Abstract
Beta cell death caused by endoplasmic reticulum (ER) stress is a key factor aggravating type 2 diabetes. Exenatide, a glucagon-like peptide (GLP)-1 receptor agonist, prevents beta cell death induced by thapsigargin, a selective inhibitor of ER calcium storage. Here, we report on our proteomic studies designed to elucidate the underlying mechanisms. We conducted comparative proteomic analyses of cellular protein profiles during thapsigargin-induced cell death in the absence and presence of exenatide in INS-1 rat insulinoma cells. Thapsigargin altered cellular proteins involved in metabolic processes and protein folding, whose alterations were variably modified by exenatide treatment. We categorized the proteins with thapsigargin initiated alterations into three groups: those whose alterations were 1) reversed by exenatide, 2) exaggerated by exenatide, and 3) unchanged by exenatide. The most significant effect of thapsigargin on INS-1 cells relevant to their apoptosis was the appearance of newly modified spots of heat shock proteins, thimet oligopeptidase and 14-3-3β, ε, and θ, and the prevention of their appearance by exenatide, suggesting that these proteins play major roles. We also found that various modifications in 14-3-3 isoforms, which precede their appearance and promote INS-1 cell death. This study provides insights into the mechanisms in ER stress-caused INS-1 cell death and its prevention by exenatide.
Highlights
Recent reports suggest that hyperglycemia results in both oxidative and endoplasmic reticulum (ER) stresses [1], suggesting that ER stress and oxidative stress are endogenous aggravating factors for type 2 diabetes
We found that levels higher than 10 nM of exenatide blocked caspase-3/7 activation caused by thapsigargin treatment (Fig. 1C, 1D)
Since the response of insulin secretion induced by exenatide continuously increased up to 10 nM, and our previous study [10] showed that exenatide completely blocked all types of beta cell death at 10 nM, but not at 1 nM, we further used 10 nM exenatide as the optimal concentration at which exenatide can lead to the distinct effects in INS-1 cells
Summary
Recent reports suggest that hyperglycemia results in both oxidative and endoplasmic reticulum (ER) stresses [1], suggesting that ER stress and oxidative stress are endogenous aggravating factors for type 2 diabetes. ER stress is a cellular state involving accumulation of unfolded proteins, perturbation of calcium ions, or disturbances of redox state [2,3]. Inhibitors of ER stress might serve as antidiabetic agents. Thapsigargin, a selective inhibitor of endoplasmic reticulum Ca2+/ATPase causing the depletion of ER Ca2+ store, is widely used in studies of ER stress-caused beta cell death. When cells fail to adapt to ER stress, beta cell apoptosis is initiated by diverse signaling molecules.
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