Issue 12, 2019

Insulin sensor cells for the analysis of insulin secretion responses in single living pancreatic β cells

Abstract

Investigation of the functions of insulin-secreting cells in response to glucose in single-living cells is essential for improving our knowledge on the pathogenesis of diabetes. Therefore, it is desired to develop a new convenient method that enables the direct detection of insulin secreted from single-living cells. Here, insulin-sensor-cells expressing a protein-based insulin-detecting probe immobilized on the extracellular membrane were developed to evaluate the insulin-secretion response in single-living pancreatic β cells. The protein-based insulin-detecting probe (NαLY) was composed of a bioluminescent protein (nano-luc), the αCT segment of the insulin receptor, L1 and CR domains of the insulin receptor, and a fluorescent protein (YPet). NαLY exhibited a bioluminescence resonance energy transfer (BRET) signal in response to insulin; thus, cells of Hepa1–6 line were genetically engineered to express NαLY on the extracellular membrane. The cells were found to act as insulin-sensor-cells, exhibiting a BRET signal in response to insulin. When the insulin-sensor-cells and pancreatic β cells (MIN6 cell line) were cocultured and stimulated with glucose, insulin-sensor-cells nearby pancreatic β cells showed the spike-shaped BRET signal response, whereas the insulin-sensor-cells close to one pancreatic β cell did not exhibit such signal response. However, all the insulin-sensor-cells showed a gradual increase in BRET signals, which were presumably attributed to the increase in insulin concentrations in the culture dish, confirming the function of these insulin-sensor-cells. Therefore, we demonstrated that heterogenetic insulin secretion in single-living pancreatic β cells could be measured directly using the insulin sensor cells.

Graphical abstract: Insulin sensor cells for the analysis of insulin secretion responses in single living pancreatic β cells

Supplementary files

Article information

Article type
Paper
Submitted
02 Mac 2019
Accepted
08 Mei 2019
First published
09 Mei 2019

Analyst, 2019,144, 3765-3772

Insulin sensor cells for the analysis of insulin secretion responses in single living pancreatic β cells

H. Shigeto, T. Ono, T. Ikeda, R. Hirota, T. Ishida, A. Kuroda and H. Funabashi, Analyst, 2019, 144, 3765 DOI: 10.1039/C9AN00405J

To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements