Fluorescent Exendin-4 Derivatives for Pancreatic β-Cell Analysis

Clardy, Susan; Keliher, Edmund J.; Mohan, James F.; Sebas, Matt; Benoist, Christophe; Mathis, Diane; Weissleder, Ralph

doi:10.1021/bc4005014

Cited by 40 publications

(64 citation statements)

References 18 publications

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“…1B). Furthermore, the binding of a fluorescently tagged exenatide peptide that was developed to probe GLP-1R expression (22) was evident in MEG-01 cells (Fig. 1C).…”

Section: Human Megakaryocytes and Platelets Express A Functional Glp-1rmentioning

confidence: 99%

“…MIN6 cells and MEG-01 cells were washed with Hanks' buffered salt solution and resuspended in binding buffer (as described previously) (22,23) with 100 nmol/L fluorescent probe (EP40-BF) or 100 nmol/L unlabeled exenatide for 1 h in the dark. Cells were washed 23 with Hanks' buffered salt solution and resuspended in PBS with 2% BSA.…”

Section: Fluorescent Exenatide Probementioning

confidence: 99%

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Glucagon-Like Peptide 1 Receptor Activation Attenuates Platelet Aggregation and Thrombosis

et al. 2016

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Short-term studies in subjects with diabetes receiving glucagon-like peptide 1 (GLP-1)-targeted therapies have suggested a reduced number of cardiovascular events. The mechanisms underlying this unexpectedly rapid effect are not known. We cloned full-length GLP-1 receptor (GLP-1R) mRNA from a human megakaryocyte cell line (MEG-01), and found expression levels of GLP-1Rs in MEG-01 cells to be higher than those in the human lung but lower than in the human pancreas. Incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in MEG-01 cells, and exenatide significantly inhibited thrombin-, ADP-, and collagen-induced platelet aggregation. Incubation with exenatide also inhibited thrombus formation under flow conditions in ex vivo perfusion chambers using human and mouse whole blood. In a mouse cremaster artery laser injury model, a single intravenous injection of exenatide inhibited thrombus formation in normoglycemic and hyperglycemic mice in vivo. Thrombus formation was greater in mice transplanted with bone marrow lacking a functional GLP-1R (Glp1r 2/2 ), compared with those receiving wild-type bone marrow. Although antithrombotic effects of exenatide were partly lost in mice transplanted with bone marrow from Glp1r 2/2 mice, they were undetectable in mice with a genetic deficiency of endothelial nitric oxide synthase. The inhibition of platelet function and the prevention of thrombus formation by GLP-1R agonists represent potential mechanisms for reduced atherothrombotic events.Type 2 diabetes (T2D) is associated with a number of risk factors that contribute to an increased risk of atherothrombotic events, including hypertension, dyslipidemia, obesity, and chronic inflammation, as well as endothelial and platelet dysfunction (1). Platelets are small, versatile, anucleate cells in the circulation that play critical roles in both early and late stages of atherothrombosis, contributing also to cell-based thrombin generation and blood coagulation (2). Subjects with T2D exhibit a prothrombotic state, including increased production of coagulation factors; decreased production of fibrinolytic factors; and a propensity to platelet activation, aggregation, and adhesion (1,3,4). Compounding the latter, subjects with T2D show reduced sensitivity to antiplatelet drugs, such as aspirin and clopidogrel (5,6), and manifest a higher incidence of cardiovascular events (1,6,7). Although the currently available antidiabetic agents have been effective at lowering blood glucose levels and preventing microvascular disease, until the recent EMPA-REG study (8), it had been exceedingly difficult to demonstrate the beneficial effects of normalizing blood glucose

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“…1B). Furthermore, the binding of a fluorescently tagged exenatide peptide that was developed to probe GLP-1R expression (22) was evident in MEG-01 cells (Fig. 1C).…”

Section: Human Megakaryocytes and Platelets Express A Functional Glp-1rmentioning

confidence: 99%

Section: Fluorescent Exenatide Probementioning

confidence: 99%

Glucagon-Like Peptide 1 Receptor Activation Attenuates Platelet Aggregation and Thrombosis

et al. 2016

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“…Many approaches have been and are being examined, as has recently been reviewed (60). One promising approach is to use fluorescent exendin-4 derivatives, which should bind preferentially to ␤-cells (61,62). However, a major issue concerns how much sensitivity and precision can be achieved by any noninvasive approach.…”

Section: Looking To the Futurementioning

confidence: 99%

β-Cell Failure in Type 2 Diabetes: Postulated Mechanisms and Prospects for Prevention and Treatment

Halban

Polonsky

Bowden

et al. 2014

The Journal of Clinical Endocrinology &Amp; Metabolism

270

216

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β-Cell failure is central to the development and progression of T2D. It antedates and predicts diabetes onset and progression, is in part genetically determined, and often can be identified with accuracy even though current tests are cumbersome and not well standardized. Multiple pathways underlie decreased β-cell function and mass, some of which may be shared and may also be a consequence of processes that initially caused dysfunction. Goals for future research include to 1) impact the natural history of β-cell failure; 2) identify and characterize genetic loci for T2D; 3) target β-cell signaling, metabolic, and genetic pathways to improve function/mass; 4) develop alternative sources of β-cells for cell-based therapy; 5) focus on metabolic environment to provide indirect benefit to β-cells; 6) improve understanding of the physiology of responses to bypass surgery; and 7) identify circulating factors and neuronal circuits underlying the axis of communication between the brain and β-cells.

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“…Receptor activity of the Cy3 and Cy5 probes were tested in a binding assay; the C-terminal fluorophore labeling maintained near-100 % receptor affinity (data not shown), in accordance with previous reports. 19 See suppl. Figure 2 for probe design.…”

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confidence: 99%

Beta cell specific probing with fluorescent exendin-4 is progressively reduced in type 2 diabetic mouse models

Lehtonen

Schäffer

Rasch

et al. 2015

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Probes based on GLP-1R agonist exendin-4 have shown promise as in vivo b cell tracers. However, questions remain regarding the b cell specificity of exendin-4 probes, and it is unclear if the expression levels of the GLP-1R are affected in a type 2 diabetic state. Using in vivo probing followed by ex vivo imaging we found fluorescent exendin-4 probes to distinctly label the pancreatic islets in mice in a Glp-1r dependent manner. Furthermore, a co-localization study revealed a near 100 percent b cell specificity with less than one percent probing in other analyzed cell types. We then tested if probing was affected in models of type 2 diabetes using the Lepr db/db (db/db) and the Diet-Induced Obese (DIO) mouse. Although nearly all b cells continued to be probed, we observed a progressive decline in probing intensity in both models with the most dramatic reduction seen in db/db mice. This was paralleled by a progressive decrease in Glp-1r protein expression levels. These data confirm b cell specificity for exendin-4 based probes in mice. Furthermore, they also suggest that GLP-1R targeting probes may provide a tool to monitor b cell function rather than mass in type 2 diabetic mouse models.

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Fluorescent Exendin-4 Derivatives for Pancreatic β-Cell Analysis

Cited by 40 publications

References 18 publications

Glucagon-Like Peptide 1 Receptor Activation Attenuates Platelet Aggregation and Thrombosis

Glucagon-Like Peptide 1 Receptor Activation Attenuates Platelet Aggregation and Thrombosis

β-Cell Failure in Type 2 Diabetes: Postulated Mechanisms and Prospects for Prevention and Treatment

Beta cell specific probing with fluorescent exendin-4 is progressively reduced in type 2 diabetic mouse models

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