The PI3K/Akt1-FoxO1 Translocation Pathway Mediates EXf Effects on NIT-1 Cell Survival

Abstract

EXf, a glucagon-like peptide 1 (GLP-1) receptor agonist, stimulates β-cell proliferation and reduces apoptosis in diabetic animal models, but the underlying mechanisms are not fully understood. We constructed a FoxO1-GFP fusion protein expression plasmid and transiently transfected it into NIT-1 cells to investigate whether FoxO1 mediates EXf effects on NIT-1 cell survival. Our results showed that EXf could increase cell viability by inhibiting apoptosis and stimulating proliferation, and it could also promote the translocation of the FoxO1-GFP fusion protein from the nucleus to the cytoplasm in NIT-1 cells. However, the above effects of EXf were suppressed by the inhibitor of PI3K. Comparative transcription analysis showed up-regulation of igf-1r, irs-2, pI3k, akt1 and pdx-1 in NIT-1 cells after EXf treatment. Moreover, the up-regulation of PI3K and phosphorylation of Akt1 upon EXf treatment was confirmed by Western blot, both phenomena were abrogated by wortmannin, an inhibitor of PI3K. In summary, FoxO1 may mediate the effects of EXf on NIT-1 cell survival by activating the PI3K/Akt1 pathway.

Key words

EXf - GLP-1 - FoxO1 - NIT-1 cells - translocation

• References

• 1 Druck DJ, Nauck MA. The incretin system: Glucagon-like-peptide-1receptor agonists and dipeptidyl peptidase-4 inhibitors in type 2 diabetes. Lancet 2006; 368: 1690-1705
• 2 Drucker DJ. Minireview: The glucagon-like peptides. Endocrinology 2001; 142: 521-527
• 3 Gutniak M, Orskov C, Holst JJ. et al. Antidiabetogenic effect of glucagon-like peptide-1 (7–36) amide in normal subjects and patients with diabetes mellitus. N Engl J Med 1992; 326: 1316-1322
• 4 Holst JJ. Therapy of type 2 diabetes mellitus based on the actions of glucagon-like peptide-1. Diabetes Metab Res Rev 2002; 18: 430-441
• 5 Fehmann HC, Habener JF. Insulinotropic hormone glucagon-like peptide-1 (7–37) stimulation of proinsulin gene expression and proinsulin biosynthesis in insulinoma beta TC-1 cells. Endocrinology 1992; 130: 159-166
• 6 Buteau J, Roduit R, Susini S. et al. Glucagon-like peptide-1 promotes DNA synthesis, activates phosphatidylinositol 3-kinase and increases transcription factor pancreatic and duodenal homeobox gene 1 (PDX-1) DNA binding activity in beta (INS-1)-cells. Diabetologia 1999; 42: 856-864
• 7 Stoffers DA, Kieffer TJ, Hussain MA. et al. Insulinotropic glucagon-like peptide 1 agonists stimulate expression of homeodomain protein IDX-1 and increase islet size in mouse pancreas. Diabetes 2004; 9: 741-748
• 8 Farilla L, Hui H, Bertolotto C. et al. Glucagon-like peptide-1 promotes islet cell growth and inhibits apoptosis in Zucker diabetic rats. Endocrinology 2002; 143: 4397-4408
• 9 Buteau J, Foisy S, Rhodes CJ. et al. Protein kinase Cζ activation mediates glucagon-like peptide-1-induced pancreatic beta-cell proliferation. Diabetes 2001; 50: 2237-2243
• 10 Buteau J, Foisy S, Joly E. et al. Glucagon-like peptide 1 induces pancreatic beta-cell proliferation via transactivation of the epidermal growth factor receptor. Diabetes 2003; 52: 124-132
• 11 Buteau J, EI-Assaad W, Rhodes CJ. et al. Glucagon-like peptide-1 prevents beta cell glucolipotoxicity. Diabetologia 2004; 47: 806-815
• 12 Wang Q, Li L, Xu E. et al. Glucagon-like peptide-1 regulates proliferation and apoptosis via activation of protein kinase B in pancreatic INS-1 beta cells. Diabetologia 2004; 47: 478-487
• 13 Kops GJ, Burgering BM. Forkhead transcription factors: New insights into protein kinase B (c-akt) signaling. J Mol Med 1999; 77: 656-665
• 14 Buteau J, Shlien A, Foisy S. et al. Metabolic diapause in pancreatic beta- cells expressing a gain-of-function mutant of the forkhead protein Foxo1. J Biol Chem 2007; 282: 287-293
• 15 Kim SJ, Winter K, Nian C. et al. Glucose- dependent insulinotropic polypeptide (GIP) stimulation of pancreatic beta-cell survival is dependent upon phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB) signaling, inactivation of the forkhead transcription factor Foxo1, and down-regulation of bax expression. J Biol Chem 2005; 280: 22297-22307
• 16 Nakae J, Biggs 3rd WH, Kitamura T. et al. Regulation of insulin action and pancreatic beta-cell function by mutated alleles of the gene encoding forkhead transcription factor Foxo1. Nat Genet 2002; 32: 245-253
• 17 Song GM, Huan Y, Sun SJ. et al. Biological activity of EXf, a peptide analogue of Ex-4. European Journal of Pharmacology 2010; 628: 261-267
• 18 Hou GJ, Li CN, Liu SN. et al. Long-term treatment with EXf, a peptide analogue of Ex-4, improves β-cell function and survival in diabetic KKAy mice. Peptides 2013; 40: 123-132
• 19 Buteau J, Accili D. Regulation of pancreatic beta-cell function by the forkhead protein FoxO1. Diabetes, Obesity and Metabolism 2007; 9: 140-146
• 20 Harbeck MC, Louie DC, Howland J. et al. Expression of insulin receptor mRNA and insulin receptor substrate 1 in pancreatic islet beta-cells. Diabetes 1996; 45: 711-717
• 21 Cornu M, Yang JY, Jaccard E. et al. Glucagon like peptide-1 protects beta-cells against apoptosis by increasing the activity of an IGF-2/IGF-1 receptor autocrine loop. Diabetes 2009; 58: 1816-1825
• 22 Kitamura T, Nakae J, Kitamura Y. et al. The forkhead transcription factor Foxo1 links insulin signaling to Pdx1 regulation of pancreatic beta cell growth. J Clin Invest 2002; 110: 1839-1847
• 23 Sharma A, Zangen DH, Reitz P. et al. The homeodomain protein IDX-1 increases after an early burst of proliferation during pancreatic regeneration. Diabetes 1999; 48: 507-513
• 24 Buteau J, Spatz ML, Accili D. Transcription Factor FoxO1 Mediates Glucagon-like peptide-1 effects on pancreatic beta-cell mass. Diabetes 2006; 55: 1190-1195