The role of mechanosensitive ion channels in the release of pancreatic hormones and type 2- diabetes

Abstract

Pancreatic beta cells play a critical role in type 2 diabetes (T2D) development and progression. Therefore, deciphering the full mechanism behind glucose-stimulated insulin secretion is of great interest. Glucose elicits insulin secretion by being metabolized to ATP, which closes the ATP-dependent K+ channel, leading to accumulation of positive charges inside the beta-cell, beta-cell depolarization, activation of voltage-gated Ca2+ channels, leading to Ca2+-dependent exocytosis of insulin-containing secretory granules.
However, it remains unknown which molecular reactions that generate the basal influx of positive charges in the beta-cell, without closure of the ATP-dependent K+ channel would not lead to beta-cell depolarization. One candidate for this reaction is mechanotransduction. This signaling modality remains, so far, unexplored in insulin secretion. However, repeated reports indicate that glucose stimulation causes an increase in beta cell volume because of metabolism of the hexose, which increases membrane tension. These mechanical signals can be transduced into electrical signals by the mechanosensitive channels. In this project, we will test whether mechanosensitive signals are active in glucose-stimulated beta cell depolarization and insulin secretion, as well as their involvement in T2D development.
Gene expression of mechanosensitive channels were investigated in human islets and clonal beta cell line. First gene expression of mechanosensitive channels was validated by RT-qPCR and protein detected by Western blotting. Thereafter, mechanosensitive channels were silenced or inhibited to establish its functional implications. Ratiometric Ca2+ imaging was used to determine the cellular cues activating channels. Finally, insulin secretion was measured at basal and stimulatory glucose concentrations.
We found mechanosensitive channels to be highly expressed in both human islets and INS-1 832/13 cells. Interestingly, silencing or inhibiting mechanosensitive channels effect both depolarization-evoked Ca2+ signaling and glucose-stimulated insulin secretion in INS-1 832/13.
Taken together, we propose that mechanosensitive channels have a fundamental function in glucose-stimulated insulin secretion.

Popular Abstract

Mechanotransduction controls insulin secretion

Glucose is the most easily accessible source of energy in our bodies. Blood glucose concentrations are maintained within a relatively narrow range by a group of hormones from the endocrine pancreatic islets of Langerhans. These microorgans consist of at least five different endocrine cell types. Each cell type produces a special hormone that has an important function. Chief among those is insulin, produced by the beta cells. After a meal, blood glucose rises, and the pancreatic beta cells are activated by a chain of metabolic and electrical reactions that culminate with the release of insulin to the blood stream. Insulin is the body’s sole glucose lowering hormone. This is due to its capacity to promote uptake and storage of glucose, primarily in skeletal muscle, adipose tissue and in the liver. However, the actions of insulin are even more far-reaching. It is considered the major anabolic hormone as it promotes storage of food macronutrients. Fatty acids are taken up into the adipose tissue and amino acids are directed to protein synthesis. Patients with type 2 diabetes cannot produce enough insulin in relation to the amounts needed in the cells that normally store glucose. As a result, the patients will have a chronically elevated blood glucose, which if long-term, causes damage in blood vessels. This is one of the main reasons for the well-known diabetic complications in large arteries, such as increased risk of heart attack, stroke, but also in small capillaries leading to blindness, kidney failure and malfunctions in the nervous system.
How does glucose evoke insulin secretion in the beta cell? The consensus model for glucose-stimulated insulin secretion is close to an axiom in diabetes research, and overall it has served 25 years of beta cells research very well, generating numerous important studies and insights. Briefly, glucose is taken up and metabolized to the energy-dense biomolecule ATP, which in turn closes ATP-sensitive ion channels. As long as these ion channels remain open, they continuously let excess amounts of positively charged ions out and this keeps the beta cell negatively charged and in resting state. However, when the ATP-sensitive channels close, the beta cell becomes more positively charged, which activates a series of reactions that culminates with insulin secretion. However, this consensus model for glucose-stimulated insulin secretion is incomplete. The constant influx of positive charges, that the ATP-sensitive ion channel must rectify, has remained unidentified for ~25 years.
Mechanotransduction (the conversion of mechanical stimuli into biological signals) is a signaling modality that so far has not been considered relevant to insulin secretion. However, mechanical signals typically produce electrical currents that make the beta cell more positively charged. Such types of current correspond to the part lacking in the present beta cell consensus model.

In this project, we have tested whether mechanosensitive channels are active in glucose-stimulated insulin secretion and may play a role in type 2 diabetes development. First, we showed that mechanosensitive channels really exist in human pancreatic islets and other beta cells used for experiments. After that, we measured a fundamentally important signal in the beta cell, the concentration of free calcium ions or (Ca2+)free . This signal is generated by opening of voltage-gated Ca2+ channels in glucose-stimulated beta cells as the cells become more positively charged after closure of the ATP-sensitive channels. The biological importance of the Ca2+ ion species is its capacity to activate the insulin release machinery. We measured (Ca2+)free by ratiometric fura-2 AM imaging and found that inhibition or silencing of mechanosensitive channels effected Ca2+ signals inside the cells. Next, we found that silencing or inhibition of mechanosensitive channels leads to change in glucose-stimulated insulin secretion.
Taken together, we propose that mechanosensitive channels have a fundamental function in glucose-stimulated insulin secretion.

Master’s Degree Project in Molecular Biology and Biotechnology/60 credits 2017
Department of Biology, Lund University.
Advisor: Prof. Erik Renström / The Unit of Islet Pathophysiology/ Department of Clinical sciences, Malmö.

• author: Barghouth, Mohammad
• supervisor: Erik Renström
• organization: Degree Projects in Molecular Biology
• course: MOBT01 20162
• year: 2017
• type: H2 - Master's Degree (Two Years)
• subject: Biology and Life Sciences
• language: English
• id: 8909645
• date added to LUP: 2017-05-30 15:54:17
• date last changed: 2017-05-30 15:54:17