Trigonella foenum-graecum Seed Extract, 4-Hydroxyisoleucine, and Metformin Stimulate Proximal Insulin Signaling and Increase Expression of Glycogenic Enzymes and GLUT2 in HepG2 Cells

Abstract

Fenugreek (Trigonella foenum-graecum) is globally recognized for its medicinal properties and hypoglycemic effects. The seed extract as well as its active compound, 4-hydroxyisoleucine (4-OH-Ile), have been shown to reduce hyperglycemic insulin resistance. The mechanism by which this occurs has not been investigated in human liver cells (HepG2) in comparison to the antihyperglycemic drug, metformin. We investigated the effects of an aqueous fenugreek seed extract (FSE), 4-OH-Ile, and metformin in HepG2 cells relative to insulin as a positive control. Cells were treated with FSE and 4-OH-Ile at 100 ng/mL under normoglycemic (5 mM glucose) and hyperglycemic (30 mM glucose) conditions for 72 hr. Tyrosine phosphorylation of insulin receptor-β (IR-β), protein kinase B (Akt), glycogen synthase kinase-3α/β (GSK-3α/β), and glucose transporter 2 (GLUT2) was determined by western blotting. Gene expression of sterol regulatory element-binding protein 1c (SREBP1c), GLUT2, glycogen synthase (GS), and glucokinase (GK) was evaluated by quantitative polymerase chain reaction, and supernatant glucose levels were measured using the Piccolo biochemistry analyzer. Under normo- and hyperglycemic conditions, FSE, 4-OH-Ile, insulin (100 ng/mL), and metformin (2 mM) caused a significant increase in phosphorylation of IR-β, Akt, GSK-3α/β, and GLUT2. Glucose uptake, however, was most significantly increased in FSE-treated cells during both conditions. FSE induced the most significant changes in downstream insulin signaling, GS, GK, SREBP1c, and GLUT2 expression compared to 4-OH-Ile, metformin, and insulin. In addition, FSE significantly increased glucose uptake. Collectively, these findings provide a mechanism by which FSE exerts antihyperglycemic effects similar to metformin and insulin that occurs via enhanced insulin signaling, gene expression, and increasing glucose uptake.