scholarly journals Exocrine pancreas proteases regulate β-cell proliferation in zebrafish ciliopathy models and in murine systems

Biology Open ◽  
2021 ◽  
Vol 10 (6) ◽  
Author(s):  
Timothy L. Hostelley ◽  
Jessica E. Nesmith ◽  
Emily Larkin ◽  
Amanda Jones ◽  
Daniel Boyes ◽  
...  
Keyword(s):  
Exocrine Pancreas ◽  
Ex Vivo ◽  
Cell Mass ◽  
Transgenic Zebrafish ◽  
Genetic Syndromes ◽  
Β Cells ◽  
Β Cell ◽  
Cell Numbers ◽  
Pancreatic Proteases

ABSTRACT Pancreatic β-cells are a critical cell type in the pathology of diabetes. Models of genetic syndromes featuring diabetes can provide novel mechanistic insights into regulation of β-cells in the context of disease. We previously examined β-cell mass in models of two ciliopathies, Alström Syndrome (AS) and Bardet-Biedl Syndrome (BBS), which are similar in the presence of metabolic phenotypes, including obesity, but exhibit strikingly different rates of diabetes. Zebrafish models of these disorders show deficient β-cells with diabetes in AS models and an increased β-cells absent diabetes in BBS models, indicating β-cell generation or maintenance that correlates with disease prevalence. Using transcriptome analyses, differential expression of several exocrine pancreas proteases with directionality that was consistent with β-cell numbers were identified. Based on these lines of evidence, we hypothesized that pancreatic proteases directly impact β-cells. In the present study, we examined this possibility and found that pancreatic protease genes contribute to proper maintenance of normal β-cell numbers, proliferation in larval zebrafish, and regulation of AS and BBS β-cell phenotypes. Our data suggest that these proteins can be taken up directly by cultured β-cells and ex vivo murine islets, inducing proliferation in both. Endogenous uptake of pancreatic proteases by β-cells was confirmed in vivo using transgenic zebrafish and in intact murine pancreata. Taken together, these findings support a novel proliferative signaling role for exocrine pancreas proteases through interaction with endocrine β-cells.

scholarly journals NF-κB activity during pancreas development regulates adult β-cell mass by modulating neonatal β-cell proliferation and apoptosis

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Dror Sever ◽  
Anat Hershko-Moshe ◽  
Rohit Srivastava ◽  
Roy Eldor ◽  
Daniel Hibsher ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Developmental Stages ◽  
Cell Mass ◽  
Diabetes Incidence ◽  
Β Cells ◽  
Β Cell ◽  
Cell Dysfunction ◽  
Regulatory Circuit ◽  
Proliferation And Apoptosis

AbstractNF-κB is a well-characterized transcription factor, widely known for its roles in inflammation and immune responses, as well as in control of cell division and apoptosis. However, its function in β-cells is still being debated, as it appears to depend on the timing and kinetics of its activation. To elucidate the temporal role of NF-κB in vivo, we have generated two transgenic mouse models, the ToIβ and NOD/ToIβ mice, in which NF-κB activation is specifically and conditionally inhibited in β-cells. In this study, we present a novel function of the canonical NF-κB pathway during murine islet β-cell development. Interestingly, inhibiting the NF-κB pathway in β-cells during embryogenesis, but not after birth, in both ToIβ and NOD/ToIβ mice, increased β-cell turnover, ultimately resulting in a reduced β-cell mass. On the NOD background, this was associated with a marked increase in insulitis and diabetes incidence. While a robust nuclear immunoreactivity of the NF-κB p65-subunit was found in neonatal β-cells, significant activation was not detected in β-cells of either adult NOD/ToIβ mice or in the pancreata of recently diagnosed adult T1D patients. Moreover, in NOD/ToIβ mice, inhibiting NF-κB post-weaning had no effect on the development of diabetes or β-cell dysfunction. In conclusion, our data point to NF-κB as an important component of the physiological regulatory circuit that controls the balance of β-cell proliferation and apoptosis in the early developmental stages of insulin-producing cells, thus modulating β-cell mass and the development of diabetes in the mouse model of T1D.

scholarly journals β-Cell Generation: Can Rodent Studies Be Translated to Humans?

2011 ◽  
Vol 2011 ◽  
pp. 1-15 ◽  
Author(s):  
Françoise Carlotti ◽  
Arnaud Zaldumbide ◽  
Johanne H. Ellenbroek ◽  
H. Siebe Spijker ◽  
Rob C. Hoeben ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Cell Mass ◽  
Cell Types ◽  
Organ Donors ◽  
Cell Replacement Therapy ◽  
Β Cell ◽  
Cell Replacement ◽  
Therapeutic Alternatives

β-cell replacement by allogeneic islet transplantation is a promising approach for patients with type 1 diabetes, but the shortage of organ donors requires new sources ofβcells. Islet regenerationin vivoand generation ofβ-cellsex vivofollowed by transplantation represent attractive therapeutic alternatives to restore theβ-cell mass. In this paper, we discuss different postnatal cell types that have been envisaged as potential sources for futureβ-cell replacement therapy. The ultimate goal being translation to the clinic, a particular attention is given to the discrepancies between findings from studies performed in rodents (bothex vivoon primary cells andin vivoon animal models), when compared with clinical data and studies performed on human cells.

scholarly journals MafB Is Critical for Glucagon Production and Secretion in Mouse Pancreatic α Cells In Vivo

2018 ◽  
Vol 38 (8) ◽  
Author(s):  
Megumi C. Katoh ◽  
Yunshin Jung ◽  
Chioma M. Ugboma ◽  
Miki Shimbo ◽  
Akihiro Kuno ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Cell Population ◽  
Pancreatic Polypeptide ◽  
Transporter Gene ◽  
Β Cells ◽  
Β Cell ◽  
Cell Numbers ◽  
Gene Expression Analyses

ABSTRACT The MafB transcription factor is expressed in pancreatic α and β cells during development but becomes exclusive to α cells in adult rodents. Mafb -null ( Mafb −/− ) mice were reported to have reduced α- and β-cell numbers throughout embryonic development. To further analyze the postnatal function of MafB in the pancreas, we generated endocrine cell-specific ( Mafb Δ Endo ) and tamoxifen-dependent ( Mafb Δ TAM ) Mafb knockout mice. Mafb Δ Endo mice exhibited reduced populations of insulin-positive (insulin + ) and glucagon + cells at postnatal day 0, but the insulin + cell population recovered by 8 weeks of age. In contrast, the Arx + glucagon + cell fraction and glucagon expression remained decreased even in adulthood. Mafb Δ TAM mice, with Mafb deleted after pancreas maturation, also demonstrated diminished glucagon + cells and glucagon content without affecting β cells. A decreased Arx + glucagon + cell population in Mafb Δ Endo mice was compensated for by an increased Arx + pancreatic polypeptide + cell population. Furthermore, gene expression analyses from both Mafb Δ Endo and Mafb Δ TAM islets revealed that MafB is a key regulator of glucagon expression in α cells. Finally, both mutants failed to respond to arginine, likely due to impaired arginine transporter gene expression and glucagon production ability. Taken together, our findings reveal that MafB is critical for the functional maintenance of mouse α cells in vivo , including glucagon production and secretion, as well as in development.

scholarly journals Dual-Reporter β-Cell-Specific Male Transgenic Rats for the Analysis of β-Cell Functional Mass and Enrichment by Flow Cytometry

Endocrinology ◽  
2015 ◽  
Vol 157 (3) ◽  
pp. 1299-1306 ◽  
Author(s):  
Julien Ghislain ◽  
Ghislaine Fontés ◽  
Caroline Tremblay ◽  
Melkam A. Kebede ◽  
Vincent Poitout
Keyword(s):  
Insulin Secretion ◽  
Fluorescent Protein ◽  
Ex Vivo ◽  
Yellow Fluorescent Protein ◽  
Cell Mass ◽  
Renilla Luciferase ◽  
Diabetes Research ◽  
Β Cells ◽  
Transgenic Rats ◽  
Β Cell

Abstract Mouse β-cell-specific reporter lines have played a key role in diabetes research. Although the rat provides several advantages, its use has lagged behind the mouse due to the relative paucity of genetic models. In this report we describe the generation and characterization of transgenic rats expressing a Renilla luciferase (RLuc)-enhanced yellow fluorescent protein (YFP) fusion under control of a 9-kb genomic fragment from the rat ins2 gene (RIP7-RLuc-YFP). Analysis of RLuc luminescence and YFP fluorescence revealed that reporter expression is restricted to β-cells in the adult rat. Physiological characteristics including body weight, fat and lean mass, fasting and fed glucose levels, glucose and insulin tolerance, and β-cell mass were similar between two RIP7-RLuc-YFP lines and wild-type littermates. Glucose-induced insulin secretion in isolated islets was indistinguishable from controls in one of the lines, whereas surprisingly, insulin secretion was defective in the second line. Consequently, subsequent studies were limited to the former line. We asked whether transgene activity was responsive to glucose as shown previously for the ins2 gene. Exposing islets ex vivo to high glucose (16.7 mM) or in vivo infusion of glucose for 24 hours increased luciferase activity in islets, whereas the fraction of YFP-positive β-cells after glucose infusion was unchanged. Finally, we showed that fluorescence-activated cell sorting of YFP-positive islet cells can be used to enrich for β-cells. Overall, this transgenic line will enable for the first time the application of both fluorescence and bioluminescence/luminescence-based approaches for the study of rat β-cells.

scholarly journals Loss of Foxd3 Results in Decreased β-Cell Proliferation and Glucose Intolerance During Pregnancy

Endocrinology ◽  
2011 ◽  
Vol 152 (12) ◽  
pp. 4589-4600 ◽  
Author(s):  
Jennifer L. Plank ◽  
Audrey Y. Frist ◽  
Alison W. LeGrone ◽  
Mark A. Magnuson ◽  
Patricia A. Labosky
Keyword(s):  
Cell Proliferation ◽  
Cell Mass ◽  
Diabetic Patients ◽  
Β Cells ◽  
Β Cell ◽  
Forkhead Box ◽  
Physiological Demands ◽  
Mass Expansion ◽  
Specific Deletion

A complete molecular understanding of β-cell mass expansion will be useful for the improvement of therapies to treat diabetic patients. During normal periods of metabolic challenges, such as pregnancy, β-cells proliferate, or self-renew, to meet the new physiological demands. The transcription factor Forkhead box D3 (Foxd3) is required for maintenance and self-renewal of several diverse progenitor cell lineages, and Foxd3 is expressed in the pancreatic primordium beginning at 10.5 d postcoitum, becoming localized predominantly to β-cells after birth. Here, we show that mice carrying a pancreas-specific deletion of Foxd3 have impaired glucose tolerance, decreased β-cell mass, decreased β-cell proliferation, and decreased β-cell size during pregnancy. In addition, several genes known to regulate proliferation, Foxm1, Skp2, Ezh2, Akt2, and Cdkn1a, are misregulated in islets isolated from these Foxd3 mutant mice. Together, these data place Foxd3 upstream of several pathways critical for β-cell mass expansion in vivo.

scholarly journals Beta-cell specific insulin resistance promotes glucose-stimulated insulin hypersecretion

2020 ◽  
Author(s):  
Søs Skovsø ◽  
Evgeniy Panzhinskiy ◽  
Jelena Kolic ◽  
Derek A. Dionne ◽  
Xiao-Qing Dai ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Glucose Tolerance ◽  
Cell Mass ◽  
Whole Body ◽  
Calcium Oscillation ◽  
Male Mice ◽  
Β Cells ◽  
Β Cell ◽  
Specific Insulin

AbstractInsulin receptor (Insr) protein can be found at higher levels in pancreatic β-cells than in most other cell types, but the consequences of β-cell insulin resistance remain enigmatic. Ins1cre allele was used to delete Insr specifically in β-cells of both female and male mice which were compared to Ins1cre-containing littermate controls at multiple ages and on multiple diets. RNA-seq of recombined β-cells revealed significant differences in multiple pathways previously implicated in insulin secretion and cellular fate, including rewired Ras and NFκB signaling. Male, but not female, βInsrKO mice had reduced oxygen consumption rate, while action potential and calcium oscillation frequencies were increased in Insr knockout β-cells from female, but not male mice. Female βInsrKO and βInsrHET mice exhibited elevated insulin release in perifusion experiments, during hyperglycemic clamps, and following i.p. glucose challenge. Deletion of Insr did not reduce β-cell mass up to 9 months of age, nor did it impair hyperglycemia-induced proliferation. Based on our data, we adapted a mathematical model to include β-cell insulin resistance, which predicted that β-cell Insr knockout would improve glucose tolerance depending on the degree of whole-body insulin resistance. Indeed, glucose tolerance was significantly improved in female βInsrKO and βInsrHET mice when compared to controls at 9, 21 and 39 weeks. We did not observe improved glucose tolerance in adult male mice or in high fat diet-fed mice, corroborating the prediction that global insulin resistance obscures the effects of β-cell specific insulin resistance. We further validated our in vivo findings using the Ins1-CreERT transgenic line and found improved glucose tolerance 4 weeks after tamoxifen-mediated Insr deletion. Collectively, our data show that loss of β-cell Insr alone is sufficient to drive glucose-induced hyperinsulinemia, thereby improving glucose homeostasis in otherwise insulin sensitive dietary and age contexts.

scholarly journals Role of the G-Protein Coupled Receptor 3-Salt Inducible Kinase 2 Pathway in Human β Cell Proliferation

2021 ◽  
Author(s):  
Caterina Iorio ◽  
Jillian L Rourke ◽  
Lisa Wells ◽  
Jun-Ichi Sakamaki ◽  
Emily Moon ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
G Protein ◽  
Cell Mass ◽  
Standard Of Care ◽  
G Protein Coupled Receptor ◽  
Β Cells ◽  
Β Cell ◽  
Cell Therapies ◽  
G Protein Coupled

Loss of pancreatic β cells is the hallmark of type 1 diabetes (T1D), for which provision of insulin is the standard of care. While regenerative and stem cell therapies hold the promise of generating single-source or host-matched tissue to obviate immune-mediated complications, these will still require surgical intervention and immunosuppression. Thus, methods that harness the innate capacity of β cells to proliferate to increase β cell mass in vivo are considered vital for future T1D treatment. However, early in life β cells enter what appears to be a permanent state of quiescence, directed by an evolutionarily selected genetic program that establishes a β cell mass setpoint to guard against development of fatal endocrine tumours. Here we report the development of a high-throughput RNAi screening approach to identify upstream pathways that regulate adult human β cell quiescence and demonstrate in a screen of the GPCRome that silencing G-protein coupled receptor 3 (GPR3) leads to human pancreatic β cell proliferation. Loss of GPR3 leads to activation of Salt Inducible Kinase 2 (SIK2), which is necessary and sufficient to drive cell cycle entry, increase β cell mass, and enhance insulin secretion in mice. Taken together, targeting the GPR3-SIK2 pathway represents a novel avenue to stimulate the regeneration of β cells.

GLP-1(28–36) improves β-cell mass and glucose disposal in streptozotocin-induced diabetic mice and activates cAMP/PKA/β-catenin signaling in β-cells in vitro

2013 ◽  
Vol 304 (12) ◽  
pp. E1263-E1272 ◽  
Author(s):  
Weijuan Shao ◽  
Zhaoxia Wang ◽  
Wilfred Ip ◽  
Yu-Ting Chiang ◽  
Xiaoquan Xiong ◽  
...  
Keyword(s):  
Cyclin D1 ◽  
Cell Line ◽  
Cell Mass ◽  
Glucose Disposal ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells ◽  
In Vitro Investigation

Recent studies have demonstrated that the COOH-terminal fragment of the incretin hormone glucagon-like peptide-1 (GLP-1), a nonapeptide GLP-1(28–36)amide, attenuates diabetes and hepatic steatosis in diet-induced obese mice. However, the effect of this nonapeptide in pancreatic β-cells remains largely unknown. Here, we show that in a streptozotocin-induced mouse diabetes model, GLP-1(28–36)amide improved glucose disposal and increased pancreatic β-cell mass and β-cell proliferation. An in vitro investigation revealed that GLP-1(28–36)amide stimulates β-catenin (β-cat) Ser675 phosphorylation in both the clonal INS-1 cell line and rat primary pancreatic islet cells. In INS-1 cells, the stimulation was accompanied by increased nuclear β-cat content. GLP-1(28–36)amide was also shown to increase cellular cAMP levels, PKA enzymatic activity, and cAMP response element-binding protein (CREB) and cyclic AMP-dependent transcription factor-1 (ATF-1) phosphorylation. Furthermore, GLP-1(28–36)amide treatment enhanced islet insulin secretion and increased the growth of INS-1 cells, which was associated with increased cyclin D1 expression. Finally, PKA inhibition attenuated the effect of GLP-1(28–36)amide on β-cat Ser675 phosphorylation and cyclin D1 expression in the INS-1 cell line. We have thus revealed the beneficial effect of GLP-1(28–36)amide in pancreatic β-cells in vitro and in vivo. Our observations suggest that GLP-1(28–36)amide may exert its effect through the PKA/β-catenin signaling pathway.

scholarly journals The Many Lives of Myc in the Pancreatic β-Cell

2020 ◽  
pp. jbc.REV120.011149
Author(s):  
Carolina Rosselot ◽  
Sharon Baumel-Alterzon ◽  
Yansui Li ◽  
Gabriel Brill ◽  
Luca Lambertini ◽  
...  
Keyword(s):  
Cell Death ◽  
Transcription Factor ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Diabetic Patients ◽  
Cell Regeneration ◽  
Diabetes Research ◽  
Β Cells ◽  
Β Cell

Diabetes results from insufficient numbers of functional pancreatic β-cells. Thus, increasing the number of available functional β-cells ex vivo for transplantation, or regenerating them in situ in diabetic patients, is a major focus of diabetes research. The transcription factor, Myc, discovered decades ago, lies at the nexus of most, if not all, known proliferative pathways. Based on this, many studies in the 1990’s and early 2000’s explored the potential of harnessing Myc expression to expand β-cells for diabetes treatment. Nearly all these studies in β-cells used pathophysiological or supraphysiological levels of Myc and reported enhanced β-cell death, de-differentiation or the formation of insulinomas if co-overexpressed with Bcl-xL, an inhibitor of apoptosis. This obviously reduced the enthusiasm for Myc as a therapeutic target for β-cell regeneration. However, recent studies indicate that “gentle” induction of Myc expression enhances β-cell replication without induction of cell death or loss of insulin secretion, suggesting that appropriate levels of Myc could have therapeutic potential for β-cell regeneration. Furthermore, although it has been known for decades that Myc is induced by glucose in β-cells very little is known about how this essential anabolic transcription factor perceives and responds to nutrients and increased insulin demand in vivo. Here we summarize the previous and recent knowledge of Myc in the β-cell, its potential for β-cell regeneration and its physiological importance for neonatal and adaptive β-cell expansion.

scholarly journals A fluorescent timer reporter enables sorting of insulin secretory granules by age

2020 ◽  
Vol 295 (27) ◽  
pp. 8901-8911 ◽  
Author(s):  
Belinda Yau ◽  
Lori Hays ◽  
Cassandra Liang ◽  
D. Ross Laybutt ◽  
Helen E. Thomas ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Cell Function ◽  
Islet Cells ◽  
Ex Vivo ◽  
Secretory Granules ◽  
Β Cells ◽  
Β Cell ◽  
Short Period ◽  
Insulin Granule

Within the pancreatic β-cells, insulin secretory granules (SGs) exist in functionally distinct pools, displaying variations in motility as well as docking and fusion capability. Current therapies that increase insulin secretion do not consider the existence of these distinct SG pools. Accordingly, these approaches are effective only for a short period, with a worsening of glycemia associated with continued decline in β-cell function. Insulin granule age is underappreciated as a determinant for why an insulin granule is selected for secretion and may explain why newly synthesized insulin is preferentially secreted from β-cells. Here, using a novel fluorescent timer protein, we aimed to investigate the preferential secretion model of insulin secretion and identify how granule aging is affected by variation in the β-cell environment, such as hyperglycemia. We demonstrate the use of a fluorescent timer construct, syncollin-dsRedE5TIMER, which changes its fluorescence from green to red over 18 h, in both microscopy and fluorescence-assisted organelle-sorting techniques. We confirm that the SG-targeting construct localizes to insulin granules in β-cells and does not interfere with normal insulin SG behavior. We visualize insulin SG aging behavior in MIN6 and INS1 β-cell lines and in primary C57BL/6J mouse and nondiabetic human islet cells. Finally, we separated young and old insulin SGs, revealing that preferential secretion of younger granules occurs in glucose-stimulated insulin secretion. We also show that SG population age is modulated by the β-cell environment in vivo in the db/db mouse islets and ex vivo in C57BL/6J islets exposed to different glucose environments.

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