scholarly journals Insulin and secretagogues differentially regulate fluid-phase pinocytosis in insulin-secreting β-cells

1996 ◽  
Vol 318 (2) ◽  
pp. 623-629 ◽  
Author(s):  
Gang XU ◽  
Jennie HOWLAND ◽  
Paul L ROTHENBERG
Keyword(s):  
T Cells ◽  
Cell Surface ◽  
Insulin Receptor ◽  
Cell Line ◽  
Chinese Hamster Ovary Cells ◽  
Insulin Receptors ◽  
Fluid Phase ◽  
Β Cells ◽  
Β Cell ◽  
Basal Rate

The physiological role of the β-cell insulin receptor is unknown. To evaluate a candidate function, the insulin regulation of fluid-phase pinocytosis was investigated in a clonal insulinoma cell line (βTC6-F7) and, for comparison, also in Chinese hamster ovary cells transfected with the human insulin receptor (CHO-T cells). In CHO-T cells, the net rate of fluid-phase pinocytosis was rapidly increased 3–4-fold over the basal rate by 100 nM insulin, with half-maximal stimulation at 2 nM insulin, as assayed by cellular uptake of horseradish peroxidase from the medium. Wortmannin, an inhibitor of phosphatidylinositol (PI)-3-kinase, blocked insulin-stimulated pinocytosis with an IC50 of 7.5 nM without affecting the basal rate of pinocytosis. In insulin-secreting βTC6-F7 cells, the secretagogues glucose and carbachol (at maximally effective concentrations of 15 mM and 0.5 mM respectively) augmented fluid-phase pinocytosis 1.65-fold over the basal rate. Wortmannin also inhibited secretagogue-stimulated pinocytosis in these β-cells with an IC50 of 7 nM but did not affect the basal rate of pinocytosis measured in the absence of secretagogues. Wortmannin did not influence either basal or secretagogue-induced insulin secretion. Although these βTC6-F7 cells have cell-surface insulin receptors, adding exogenous insulin or insulin-like growth factor 1 did not affect their rate of fluid-phase pinocytosis, either in the absence or presence of secretagogues. From these observations, we conclude that: (1) in both insulin-secreting β-cells and in conventional, insulin-responsive CHO-T cells, a common, wortmannin-sensitive reaction, which probably involves PI-3-kinase, regulates fluid-phase pinocytosis; (2) the insulin-receptor signal transduction pathway is dissociated from the regulation of fluid-phase pinocytosis in the insulin-secreting β-cell line we studied; and (3) the enhancement of fluid-phase pinocytosis associated with secretagogue-induced insulin release in βTC6-F7 cells is not attributable to autocrine activation of β-cell surface insulin receptors.

scholarly journals Glucose Effects on Beta-Cell Growth and Survival Require Activation of Insulin Receptors and Insulin Receptor Substrate 2

2009 ◽  
Vol 29 (11) ◽  
pp. 3219-3228 ◽  
Author(s):  
Anke Assmann ◽  
Kohjiro Ueki ◽  
Jonathon N. Winnay ◽  
Takahashi Kadowaki ◽  
Rohit N. Kulkarni
Keyword(s):  
Insulin Receptor ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Insulin Signaling ◽  
Insulin Receptors ◽  
Insulin Signaling Pathway ◽  
Insulin Receptor Substrate ◽  
Β Cells ◽  
Β Cell ◽  
Growth And Survival

ABSTRACT Insulin and insulin-like growth factor I (IGF-I) are ubiquitous hormones that regulate growth and metabolism of most mammalian cells, including pancreatic β-cells. In addition to being an insulin secretagogue, glucose regulates proliferation and survival of β-cells. However, it is unclear whether the latter effects of glucose occur secondary to autocrine activation of insulin signaling proteins by secreted insulin. To examine this possibility we studied the effects of exogenous glucose or insulin in β-cell lines completely lacking either insulin receptors (βIRKO) or insulin receptor substrate 2 (βIRS2KO). Exogenous addition of either insulin or glucose activated proteins in the insulin signaling pathway in control β-cell lines with the effects of insulin peaking earlier than glucose. Insulin stimulation of βIRKO and βIRS2KO cells led to blunted activation of phosphatidylinositol 3-kinase and Akt kinase, while surprisingly, glucose failed to activate either kinase but phosphorylated extracellular signal-regulated kinase. Control β-cells exhibited low expression of IGF-1 receptors compared to compensatory upregulation in βIRKO cells. The signaling data support the slow growth and reduced DNA and protein synthesis in βIRKO and βIRS2KO cells in response to glucose stimulation. Together, these studies provide compelling evidence that the growth and survival effects of glucose on β-cells require activation of proteins in the insulin signaling pathway.

scholarly journals Nonlinear Analysis for a Type-1 Diabetes Model with Focus on T-Cells and Pancreatic β-Cells Behavior

2020 ◽  
Vol 25 (2) ◽  
pp. 23
Author(s):  
Diana Gamboa ◽  
Carlos E. Vázquez ◽  
Paul J. Campos
Keyword(s):  
T Cells ◽  
Type 1 Diabetes ◽  
Cell Population ◽  
Closed Loop ◽  
Pancreatic Β Cell ◽  
Activated Macrophages ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells

Type-1 diabetes mellitus (T1DM) is an autoimmune disease that has an impact on mortality due to the destruction of insulin-producing pancreatic β -cells in the islets of Langerhans. Over the past few years, the interest in analyzing this type of disease, either in a biological or mathematical sense, has relied on the search for a treatment that guarantees full control of glucose levels. Mathematical models inspired by natural phenomena, are proposed under the prey–predator scheme. T1DM fits in this scheme due to the complicated relationship between pancreatic β -cell population growth and leukocyte population growth via the immune response. In this scenario, β -cells represent the prey, and leukocytes the predator. This paper studies the global dynamics of T1DM reported by Magombedze et al. in 2010. This model describes the interaction of resting macrophages, activated macrophages, antigen cells, autolytic T-cells, and β -cells. Therefore, the localization of compact invariant sets is applied to provide a bounded positive invariant domain in which one can ensure that once the dynamics of the T1DM enter into this domain, they will remain bounded with a maximum and minimum value. Furthermore, we analyzed this model in a closed-loop scenario based on nonlinear control theory, and proposed bases for possible control inputs, complementing the model with them. These entries are based on the existing relationship between cell–cell interaction and the role that they play in the unchaining of a diabetic condition. The closed-loop analysis aims to give a deeper understanding of the impact of autolytic T-cells and the nature of the β -cell population interaction with the innate immune system response. This analysis strengthens the proposal, providing a system free of this illness—that is, a condition wherein the pancreatic β -cell population holds and there are no antigen cells labeled by the activated macrophages.

scholarly journals Differentiation of Sendai Virus-Reprogrammed iPSC into β Cells, Compared with Human Pancreatic Islets and Immortalized β Cell Line

2018 ◽  
Vol 27 (10) ◽  
pp. 1548-1560 ◽  
Author(s):  
Silvia Pellegrini ◽  
Fabio Manenti ◽  
Raniero Chimienti ◽  
Rita Nano ◽  
Linda Ottoboni ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pancreatic Islets ◽  
Cell Line ◽  
Expression Profile ◽  
Sendai Virus ◽  
Secretory Function ◽  
Β Cells ◽  
Β Cell ◽  
Insulin Producing Cells ◽  
Gene And Protein Expression

Background: New sources of insulin-secreting cells are strongly in demand for treatment of diabetes. Induced pluripotent stem cells (iPSCs) have the potential to generate insulin-producing cells (iβ). However, the gene expression profile and secretory function of iβ still need to be validated in comparison with native β cells. Methods: Two clones of human iPSCs, reprogrammed from adult fibroblasts through integration-free Sendai virus, were differentiated into iβ and compared with donor pancreatic islets and EndoC-βH1, an immortalized human β cell line. Results: Both clones of iPSCs differentiated into insulin+ cells with high efficiency (up to 20%). iβ were negative for pluripotency markers (Oct4, Sox2, Ssea4) and positive for Pdx1, Nkx6.1, Chromogranin A, PC1/3, insulin, glucagon and somatostatin. iβ basally secreted C-peptide, glucagon and ghrelin and released insulin in response either to increasing concentration of glucose or a depolarizing stimulus. The comparison revealed that iβ are remarkably similar to donor derived islets in terms of gene and protein expression profile and similar level of heterogeneity. The ability of iβ to respond to glucose instead was more related to that of EndoC-βH1. Discussion: We demonstrated that insulin-producing cells generated from iPSCs recapitulate fundamental gene expression profiles and secretory function of native human β cells.

Insulin Glulisine—A Comprehensive Preclinical Evaluation

2006 ◽  
Vol 25 (1) ◽  
pp. 25-33 ◽  
Author(s):  
Ingo Stammberger ◽  
Gerhard Seipke ◽  
Thomas Bartels
Keyword(s):  
Dna Synthesis ◽  
Insulin Receptor ◽  
Cell Line ◽  
Binding Affinity ◽  
Human Insulin ◽  
Receptor Binding ◽  
Insulin Receptors ◽  
Preclinical Evaluation ◽  
Receptor Binding Affinity ◽  
Insulin Glulisine

Receptor binding and signaling and the mitogenic potential of insulin glulisine (glulisine), regular human insulin (RHI), and Asp(B10) were compared in vivo and in vitro. Insulin and insulin-like growth factor 1 (IGF-1) receptor binding was studied with human insulin receptors (293HEK cells) and the human osteosarcoma-derived cell line B10. Insulin receptor–mediated signaling was assessed in rat-1 fibroblasts overexpressing insulin receptors. Activation of insulin receptor substrates 1 and 2 (IRS-1/IRS-2) was studied in rat and human myoblasts and rat cardiomyocytes. DNA synthesis induction was assessed by [3H] thymidine incorporation in the human epithelial breast cell line MCF10. Interaction with the IGF-1 receptor, DNA synthesis, and intracellular signal transduction were assessed in cardiac K6 myoblasts. Immunohistochemical examination of Sprague-Dawley rat tissue treated with glulisine for 6 months ( n = 40), and glulisine and RHI for 12 months ( n = 60), was performed. Steady-state insulin receptor binding affinity was slightly lower for glulisine versus RHI (~0.70). IGF-1 receptor binding affinity was lower (four-to fivefold) for glulisine, but significantly higher (four-fold) for Asp(B10) versus RHI. Glulisine, Asp(B10), and RHI showed similar insulin receptor–association kinetics; however, Asp(B10) revealed increased insulin receptor affinity. Glulisine and RHI showed similar insulin receptor–mediated phosphorylation and IRS-2 activation. Activation of IRS-1 was 6- to 10-fold lower with glulisine; glulisine was less potent and Asp(B10) slightly more potent in stimulating DNA synthesis versus RHI. Stimulation of DNA synthesis was comparable for glulisine and RHI in K6 myoblasts. At 12 months, there was no significant difference between glulisine and RHI in proliferative activity. This preclinical evaluation suggests that structural changes in glulisine versus RHI are not associated with any safety issues.

scholarly journals Dextran Sulfate Protects Pancreatic β-Cells, Reduces Autoimmunity and Ameliorates Type 1 Diabetes

2020 ◽  
Author(s):  
Ada Admin ◽  
Geming Lu ◽  
Francisco Rausell-Palamos ◽  
Jiamin Zhang ◽  
Zihan Zheng ◽  
...  
Keyword(s):  
T Cells ◽  
Type 1 Diabetes ◽  
Heparan Sulfate ◽  
Dextran Sulfate ◽  
Nod Mice ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells

A failure in self-tolerance leads to autoimmune destruction of pancreatic β-cells and type 1 diabetes (T1D). Low molecular weight dextran sulfate (DS) is a sulfated semi-synthetic polysaccharide with demonstrated cytoprotective and immunomodulatory properties <i>in vitro</i>. However, whether DS can protect pancreatic β-cells, reduce autoimmunity and ameliorate T1D is unknown. Here we report that DS, but not dextran, protects human β-cells against cytokine-mediated cytotoxicity <i>in vitro</i>. DS also protects mitochondrial function and glucose-stimulated insulin secretion and reduces chemokine expression in human islets in a pro-inflammatory environment. Interestingly, daily treatment with DS significantly reduces diabetes incidence in pre-diabetic non-obese diabetic (NOD) mice, and most importantly, reverses diabetes in early-onset diabetic NOD mice. DS decreases β-cell death, enhances islet heparan sulfate (HS)/heparan sulfate proteoglycan (HSPG) expression and preserves β-cell mass and plasma insulin in these mice. DS administration also increases the expression of the inhibitory co-stimulatory molecule programmed death-1 (PD-1) in T-cells, reduces interferon-γ+ CD4+ and CD8+ T-cells and enhances the number of FoxP3+ cells. Collectively, these studies demonstrate that the action of one single molecule, DS, on β-cell protection, extracellular matrix preservation and immunomodulation can reverse diabetes in NOD mice highlighting its therapeutic potential for the treatment of T1D.

scholarly journals Primary cilia sensitize insulin receptor-mediated negative feedback in pancreatic β cells

2020 ◽  
Author(s):  
Yuezhe Li ◽  
Prem. K. Shrestha ◽  
Yi I. Wu
Keyword(s):  
Insulin Receptor ◽  
Negative Feedback ◽  
Primary Cilia ◽  
Insulin Receptors ◽  
Cytosolic Calcium ◽  
Ciliated Cells ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Concentration Changes ◽  
Calcium Elevation

AbstractInsulin receptors (IR) can localize to the primary cilia of pancreatic β cells. Because primary cilia are known to sensitize or bias the signaling of cell surface receptors, we investigate how ciliary insulin receptors influence glucose-stimulated insulin secretion (GSIS) in β cells by gauging how cytosolic calcium concentration changes in a mouse insulinoma cell line (MIN6). Purified recombinant insulin suppresses calcium elevation in response to glucose in these cells. Interestingly, ciliated cells show attenuated cytosolic calcium elevation compared to cilium-free cells after glucose stimulation even in the absence of exogenous insulin. We observe that ciliary IR is highly phosphorylated, and the phospho-IR density decreases when cells are either treated with an insulin receptor (IR) inhibitor, BMS536924, or ciliary function is disrupted through either IFT88 or BBS1 knockdown. Consistently, the attenuation of calcium elevation in ciliated cells is abrogated when cells are either treated with IR inhibitor or when primary cilia are impaired. We further demonstrate that ciliary IR signaling hyperpolarizes the plasma membrane but has no apparent impact on glucose-induced ATP production. Thus, our results argue that primary cilia sensitize insulin receptor signaling and mediate negative feedback in GSIS in pancreatic β cells.

scholarly journals Dextran Sulfate Protects Pancreatic β-Cells, Reduces Autoimmunity and Ameliorates Type 1 Diabetes

2020 ◽  
Author(s):  
Ada Admin ◽  
Geming Lu ◽  
Francisco Rausell-Palamos ◽  
Jiamin Zhang ◽  
Zihan Zheng ◽  
...  
Keyword(s):  
T Cells ◽  
Type 1 Diabetes ◽  
Heparan Sulfate ◽  
Dextran Sulfate ◽  
Nod Mice ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells

A failure in self-tolerance leads to autoimmune destruction of pancreatic β-cells and type 1 diabetes (T1D). Low molecular weight dextran sulfate (DS) is a sulfated semi-synthetic polysaccharide with demonstrated cytoprotective and immunomodulatory properties <i>in vitro</i>. However, whether DS can protect pancreatic β-cells, reduce autoimmunity and ameliorate T1D is unknown. Here we report that DS, but not dextran, protects human β-cells against cytokine-mediated cytotoxicity <i>in vitro</i>. DS also protects mitochondrial function and glucose-stimulated insulin secretion and reduces chemokine expression in human islets in a pro-inflammatory environment. Interestingly, daily treatment with DS significantly reduces diabetes incidence in pre-diabetic non-obese diabetic (NOD) mice, and most importantly, reverses diabetes in early-onset diabetic NOD mice. DS decreases β-cell death, enhances islet heparan sulfate (HS)/heparan sulfate proteoglycan (HSPG) expression and preserves β-cell mass and plasma insulin in these mice. DS administration also increases the expression of the inhibitory co-stimulatory molecule programmed death-1 (PD-1) in T-cells, reduces interferon-γ+ CD4+ and CD8+ T-cells and enhances the number of FoxP3+ cells. Collectively, these studies demonstrate that the action of one single molecule, DS, on β-cell protection, extracellular matrix preservation and immunomodulation can reverse diabetes in NOD mice highlighting its therapeutic potential for the treatment of T1D.

scholarly journals Photolysis of cell-permeant caged inositol pyrophosphates controls oscillations of cytosolic calcium in a β-cell line

2019 ◽  
Vol 10 (9) ◽  
pp. 2687-2692 ◽  
Author(s):  
S. Hauke ◽  
A. K. Dutta ◽  
V. B. Eisenbeis ◽  
D. Bezold ◽  
T. Bittner ◽  
...  
Keyword(s):  
Cell Line ◽  
Cytosolic Calcium ◽  
Β Cells ◽  
Β Cell ◽  
Inositol Pyrophosphate ◽  
Inositol Pyrophosphates ◽  
Line P

β-Cells respond directly to the intracellular photochemical release of caged inositol pyrophosphate isomers with modulations of oscillations in cytosolic Ca2+.

scholarly journals Differential Mitogenic Signaling in Insulin Receptor-Deficient Fetal Pancreatic β-Cells

Endocrinology ◽  
2006 ◽  
Vol 147 (4) ◽  
pp. 1959-1968 ◽  
Author(s):  
C. Guillen ◽  
P. Navarro ◽  
M. Robledo ◽  
A. M. Valverde ◽  
M. Benito
Keyword(s):  
Insulin Receptor ◽  
Cell Lines ◽  
Cell Mass ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Phosphatidylinositol 3 Kinase ◽  
Β Cells ◽  
Β Cell ◽  
Phosphatidylinositol 3

Insulin receptor (IR) may play an essential role in the development of β-cell mass in the mouse pancreas. To further define the function of this signaling system in β-cell development, we generated IR-deficient β-cell lines. Fetal pancreata were dissected from mice harboring a floxed allele of the insulin receptor (IRLoxP) and used to isolate islets. These islets were infected with a retrovirus to express simian virus 40 large T antigen, a strategy for establishing β-cell lines (β-IRLoxP). Subsequently, these cells were infected with adenovirus encoding cre recombinase to delete insulin receptor (β-IR−/−). β-Cells expressed insulin and Pdx-1 mRNA in response to glucose. In β-IRLoxP β-cells, p44/p42 MAPK and phosphatidylinositol 3 kinase pathways, mammalian target of rapamycin (mTOR), and p70S6K phosphorylation and β-cell proliferation were stimulated in response to insulin. Wortmannin or PD98059 had no effect on insulin-mediated mTOR/p70S6K signaling and the corresponding mitogenic response. However, the presence of both inhibitors totally impaired these signaling pathways and mitogenesis in response to insulin. Rapamycin completely blocked insulin-activated mTOR/p70S6K signaling and mitogenesis. Interestingly, in β-IR−/− β-cells, glucose failed to stimulate phosphatidylinositol 3 kinase activity but induced p44/p42 MAPKs and mTOR/p70S6K phosphorylation and β-cell mitogenesis. PD98059, but not wortmannin, inhibited glucose-induced mTOR/p70S6K signaling and mitogenesis in those cells. Finally, rapamycin blocked glucose-mediated mitogenesis of β-IR−/− cells. In conclusion, independently of glucose, insulin can mediate mitogenesis in fetal pancreatic β-cell lines. However, in the absence of the insulin receptor, glucose induces β-cell mitogenesis.

scholarly journals Differences in internalization and intracellular processing of interleukin-1 associated with the two forms of interleukin-1 receptor found in B-cells and T-cells

1991 ◽  
Vol 273 (1) ◽  
pp. 79-83 ◽  
Author(s):  
R Horuk
Keyword(s):  
T Cells ◽  
B Cells ◽  
Cell Surface ◽  
Cell Line ◽  
Interleukin 1 ◽  
Cell Types ◽  
Single Chain ◽  
Surface Binding ◽  
Raji Cells ◽  
Kinetics Of

There are at least two classes of interleukin-1 (IL-1) receptor, namely p80, and 80 kDa single-chain protein found in T-cells, fibroblasts and many other cell types, and p68, a 68 kDa protein expressed in B-cells and macrophages. The to classes of IL-1 receptor show distinct differences in their substrate-binding site and molecular properties. In this study we show that the kinetics of IL-1 internalization and the consequences of ligand processing in the two subtypes of IL-1 receptor are also very different. The Raji cell line was used as a source of the p68 form of the IL-1 receptor, whereas the YT cell line was used as a source of p80 receptor. Under conditions of steady-state binding the IL-1 was equally distributed between cell-surface and intracellular sites in YT cells, compared with predominantly cell-surface binding (85%) in Raji cells. The mechanism of IL-1 processing was also different in the two cell types. In Raji cells 60% of internalized IL-1 was released from the cells in an intact form, whereas the remainder was degraded. All of the IL-1 extruded from YT cells was intact. The kinetics of IL-1 release was faster in Raji cells, with a half-time of 4.5 h compared with over 15 h in YT cells. SDS/PAGE analysis of internalized IL-1 in Raji cells revealed that the ligand was sequentially processed to trichloroacetic acid-soluble products. The YT receptor-ligand complex was resistant to dissociation at pH 5, whereas that in Raji cells rapidly dissociated at this pH. Treatment of Raji cells with the lysosomotropic agent chloroquine inhibited the degradation of IL-1 without having any effect on the amount of intact IL-1 in the intracellular compartments. From these data we conclude that the pathways of internalization, intracellular trafficking and overall processing of IL-1 are different for p68 IL-1 receptors compared with p80. This could have direct consequences for IL-1 action and IL-1 receptor regulation in the cell.

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