Pro-Survival Role of Gelsolin in Mouse β-Cells

Yermen, Barbara; Tomás, Alejandra; Halban, Philippe A.

doi:10.2337/db06-0769

Cited by 25 publications

(24 citation statements)

References 40 publications

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“…For the MIN6B1 assays, the rate of cell death was analysed using TUNEL staining (Roche, Mannheim, Germany), which detects nuclei in cells undergoing apoptosis or necrosis by immunofluorescence. It has been shown previously that under these conditions of culture, cell death detected by TUNEL is due predominantly to apoptosis [17]. The percentage of apoptotic cells was quantified using ImageJ-based software (NIH, Bethesda, MD, USA), where a brightness threshold detection system quantifies apoptotic cells (TUNEL-positive cells) in relation to the total number of cells (stained with 4′,6-diamidino-2-phenylindole [DAPI]; Vector Labs, Peterborough, UK).…”

Section: Methodsmentioning

confidence: 99%

Cardiotrophin 1 protects beta cells from apoptosis and prevents streptozotocin-induced diabetes in a mouse model

et al. 2013

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Aims/hypothesis Cardiotrophin 1 (CT-1) is a recently described cytokine originally isolated from the heart where it has been shown to play an important role in apoptotic protection of cardiomyocytes and heart hypertrophy. Its beneficial properties have also been described in other organs such as liver and neuromuscular tissue. In the present study, we investigated whether CT-1 can confer protection against pro-apoptotic stimuli in pancreatic beta cells, and its role in insulin secretion and diabetes development.Methods The effects of CT-1 on apoptosis and function were studied using MIN6B1 cells and freshly isolated murine pancreatic islets. The impact on the development of diabetes was evaluated in Ct1-null (Ct1 −/− ) mice (the gene Ct1 is also known as Ctf1) using two streptozotocin (STZ)-induced models of diabetes. Results CT-1 has a protective effect in MIN6B1 cells and murine islets under the pro-apoptotic stimulus of serum deprivation, which correlates with the expression of B cell lymphoma-extra large, or following exposure to a mixture of cytokines. In addition, CT-1 enhances glucose-stimulated insulin secretion in MIN6B1 cells and this was repressed by inhibitors of phospholipase C. Furthermore, Ct1 −/− mice were more prone to develop diabetes, and their glucose tolerance test showed impaired plasma glucose clearance which correlated with decreased pancreatic insulin secretion. Conclusions/interpretation The results obtained from both in vitro and in vivo experiments show that CT-1 improves beta cell function and survival, and protects mice against STZ-induced diabetes.

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Section: Methodsmentioning

confidence: 99%

Cardiotrophin 1 protects beta cells from apoptosis and prevents streptozotocin-induced diabetes in a mouse model

et al. 2013

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“…Detection of apoptosis and proliferation. For MIN6B1 cells, apoptosis was measured by transferase-mediated dUTP nick-end labeling (TUNEL) assay or caspase-3 activation, and proliferation was assessed by 5-bromo-2Ј-deoxyuridine (BrdU) incorporation as previously described (19). For primary mouse islets cells, the cells were established in monolayer on dishes coated with 804G-ECM according to established procedures (18).…”

Section: Methodsmentioning

confidence: 99%

“…Finally, to assess cell death, MIN6B1 cells lacking IRS-2 and/or AS160 were cultured for 72 h in either standard (25 mmol/l glucose and 15% FCS) or deprived (5 mmol/l glucose and 1% FCS) condition, the latter having been shown previously to increase apoptosis in these cells (19). Cell death was quantified using either TUNEL assay (28), which detects DNA strand breaks without distinguishing between apoptosis and necrosis, or active caspase-3, a main mediator of apoptosis.…”

Section: As160 In ␤-Cellsmentioning

confidence: 99%

Rab GTPase-Activating Protein AS160 Is a Major Downstream Effector of Protein Kinase B/Akt Signaling in Pancreatic β-Cells

Bouzakri¹,

Ribaux²,

Tomás³

et al. 2008

Diabetes

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OBJECTIVE-Protein kinase B/Akt plays a central role in ␤-cells, but little is known regarding downstream Akt substrates in these cells. Recently, Rab GTPase-activating protein AS160, a substrate of Akt, was shown to be involved in insulin modulation of GLUT4 trafficking in skeletal muscle and adipose tissue. The aim of this study was to investigate the expression and potential role of AS160 in ␤-cells.RESEARCH DESIGN AND METHODS-AS160 mRNA expression was measured in mouse and human islets and fluorescenceactivated cell sorted ␤-cells and compared in islets from control subjects versus individuals with type 2 diabetes. For knockdown experiments, transformed mouse insulin-secreting MIN6B1 cells were transfected with pSUPER-GFP plasmid encoding a small hairpin RNA against insulin receptor substrate (IRS)-2, AS160, or a negative control. Primary mouse islet cells were transfected with AS160 small interfering RNA. RESULTS-AS160was expressed in human and mouse pancreatic ␤-cells and phosphorylated after glucose stimulation. AS160 mRNA expression was downregulated in pancreatic islets from individuals with type 2 diabetes. In MIN6B1 cells, glucose induced phosphorylation of Akt and AS160, and this was mediated by insulin receptor/IRS-2/phosphatidylinositol 3-kinase independently of changes in cytosolic Ca 2ϩ . Knockdown of AS160 resulted in increased basal insulin secretion, whereas glucosestimulated insulin release was abolished. Furthermore, ␤-cells with decreased AS160 showed increased apoptosis and loss of glucose-induced proliferation.CONCLUSIONS-This study shows for the first time that AS160, previously recognized as a key player in insulin signaling in skeletal muscle and adipose tissue, is also a major effector of protein kinase B/Akt signaling in the ␤-cell.

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“…Cells were then incubated for 1 h at 37°C with KRBH containing 2.8 mM glucose (basal), followed by 1 h at 16.7 mM glucose (stimulated). The buffers were recovered and cellular insulin extracted with acid/ethanol (27). Insulin in buffers and extracts was measured by radioimmunoassay using rat or human insulin standard.…”

Section: Antibodies and Reagents-l-nmentioning

confidence: 99%

“…Detection of Apoptosis and Proliferation-Cell death was measured by TUNEL assay and proliferation was assessed by BrdU incorporation as previously described (27).…”

Section: Antibodies and Reagents-l-nmentioning

confidence: 99%

Silencing Mitogen-activated Protein 4 Kinase 4 (MAP4K4) Protects Beta Cells from Tumor Necrosis Factor-α-induced Decrease of IRS-2 and Inhibition of Glucose-stimulated Insulin Secretion

Bouzakri

Ribaux

Halban

2009

Journal of Biological Chemistry

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Obesity and type 2 diabetes present partially overlapping phenotypes with systemic inflammation as a common feature, raising the hypothesis that elevated cytokine levels may contribute to peripheral insulin resistance as well as the decreased beta cell functional mass observed in type 2 diabetes. In healthy humans, TNF-␣ infusion induces skeletal muscle insulin resistance. We now explore the impact of TNF-␣ on primary beta cell function and the underlying signaling pathways. Human and rat primary beta cells were sorted by FACS and cultured for 24 h ؎ 20 ng/ml TNF-␣ to explore the impact on apoptosis, proliferation, and short-term insulin secretion (1 h, 2.8 mM glucose followed by 1 h, 16.7 mM glucose at the end of the 24-h culture period) as well as key signaling protein phosphorylation and expression. Prior exposure to TNF-␣ for 24 h inhibits glucose-stimulated insulin secretion from primary beta cells. This is associated with a decrease in glucose-stimulated phosphorylation of key proteins in the insulin signaling pathway including Akt, AS160, and other Akt substrates, ERK as well as the insulin receptor. Strikingly, TNF-␣ treatment decreased IRS-2 protein level by 46 ؎ 7% versus control, although mRNA expression was unchanged. While TNF-␣ treatment increased MAP4K4 mRNA expression by 33 ؎ 5%, knockdown of MAP4K4 by siRNA-protected beta cells against the detrimental effects of TNF-␣ on both insulin secretion and signaling. We thus identify MAP4K4 as a key upstream mediator of TNF-␣ action on the beta cell, making it a potential therapeutic target for preservation of beta cell function in type 2 diabetes.

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Pro-Survival Role of Gelsolin in Mouse β-Cells

Cited by 25 publications

References 40 publications

Cardiotrophin 1 protects beta cells from apoptosis and prevents streptozotocin-induced diabetes in a mouse model

Cardiotrophin 1 protects beta cells from apoptosis and prevents streptozotocin-induced diabetes in a mouse model

Rab GTPase-Activating Protein AS160 Is a Major Downstream Effector of Protein Kinase B/Akt Signaling in Pancreatic β-Cells

Silencing Mitogen-activated Protein 4 Kinase 4 (MAP4K4) Protects Beta Cells from Tumor Necrosis Factor-α-induced Decrease of IRS-2 and Inhibition of Glucose-stimulated Insulin Secretion

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