3T3-L1 adipocytes as a model of Glut4 translocation

Abstract

Type 2 diabetes (T2D), a lethal disease, reveals an alarming increase of epidemic proportions and, by 2050, an estimated 1 in 3 adults in the US will suffer from it. Persistent and systemic inflammatory and oxidative stress in adipose tissue and skeletal muscle are recognized as key players in the progression of T2D. T2D is characterized by insulin resistance, the inability of cells to respond to standard circulating levels of insulin, resulting in hyperglycemia. In adipocytes and skeletal muscle, glucose uptake, a vital step in blood glucose regulation, predominantly occurs via the glucose transporter 4 (Glut4) in an insulindependent recruitment from a cytosolic vesicle. Under inflammatory stress, these cells have demonstrated a lack of Glut4 presence in the plasma membrane, leading to a lack of glucose transport within these cells. 3T3-L1 murine adipocytes are a key model to study insulin signaling in vitro. Mature adipocytes are converted from fibroblasts in a lengthy process requiring chemical induction. Several inducers have been tested in hopes to shorten the time span required for differentiation. However, the use of different inducers and changes in the differentiation programming could have consequences on the phenotype of the mature adipocytes. For this reason, we sought to evaluate the differences in 3T3-L1 fibroblasts induced with either 3-isobutyl-1-methylxanthine (IBMX), or troglitazone. In addition to pharmaceutical recourse to restore insulin signaling and glucose uptake in cells under inflammatory duress, a dietary approach has been considered. Natural products such as blueberries have been of particular interest due to their health benefit including antioxidant benefits and their ability to modulate biochemical pathways. Wild Alaskan blueberries are of particular interest due to their link to a reduced prevalence of T2D in Alaskan Native populations. Here, we report differences in the phenotype of 3T3-L1 adipocytes based on the inducer of adipogenesis and a lack of increase in plasma membrane Glut4. Our studies demonstrate a higher expression of adipocyte biomarkers in cells treated with troglitazone compared to IBMX, but a higher expression of total Glut4 in cells treated with IBMX compared to troglitazone. In both phenotypes, we note a lack of insulin-induced increase of plasma membrane levels of Glut4 in 3T3-L1 mature adipocytes. Finally, we demonstrate an insulin-like benefit of Alaskan blueberry extracts on plasma membrane Glut4.