scholarly journals EndoC-βH1 multi-genomic profiling defines gene regulatory programs governing human pancreatic β cell identity and function

2018 ◽  
Author(s):  
Nathan Lawlor ◽  
Eladio J. Márquez ◽  
Peter Orchard ◽  
Narisu Narisu ◽  
Muhammad Saad Shamim ◽  
...  
Keyword(s):  
Beta Cell ◽  
Web Application ◽  
Regulatory Networks ◽  
Cell Function ◽  
Cell Model ◽  
Regulatory Element ◽  
Human Islets ◽  
Chromatin Interaction ◽  
Cell Identity ◽  
Molecular Landscape

SUMMARYEndoC-βH1 is emerging as a critical human beta cell model to study the genetic and environmental etiologies of beta cell function, especially in the context of diabetes. Comprehensive knowledge of its molecular landscape is lacking yet required to fully take advantage of this model. Here, we report extensive chromosomal (spectral karyotyping), genetic (genotyping), epigenetic (ChIP-seq, ATAC-seq), chromatin interaction (Hi-C, Pol2 ChIA-PET), and transcriptomic (RNA-seq, miRNA-seq) maps of this cell model. Integrated analyses of these maps define known (e.g.,PDX1, ISL1) and putative (e.g.,PCSK1, mir-375) beta cell-specific chromatin interactions and transcriptionalcis-regulatory networks, and identify allelic effects oncis-regulatory element use and expression.Importantly, comparative analyses with maps generated in primary human islets/beta cells indicate substantial preservation of chromatin looping, but also highlight chromosomal heterogeneity and fetal genomic signatures in EndoC-βH1. Together, these maps, and an interactive web application we have created for their exploration, provide important tools for the broad community in the design and success of experiments to probe and manipulate the genetic programs governing beta cell identity and (dys)function in diabetes.

Persistent expression of HNF6 in islet endocrine cells causes disrupted islet architecture and loss of beta cell function

Development ◽  
2000 ◽  
Vol 127 (13) ◽  
pp. 2883-2895 ◽  
Author(s):  
M. Gannon ◽  
M.K. Ray ◽  
K. Van Zee ◽  
F. Rausa ◽  
R.H. Costa ◽  
...  
Keyword(s):  
Beta Cell ◽  
Beta Cells ◽  
Endocrine Cells ◽  
Spatial Organization ◽  
Glucose Transporter ◽  
Cell Function ◽  
Regulatory Element ◽  
Cell Physiology ◽  
Pancreatic Ducts ◽  
And Function

We used transgenesis to explore the requirement for downregulation of hepatocyte nuclear factor 6 (HNF6) expression in the assembly, differentiation, and function of pancreatic islets. In vivo, HNF6 expression becomes downregulated in pancreatic endocrine cells at 18. 5 days post coitum (d.p.c.), when definitive islets first begin to organize. We used an islet-specific regulatory element (pdx1(PB)) from pancreatic/duodenal homeobox (pdx1) gene to maintain HNF6 expression in endocrine cells beyond 18.5 d.p.c. Transgenic animals were diabetic. HNF6-overexpressing islets were hyperplastic and remained very close to the pancreatic ducts. Strikingly, alpha, delta, and PP cells were increased in number and abnormally intermingled with islet beta cells. Although several mature beta cell markers were expressed in beta cells of transgenic islets, the glucose transporter GLUT2 was absent or severely reduced. As glucose uptake/metabolism is essential for insulin secretion, decreased GLUT2 may contribute to the etiology of diabetes in pdx1(PB)-HNF6 transgenics. Concordantly, blood insulin was not raised by glucose challenge, suggesting profound beta cell dysfunction. Thus, we have shown that HNF6 downregulation during islet ontogeny is critical to normal pancreas formation and function: continued expression impairs the clustering of endocrine cells and their separation from the ductal epithelium, disrupts the spatial organization of endocrine cell types within the islet, and severely compromises beta cell physiology, leading to overt diabetes.

scholarly journals Reprogramming of pancreatic exocrine cells towards a beta (β) cell character using Pdx1, Ngn3 and MafA

2012 ◽  
Vol 442 (3) ◽  
pp. 539-550 ◽  
Author(s):  
Ersin Akinci ◽  
Anannya Banga ◽  
Lucas V. Greder ◽  
James R. Dutton ◽  
Jonathan M. W. Slack
Keyword(s):  
Beta Cell ◽  
Cell Function ◽  
Cell Model ◽  
Severe Combined Immunodeficiency ◽  
Rapid Change ◽  
Cell Phenotype ◽  
Gene Promoters ◽  
Exocrine Cells ◽  
Pancreatic Exocrine

Pdx1 (pancreatic and duodenal homeobox 1), Ngn3 (neurogenin 3) and MafA (v-maf musculoaponeurotic fibrosarcoma oncogene family, protein A) have been reported to bring about the transdifferentiation of pancreatic exocrine cells to beta (β) cells in vivo. We have investigated the mechanism of this process using a standard in vitro model of pancreatic exocrine cells, the rat AR42j-B13 cell line. We constructed a new adenoviral vector encoding all three genes, called Ad-PNM (adenoviral Pdx1, Ngn3, MafA construct). When introduced into AR42j-B13 cells, Ad-PNM caused a rapid change to a flattened morphology and a cessation of cell division. The expression of exocrine markers is suppressed. Both insulin genes are up-regulated as well as a number of transcription factors normally characteristic of beta cells. At the chromatin level, histone tail modifications of the Pdx1, Ins1 (insulin 1) and Ins2 (insulin 2) gene promoters are shifted in a direction associated with gene activity, and the level of DNA CpG methylation is reduced at the Ins1 promoter. The transformed cells secrete insulin and are capable of relieving diabetes in streptozotocin-treated NOD-SCID (non-obese diabetic severe combined immunodeficiency) mice. However the transformation is not complete. The cells lack expression of several genes important for beta cell function and they do not show glucose-sensitive insulin secretion. We conclude that, for this exocrine cell model, although the transformation is dramatic, the reprogramming is not complete and lacks critical aspects of the beta cell phenotype.

scholarly journals Unbiased profiling of the human proinsulin biosynthetic interaction network reveals a role for peroxiredoxin 4 in proinsulin folding

2020 ◽  
Author(s):  
Ada Admin ◽  
Duc T. Tran ◽  
Anita Pottekat ◽  
Saiful A. Mir ◽  
Salvatore Loguercio ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Beta Cell ◽  
Secretory Pathway ◽  
Cell Function ◽  
Affinity Purification ◽  
Interaction Network ◽  
Human Islets ◽  
Cell Protein ◽  
Insulin Biosynthesis ◽  
Human Proinsulin

The beta cell protein synthetic machinery is dedicated to the production of mature insulin, which requires the proper folding and trafficking of its precursor, proinsulin. The complete network of proteins that mediate proinsulin folding and advancement through the secretory pathway, however, remains poorly defined. Here we used affinity purification and mass spectrometry to identify for the first time, the proinsulin biosynthetic interaction network in human islets. Stringent analysis established a central node of proinsulin interactions with ER folding factors, including chaperones and oxidoreductases, that is remarkably conserved in both sexes and across three ethnicities. The ER-localized peroxiredoxin PRDX4 was identified as a prominent proinsulin interacting protein. In beta cells, gene silencing of PRDX4 rendered proinsulin susceptible to misfolding, particularly in response to oxidative stress, while exogenous PRDX4 improved proinsulin folding. Moreover, proinsulin misfolding induced by oxidative stress or high glucose was accompanied by sulfonylation of PRDX4, a modification known to inactivate peroxiredoxins. Notably, islets from patients with Type II diabetes (T2D) exhibited significantly higher levels of sulfonylated PRDX4 than islets from healthy individuals. In conclusion, we have generated the first reference map of the human proinsulin interactome to identify critical factors controlling insulin biosynthesis, beta cell function, and T2D.

scholarly journals Type 2 Diabetes Risk Alleles Reveal a Role for Peptidylglycine Alpha-amidating Monooxygenase in Beta Cell Function

2017 ◽  
Author(s):  
Anne Raimondo ◽  
Soren K. Thomsen ◽  
Benoit Hastoy ◽  
Mahesh M. Umapathysivam ◽  
Xiao-Qing Dai ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Beta Cell ◽  
Beta Cell Function ◽  
Molecular Mechanisms ◽  
Cell Function ◽  
Cell Model ◽  
Catalytic Function ◽  
Risk Alleles ◽  
Human Beta Cells

ABSTRACTMolecular mechanisms underpinning the genetic risk for type 2 diabetes (T2D) remain poorly understood, hindering translation into new therapies. Recently, genome-wide studies identified two coding variants in Peptidylglycine Alpha-amidating Monooxygenase (PAM) associated with T2D risk and measures of beta cell dysfunction. Here, we demonstrate that both risk alleles impact negatively on overall PAM activity, but via distinct effects on expression and catalytic function. In a human beta cell model, PAM silencing caused decreased insulin content and altered dynamics of granule exocytosis. Analysis of primary human beta cells from cadaveric donors confirmed an effect on exocytosis in carriers of the p.D563G T2D-risk allele. Finally, we show that the granular packaging protein Chromogranin A is a PAM substrate and a strong candidate for mediating downstream effects on insulin secretion. Taken together, our results establish a role for PAM in beta cell function, and uncover a novel mechanism for T2D-associated PAM alleles.

scholarly journals OR14-01 FSTL3 Neutralizing Antibodies Restore Function to Diabetic Mouse and Human Islets: A New Approach for Treating Diabetes

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Alan Schneyer ◽  
Melissa L Brown ◽  
Nolan Meyer ◽  
Alexa Lopez ◽  
Alden Richter
Keyword(s):  
Beta Cell ◽  
Beta Cells ◽  
High Glucose ◽  
Beta Cell Function ◽  
Neutralizing Antibodies ◽  
Cell Function ◽  
Human Islets ◽  
Extracellular Proteins ◽  
Chow Diet ◽  
Human Diabetes

Abstract Activin, GDF11 and myostatin are structurally related members of the TGFbeta superfamily of growth factors with many biological roles in animal models and humans. Their actions are neutralized by extracellular proteins such as follistatin and follistatin like-3 (FSTL3). We have previously demonstrated that genetic inactivation of Fstl3 results in enlarged pancreatic islets containing increased numbers of beta cells that produce more insulin in response to glucose compared to wild type litter mates. We further discovered that at least some of these new beta cells arise via transdifferentiation from alpha cells. We also demonstrated that functional human islets from normal donors produce very high levels of activin. In contrast, activin biosynthesis is vastly reduced and FSTL3 synthesis is significantly increased in human islets from diabetic donors suggesting that activin is critical for normal insulin production. This was substantiated by direct treatment of human diabetic islets with activin which restored their response to glucose. These observations support the hypothesis that an FSTL3 neutralizing antibody would constitute a novel therapeutic approach to curing diabetes through restoring beta cell function as well as accelerating generation of new beta cells through transdifferentiation. To test this hypothesis, we produced a mouse monoclonal antibody that neutralized hFSTL3 (FP-101), thereby releasing bioactive activin, GDF11, and myostatin. We have now tested this antibody for biological activity in vitro on mouse and human islets. We used islets from high fat diet (HFD) treated mice to model diabetes-inducing effects of obesity as well as 24-hour incubation in hyperglycemic (33 mM glucose) medium to create human islets that lose responsiveness to high glucose as a model for human diabetes. In mouse islets we found that stimulation of normal (chow diet) islets by high glucose produced a stimulation index (SI) of 3.5 that was reduced to 2 in HFD islets. Treatment with activin, FP-101, or a commercial polyclonal antibody to mFSTL3 all increased response of HFD islets to elevated glucose and partially restored SI to normal levels. In human islets, hyperglycemia eliminated the normal (2.5 SI) response to high glucose while activin or FP-101 treatments dose-responsively restored this response. These results demonstrate that anti-FSTL3 therapy can restore function to compromised beta cells from mouse and human diabetes models. The observation that activin has the same action as anti-FSTL3 antibody indicates that FP-101 works through enhancing the activin signaling pathway. Finally, these results demonstrate that the FSTL3-activin pathway is an important regulator of beta cell function in humans as well as mice, supporting further development of this therapy as a diabetes treatment.

scholarly journals The diet-derived short chain fatty acid propionate improves beta-cell function in humans and stimulates insulin secretion from human islets in vitro

2016 ◽  
Vol 19 (2) ◽  
pp. 257-265 ◽  
Author(s):  
Attilio Pingitore ◽  
Edward S. Chambers ◽  
Thomas Hill ◽  
Inmaculada Ruz Maldonado ◽  
Bo Liu ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Fatty Acid ◽  
Beta Cell ◽  
Beta Cell Function ◽  
Short Chain Fatty Acid ◽  
Cell Function ◽  
Human Islets ◽  
Chain Fatty Acid ◽  
Short Chain

scholarly journals IL-1 produced and released endogenously within human islets inhibits beta cell function.

1998 ◽  
Vol 102 (3) ◽  
pp. 516-526 ◽  
Author(s):  
M Arnush ◽  
M R Heitmeier ◽  
A L Scarim ◽  
M H Marino ◽  
P T Manning ◽  
...  
Keyword(s):  
Beta Cell ◽  
Beta Cell Function ◽  
Cell Function ◽  
Human Islets
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