Obestatin promotes survival of pancreatic beta-cells and human islets and induces expression of genes involved in the regulation of beta-cell mass and function.
OBJECTIVE
Obestatin is a newly discovered peptide encoded by the ghrelin gene whose biological functions are poorly understood. We investigated obestatin effect on survival of beta-cells and human pancreatic islets and the underlying signaling pathways.


METHODS
beta-Cells and human islets were used to assess obestatin effect on cell proliferation, survival, apoptosis, intracellular signaling, and gene expression.


RESULTS
Obestatin showed specific binding on HIT-T15 and INS-1E beta-cells, bound to glucagon-like peptide-1 receptor (GLP-1R), and recognized ghrelin binding sites. Obestatin exerted proliferative, survival, and antiapoptotic effects under serum-deprived conditions and interferon-gamma/tumor necrosis factor-alpha/interleukin-1 beta treatment, particularly at pharmacological concentrations. Ghrelin receptor antagonist [D-Lys(3)]-growth hormone releasing peptide-6 and anti-ghrelin antibody prevented obestatin-induced survival in beta-cells and human islets. beta-Cells and islet cells released obestatin, and addition of anti-obestatin antibody reduced their viability. Obestatin increased beta-cell cAMP and activated extracellular signal-related kinase 1/2 (ERK1/2) and phosphatidylinositol 3-kinase (PI 3-kinase)/Akt; its antiapoptotic effect was blocked by inhibition of adenylyl cyclase/cAMP/protein kinase A (PKA), PI 3-kinase/Akt, and ERK1/2 signaling. Moreover, obestatin upregulated GLP-1R mRNA and insulin receptor substrate-2 (IRS-2) expression and phosphorylation. The GLP-1R antagonist exendin-(9-39) reduced obestatin effect on beta-cell survival. In human islets, obestatin, whose immunoreactivity colocalized with that of ghrelin, promoted cell survival and blocked cytokine-induced apoptosis through cAMP increase and involvement of adenylyl cyclase/cAMP/PKA signaling. Moreover, obestatin 1) induced PI 3-kinase/Akt, ERK1/2, and also cAMP response element-binding protein phosphorylation; 2) stimulated insulin secretion and gene expression; and 3) upregulated GLP-1R, IRS-2, pancreatic and duodenal homeobox-1, and glucokinase mRNA.


CONCLUSIONS
These results indicate that obestatin promotes beta-cell and human islet cell survival and stimulates the expression of main regulatory beta-cell genes, identifying a new role for this peptide within the endocrine pancreas.
Granata
Riccarda
R
Laboratory of Molecular and Cellular Endocrinology, Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Turin, Corso Dogliotti, 14-10126 Turin, Italy. riccarda.granata@unito.it
Settanni
Fabio
F
Gallo
Davide
D
Trovato
Letizia
L
Biancone
Luigi
L
Cantaluppi
Vincenzo
V
Nano
Rita
R
Annunziata
Marta
M
Campiglia
Pietro
P
Arnoletti
Elisa
E
Ghè
Corrado
C
Volante
Marco
M
Papotti
Mauro
M
Muccioli
Giampiero
G
Ghigo
Ezio
E
D053148:Caspase 3 / Q000378:metabolism; D018929:Cell Culture Techniques; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects*; D000242:Cyclic AMP / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D054439:Ghrelin / Q000378:metabolism; D052216:Glucagon-Like Peptide 1 / Q000378:metabolism; D006801:Humans; D055504:Insulin Receptor Substrate Proteins; D050417:Insulin-Secreting Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins / Q000187:drug effects / Q000378:metabolism; D007515:Islets of Langerhans / Q000166:cytology* / Q000187:drug effects; D036361:Peptide Hormones / Q000378:metabolism / Q000494:pharmacology*; D010750:Phosphoproteins / Q000187:drug effects / Q000378:metabolism
D054439:Ghrelin; C526297:IRS2 protein, human; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D036361:Peptide Hormones; D010750:Phosphoproteins; C505799:obestatin, human; D052216:Glucagon-Like Peptide 1; D000242:Cyclic AMP; D053148:Caspase 3