In vitro protective effects of a Glycyrrhiza glabra L. leaf extract on palmitate-induced insulin resistance in endothelial cells

Liquorice (Glycyrrhiza glabra L., family Leguminosae) is one of the most popular medicinal plants known worldwide. The major value of this plant lies in the valuable compounds found in its roots, some of which are associated with positive health effects. However, little attention have been given to liquorice aerial parts, always been considered merely a waste, known to contain inositols and dihydrostilbenes, among the others, able to exert anti-inflammatory and antioxidant effects. Recently, inositols are receiving particular and growing interest owing to their several biological properties such as insulin sensitizing agent. In this study we chemically characterized a methanolic extract of Glycyrrhiza glabra leaf (GGLME) and then evaluated the in vitro effects on insulin resistance in human endothelial cells (HUVECs) exposed to palmitic acid (PA). Liquid chromatography analysis of GGLME reported identification and characterization of over thirty components, including new dihydrostilbenes, and a consistent amount of the inositol D-pinitol. To investigate the protective effect of GGLME and D-pinitol on PA-induced insulin resistance, HUVECs were pretreated for 24 h with GGLME (40-80 μg/mL) or D-pinitol (40 μM), then exposed for 3 h to 100 µM PA, and finally treated with 100 nM insulin for 15 min. D-pinitol or GGLME pretreatment significantly protected HUVECs from PA-induced toxicity at all the tested concentrations. Our results indicate that PA exposure effectively inhibited insulin-mediated IRS-1 tyrosine phosphorylation, thus blocking the PI3K/Akt/eNOS pathway with subsequent insulin resistance. Both GGLME (in a dose-dependent way) and D-pinitol pretreatments were associated to an increased tyrosine phosphorylation of IRS-1. Interestingly, this effect was observed also without PA exposure, thus demonstrating an insulin sensitizing activity of the tested compounds. Furthermore, we demonstrated an impaired PI3K/Akt/eNOS insulin signaling following PA exposure. GGLME (in a dose-dependent way) or D-pinitol pretreatments effectively increased PI3K, Akt, and eNOS activation, and these effects were observed also without PA exposure. Finally, our data suggest that the methanolic extract of Glycyrrhiza glabra leaf and its main compound D-pinitol are potentially able to ameliorate insulin resistance-related endothelial dysfunction caused by lipotoxicity. These observations contribute to provide a molecular background to the beneficial effects of Glycyrrhiza glabra leaf, from vegetable waste to new agent in the prevention and treatment of cardiovascular diseases related to insulin resistance, diabetes, and metabolic syndrome.