, insulin secretion, and cross regulation of α- and δ-cells. Protein expression profiling in islets confirms perturbation in these cellular processes and reveals additional targets of cilia-dependent signaling. At the organism level, the deletion of β-cell cilia disrupts circulating hormone levels, impairs glucose homeostasis and fuel usage, and leads to the development of diabetes. Together, these findings demonstrate that primary cilia not only orchestrate β-cell–intrinsic activity but also mediate cross talk both within the islet and from islets to other metabolic tissues, thus providing a unique role of cilia in nutrient metabolism and insight into the pathophysiology of diabetes. The pancreatic islet secretes hormones required for metabolic homeostasis. Common to all forms of diabetes are a relative or absolute insulin deficiency and metabolic imbalance associated with β-cell dysfunction ( 1 ). Islet hormone secretion is a dynamic process determined by not only cell-intrinsic properties, e.g., ion channels, but also cell–cell connectivity and communication ( 2 , 3 ). Primary cilia are a unique regulator of islet cells; a single primary cilium protrudes from each cell body and occupies the common luminal space between neighboring islet cells ( 4 , 5 ). These hairlike organs are rich with G protein-coupled receptors (GPCRs) and chemosensory receptors and act as a signaling hub to direct cellular functions. Structurally, IFT88 is a component of the intraflagellar transport (IFT) complex and is required for cilia assembly ( 6 , 7 ). Loss of IFT88 causes the absence of cilia and leads to cystic kidney disease in both mice and humans ( 8 , 9 ). Primary cilia have been shown to regulate insulin secretion ( 10 ), but it is unclear which events during β-cell glucose-stimulated insulin secretion are under cilia control and how this relates to whole-body physiology. A high incidence of obesity and diabetes is found in two human ciliopathies, Bardet–Biedl and Alström syndromes ( 11 , 12 ). The pathophysiology of cilia-related diabetes is incompletely understood and likely encompasses combined effects on feeding behavior, pancreatic development, and glucose handling. Most animal models of ciliopathy-related diabetes to date have been global or whole-pancreas knockouts with mixed phenotypes that cannot be attributed to defects in any specific tissue or cell type ( 10 , 13 , 14 ). Accordingly, there is a lack of mechanistic understanding of cilia-dependent regulation of the endocrine pancreas. To specifically examine the role of cilia in β-cell and islet function, we generated an Ins1-Cre β-cell cilia knockout (βCKO) mouse and studied its phenotype at the cellular, tissue, and organismal level. We find that targeted deletion of β-cell cilia causes not only β-cell secretory failure, as also seen in a recent Pdx1-Cre cilia KO model ( 15 ), but also aberrant α- and δ-cell hormone secretion and altered systemic energy metabolism. Our studies implicate primary cilia as a key regulator of glucose-sensing, cellular synchronicity, and both intra- and intercellular signaling pathways that govern core islet functions, demonstrating that primary cilia are required for islet function as a unit and for the maintenance of energy homeostasis. Results INS1-Cre/IFT88-Flox Mice Lack β-Cell Cilia. To determine the role of primary cilia in β-cell function, we generated βCKO mice by crossing INS1-Cre ( 16 ) with IFT88-Flox mice ( 17 ). The INS1-Cre strain was chosen based on efficient and selective recombination in β-cells and established lack of expression in the central nervous system ( 16 ). A parallel tamoxifen-inducible line, βCKO-ERT2, was generated by crossing IFT88-Flox with INS1-CreERT2 ( 16 ) and was used as a control. Both βCKO and βCKO-ERT2 mice are on the C57BL/6 background, are fertile and carry normal-size litters in Mendelian ratios, and produce pups with no obvious developmental defects at the time of weaning. Successful cilia knockout was confirmed using immunohistochemistry, qPCR, and immunoblotting. To visualize primary cilia morphology and distribution in normal islets, we stained healthy human and mouse islets with three independent cilia markers, including Arl13b, polyglutamylated tubulin, and acetylated α-tubulin ( Fig. 1 A and C ). We also captured an electron micrograph of a human β-cell cilium in ultrastructural detail ( Fig. 1 B ). Compared to wild-type (WT) mice, βCKO mice lack cilia exclusively on islet β-cells ( Fig. 1 C ), with deletion of IFT88 gene expression confirmed by qPCR ( Fig. 1 D ) and by immunoblot of IFT88 ( Fig. 1 E ). Residual IFT88 expression in knockout islets was likely from α-cells and other non–β-cells. As a control, we induced β-cell cilia knockout by treating βCKO-ERT2 mice with tamoxifen and showed that this depleted cilia in adult islet β-cells and induced hyperglycemia and glucose intolerance within days of treatment ( SI Appendix , Fig. S1 ). Primary cilia expression on βCKO-ERT2 islet α-cells and other non–β-cells was unaffected by tamoxifen, confirming β-cell specificity of Cre-ERT2 recombination.
