ORIGINAL ARTICLE Amino Acids Potentiate Insulin Signaling in CHO-K1 at High Glucose Conditions Radhakrishnan Selvi, Renganathan Bhuvanasundar, Aluru Venkata Saijyothi, Konerirajapuram Natarajan Sulochana, and Narayanasamy Angayarkanni Department of Biochemistry and Cell Biology, Vision Research Foundation, Chennai, India Received for publication December 26, 2011; accepted March 26, 2012 (ARCMED-D-11-00663). Background and Aims. Amino acids reportedly increase the glucose uptake under high glucose conditions. However, there are controversies in the role of amino acids in dia- betes mellitus. The present study explores the insulin signaling pathway involved in glucose uptake mediated by amino acids in CHO-K1 cells. Methods. CHO-K1 cells were exposed to normal (7 mM) and high glucose (17 and 27 mM) with 100 nM insulin in the presence and absence of amino acid mixtures (AAM) in varying concentration (5 and 20 mM) followed by the assays, insulin receptor tyrosine kinase (IRTK) and phosphatidylinositol 3 kinase (PI3K) by autoradiography, protein kinase B (Akt) and glucose transporter (GLUT4) by Western blot and glycogen synthase (GS) by HPLC. Results. The addition of 5 and 20 mM AAM significantly increased IRTK and PI3K activity (ANOVA p 5 0.025, p 5 0.003, respectively) with increasing glucose concentra- tion. Addition of 5 mM AAM in the presence of normal glucose significantly increased the levels of phosphorylated Akt Ser473 ( p 5 0.02) with no significant change at high glucose. At 20 mM AAM there was a significant decrease in Akt phosphorylation ( p 5 0.035) that was increased by high glucose concentration. GLUT4 protein levels were increased with AAM (5 mM) along with increase in glycogen synthase activity at all glucose concentrations ( p !0.05). Conclusions. Amino acids as a mixture is beneficial in augmenting insulin signaling pathway via IRTK/PI3K/GLUT4 pathway along with activation of GS in CHO-K1 cells, thereby ensuring increased intracellular glucose availability. Ó 2012 IMSS. Published by Elsevier Inc. Key Words: Amino acids, Glucose, IRTK, PI3K, Akt, GLUT4. Introduction Amino acid levels in plasma have been shown to have disease correlations in diabetes mellitus (1). In the context of hyperglycemia, amino acids such as lysine, glycine, alanine, glutamic acid and aspartic acid have been reported to have antiglycating properties (2) and anticataract proper- ties (3). Oral supplementation with amino acids for patients with type 2 diabetes mellitus (T2DM) under anti-diabetic therapy produced a further decrease of postprandial plasma glucose without change in plasma insulin levels. This decrease was attributed to the amino acid supplementation that improved insulin sensitivity (4). AA as a mixture (AAM) was also found to increase glucose transport in CHO-K1 cells (5). The mechanism through which AAM augments GLUT4 recruitment is not yet clear and is ad- dressed in this study. Glucose uptake initiated by insulin is receptor mediated. Insulin binds to the insulin receptor (IR) on the plasma membrane, increasing its tyrosine kinase activity that phos- phorylates insulin receptor substrates (IRS). Tyrosine phos- phorylated IRS-1 recruits and activates PI3K, which increases serine phosphorylation of the downstream Akt. Activation of Akt promotes translocation of GLUT4 to Address reprint requests to: Narayanasamy Angayarkanni, Head, Depart- ment of Biochemistry and Cell Biology, Sankara Nethralaya, Vision Research Foundation, 18, College Road, Chennai 600 006, India; Phone: 91-44- 28271616; FAX: 91-44-28254180; E-mail: drak@snmail.org 0188-4409/$ - see front matter. Copyright Ó 2012 IMSS. Published by Elsevier Inc. doi: 10.1016/j.arcmed.2012.04.008 Archives of Medical Research 43 (2012) 173e182